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  • 1
    Electronic Resource
    Electronic Resource
    Springer
    Cell & tissue research 296 (1999), S. 439-442 
    ISSN: 1432-0878
    Keywords: Key words Spleen ; Sinus endothelial cells ; Surface-connected canalicular system ; Lanthanum ; Three-dimensional reconstruction ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  The existence of a surface-connected canalicular system in the splenic sinus endothelial cells of the rat has been demonstrated by transmission electron microscopy with lanthanum nitrate acting as a tracer for the extracellular space. In addition, the three-dimensional arrangement of the canaliculi has been revealed by computer-aided reconstruction. The surface-connected canalicular system of the sinus endothelial cells consists of slender canaliculi that are branched, anastomosed, and that show continuity with the plasma membrane. They twist in and out among the organelles and are often found in close apposition to the spherical invaginations of the plasma membrane and run alongside them. Canaliculi which are not infiltrated by lanthanum nitrate take the form of electron-lucent tubules and are accompanied by numerous spherical invaginations of the plasma membrane. From a computer-aided reconstruction, the canaliculi, which invaginate from various sites of the plasma membrane, have been found to be continuous with each other and to penetrate to the surface of the sinus endothelial cell; they also branch and anastomose to form a complex network in the cytoplasm. Although the surface-connected canalicular system in blood platelets and thrombocytes is believed to function as the main route for the discharge of granules and the uptake of foreign materials and also to take part in the storage and transport of calcium, it is unclear at present whether the network of the surface-connected canalicular system in splenic sinus endothelial cells has any physiological significance.
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  • 2
    ISSN: 1432-0878
    Keywords: Key words Basal lamina ; Immunohistochemistry ; Confocal laser microscopy ; Cochlea ; Mongolian gerbil ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Bridging structures between discrete capillaries in the stria vascularis of the cochlea were studied morphologically in gerbils and rats. Serial thin sections for transmission electron microscopy revealed (1) that elongated cells surrounded by the basal lamina provided the structural basis for the bridging structure, (2) that the basal lamina surrounding the elongated cell extended to the basal lamina around the capillary endothelial cell, (3) that the electron density of the cytoplasm was similar to that of the pericytes around the capillaries, and (4) that the cell was attached to the capillaries at both ends only. Visualization of the basal lamina by immunofluorescent methods revealed (1) that capillaries were often bent at the site of attachment of the bridging cell, (2) that the bridging cell bifurcated occasionally, and (3) that the density of the bridging cell was much higher in the stria vascularis than in the underlying spiral ligament. Filamentous actin visualized by fluorescent phalloidin was not apparent in the bridging cell. We propose that the bridging cell provides mechanical strength to the tortuous capillary network in the stria vascularis and participates in the specific function of the stria vascularis in cooperation with other types of cells.
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  • 3
    ISSN: 1432-0878
    Keywords: Key words Harderian gland ; Tryptophan hydroxylase ; Serotonin ; Immunohistochemistry ; Rat (Wistar) ; Syrian hamster ; Mesocricetus auratus ; Djungarian hamster ; Phodopus sungorus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  The Harderian gland is considered as being an extrapineal source of melatonin. In most rodents, the Harderian gland contains two epithelial cell types (I and II). The aim of this study has been to define which cell type is involved in indoleamine synthesis. The presence and localization of serotonin (melatonin precursor) and tryptophan hydroxylase (the rate-limiting enzyme for serotonin synthesis) have been investigated by immunohistochemistry in male Wistar rats, Syrian hamsters and Djungarian hamsters. The results of the present study show that immunoreactivity for tryptophan hydroxylase and serotonin is confined to the type I cell, suggesting that this cell type is involved in indoleamine synthesis in the rodent Harderian gland.
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  • 4
    ISSN: 1432-0878
    Keywords: Key words Brain-derived neurotrophic factor ; Neurotrophin-3 ; Sympathetic preganglionic neurons ; Chromaffin cells ; Adrenal cortex ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Neurotrophins and their trk receptors constitute major classes of signaling molecules with important actions in the developing and adult nervous system. With regard to the sympathoadrenal cell lineage, which gives rise to sympathetic neurons and chromaffin cells, neurotrophin-3 (NT-3) and nerve growth factor (NGF) are thought to influence developing sympathetic neurons. Neurotrophin requirements of chromaffin cells of the adrenal medulla are less well understood than those for NGF. In order to provide the bases for understanding of putative functions of neurotrophins for the development and maintenance of chromaffin cells and their preganglionic innervation, in situ hybridization has been used to study the expression of brain-derived neurotrophic factor (BDNF) and NT-3, together with their cognate receptors trkB and trkC, in the adrenal gland and in the intermediolateral column (IML) of the spinal cord. BDNF is highly expressed in the embryonic adrenal cortex and later in cells of the cortical reticularis zone. Adrenal medullary chromaffin cells fail to express detectable levels of mRNAs for BDNF, NT-3, and their cognate receptors trkB and trkC. Neurons in the IML express BDNF and trkB, and low levels of NT-3 and trkC. Our data make it unlikely that BDNF and NT-3 serve as retrograde trophic factors for IML neurons but suggest roles of BDNF and NT-3 locally within the spinal cord and possibly for sensory nerves of the adrenal cortex.
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  • 5
    ISSN: 1432-0878
    Keywords: Key words Calcitonin gene-related peptide ; Vasoactive intestinal peptide ; Neuropeptide Y ; Colocalization ; Trigeminal ganglion ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Pituitary adenylate cyclase-activating polypeptide (PACAP)-immunoreactive nerve fibres were demonstrated in the rat pineal gland. These fibres entered the pineal gland through the conarian nerve at the distal tip of the gland. A high density of the fibres was observed in the capsule of the gland, from where the immunoreactive elements penetrated into the pineal perivascular spaces and parenchyma. The majority of PACAP-immunoreactive nerve fibres also contained calcitonin gene-related peptide (CGRP). Some PACAP-immunoreactive nerve fibres contained neuropeptide Y (NPY), but only occasionally was PACAP colocalized with vasoactive intestinal peptide (VIP). After removal of both superior cervical ganglia, a high number of PACAP-containing nerve fibres were still present in the gland. In the nervous system PACAP is present in two isoforms, PACAP-38 and PACAP-27. The concentration of PACAP-38 in the superficial pineal gland was determined by radioimmunoassay to be 20.4 pmol/g tissue at midday and 18.9 pmol/g tissue at midnight. The concentration of PACAP-27 was only about 3% of the concentration of PACAP-38. In summary, this study is the first demonstration of a PACAP-containing innervation of the rat pineal gland. The PACAP concentration in the pineal gland does not exhibit a day-night difference. The colocalization of PACAP with calcitonin gene-related peptide in the pinealopetal nerve fibres indicates that the majority of PACAP-immunoreactive nerve fibres might originate from the trigeminal ganglion.
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  • 6
    Electronic Resource
    Electronic Resource
    Springer
    Cell & tissue research 296 (1999), S. 471-477 
    ISSN: 1432-0878
    Keywords: Key words Skin ; Sensory innervation ; Capsaicin ; Protein gene product 9.5 ; Neurogenic inflammation ; Sensory neuropeptides ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Perineural application of capsaicin results in a selective and permanent reduction in the sensitivity to noxious chemical and heat stimuli and elimination of the neurogenic inflammatory response. The present quantitative immunohistochemical study has been undertaken to reveal the populations of cutaneous afferent nerves that are affected by perineural capsaicin treatment. Areas of intact and chemodenervated skin were determined with the aid of the vascular labelling technique. In sections taken from intact skin areas, staining with antibodies against protein gene product 9.5 revealed a rich epidermal innervation. Fibres immunoreactive for growth-associated protein 43 were also abundant; nerve fibres immunoreactive for substance P and calcitonin gene-related peptide were less numerous. Somatostatin- and RT97-immunoreactive fibres were seen only in the subepidermal layer. In sections taken from skin areas supplied by the sciatic nerve treated with capsaicin 3 days previously, the number of epidermal nerve fibres immunoreactive to protein gene product 9.5, growth-associated protein 43, substance P and calcitonin gene-related peptide was reduced by 90%, 95%, 97% and 66%, respectively. These changes persisted for at least 42 days. The findings reveal that the majority of epidermal axons are capsaicin-sensitive and comprise a chemically heterogeneous population. Reductions in cutaneous fibre populations following perineural capsaicin treatment may result from both the degeneration of sensory axons and the depletion of neuron-specific macromolecules. In addition, most cutaneous nociceptive axons may not use the major sensory neuropeptides substance P and calcitonin gene-related peptide as afferent neurotransmitters.
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  • 7
    ISSN: 1432-0878
    Keywords: Key words Chondrocyte ; Synoviocyte ; Co-culture ; Proliferation ; Lipid peroxidation ; Cytotoxicity ; Ultrastructure ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Objective: A new co-culture system of rat articular chondrocytes and synoviocytes (HIG-82; cell line) was incubated with phorbol myristate acetate (PMA), H2O2 or a combination of Fe2+ and ascorbic acid to simulate inflammation-like radical attacks in articular joints. Methods: Chondrocytes were characterized by immunocytochemistry against collagen type II, transmission electron (TEM) and light microscopy. Lipid peroxidation was investigated by measuring thiobarbituric-acid-reactive material in the supernatants, cytotoxicity by determining release of lactate dehydrogenase and proliferation by measuring [3H]thymidine incorporation, culture protein and DNA. Results: PMA or Fe2+ and ascorbic acid induced lipid peroxidation in chondrocytes and synoviocytes that was decreased significantly in co-cultures. PMA and H2O2 dose dependently induced release of lactate dehydrogenase in chondrocytes, which was lowered in co-cultures or in previously co-cultured chondrocytes to a nearly basal level. In contrast, conditioned media of synoviocyte cultures showed no lowering effect on the radical-induced toxicity. Protection against H2O2-induced damage of cellular membranes by co-culturing was also shown by TEM. Synoviocytes released chondrocyte-stimulating growth factors spontaneously without previous interaction. Conclusion: Chondrocytes establish protective mechanisms against reactive oxygen species via an interaction with synoviocytes. Our co-culture model presents a possible way to study mechanisms of inflammation in articular joints under defined conditions.
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  • 8
    ISSN: 1432-0878
    Keywords: Key words D-aspartate ; Development ; Glutamate ; Retina ; Glutamate transporter (GLAST) ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  The patterns of expression of the glutamate transporter GLAST were compared with the patterns of uptake of exogenous D-aspartate, which is a substrate for all glutamate transporters. At postnatal day 0, fine radial processes and end feet of presumptive Müller cells were weakly immunoreactive for GLAST. At postnatal day 3, intense labelling was associated with astrocytes enveloping newly formed blood vessels on the vitread surface of the retina. Between postnatal days 7 and 10, there was a rapid increase in the intensity of labelling in the Müller cells but clear stratification of GLAST-immunoreactive processes in the inner plexiform layer was not observed until postnatal day 14. By comparison, D-aspartate uptake was initially associated with a wide variety of cellular elements including most neuroblasts, presumptive Müller cells, and astrocytes associated with blood vessels but was absent from the somata of many neurons in the ganglion cell layer and amacrine cell layer. There was a gradual contraction in the numbers of cells that were able to take up D-aspartate, such that, by adulthood, uptake was restricted mainly to Müller cells and astrocytes. We conclude that, during early retinal development, the low levels of GLAST expression by Müller cells permit D-aspartate, and by inference, glutamate, to permeate the retina freely, thus allowing uptake by other glutamate transporters on other cell types. As the retina matures, increased expression of GLAST by Müller cells restricts the access of D-aspartate to other cellular compartments in the retina. This changing pattern of spatial buffering of glutamate by GLAST probably has significant implications regarding our understanding of the role of glutamate during processes such as retinal synaptogenesis.
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  • 9
    ISSN: 1432-0878
    Keywords: Key words Bone sialoprotein ; Osteopontin ; Osteocalcin ; Osteoblasts ; In situ hybridization ; Dexamethasone ; 1 ; 25 dihydroxyvitamin D3 ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  We have used in situ hybridization to evaluate the effects of 1,25 dihydroxyvitamin D3 (1,25 (OH)2 D3) on the expression of mRNA for bone-matrix proteins and to determine whether mature osteoblasts respond differently to 1,25 (OH)2 D3 than younger, newly differentiated osteoblasts. Rat calvaria cells were cultured for 7, 12, 15, and 19 days to obtain a range of nodules from very young to very mature. At each time point, some cultures were treated with 10 nM 1,25 (OH)2 D3 for 24 h prior to fixation. In control cultures, type-I collagen mRNA was detectable in osteoblastic cells in very young nodules and increased with increasing maturity of the nodules and the osteoblasts lining them. The bone sialoprotein mRNA signal was weak in young osteoblasts, increased in older osteoblasts, and decreased in mature osteoblasts. Weak osteocalcin and osteopontin signals were seen only in osteoblasts of intermediate and mature nodules. 1,25 (OH)2 D3 treatment markedly upregulated osteocalcin and osteopontin mRNAs and downregulated mRNA levels of bone sialoprotein and, to a lesser extent, type-I collagen in both young and mature osteoblasts. However, a marked diversity of signal levels for bone sialoprotein, osteocalcin, and osteopontin existed between neighboring mature osteoblasts, particularly after 1,25 (OH)2 D3 treatment, which may therefore selectively affect mature osteoblasts, depending on their differentiation status or functional stage of activity.
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  • 10
    ISSN: 1432-0878
    Keywords: Key words Thyroid gland ; Calcitonin ; C-cell hyperplasia ; C-cell carcinoma ; Medullary thyroid carcinoma ; Sexual dimorphism ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  In many rat strains, C-cell hyperplasia occurs in an age-dependent manner and is often associated with multifocal C-cell carcinoma. The purpose of this study was to investigate the spectrum of spontaneous, proliferative C-cell disorders by gender in Wistar rats throughout their lifespan. The incidence of C-cell hyperplasia shows a significant increase with age (P〈0.001) and is much higher in female rats than in male rats (P〈0.05). From 3 to 24 months of life, 27.5% of female rats showed a normal C-cell pattern, 55.0% showed C-cell hyperplasia, and 17.5% showed C-cell tumors; while 57.5% of male rats showed a normal C-cell pattern, 32.5% showed C-cell hyperplasia, and 10% showed C-cell tumors. Although the overall frequency of C-cell neoplasms in females was nearly double that in males, these data are not statistically significant. However, the number of C-cell tumors showed a significant increase with age (P〈0.05). Therefore, we can conclude that there were significant differences in the incidence of the total spectrum of C-cell proliferative abnormalities in the thyroid gland of Wistar rats that were both age-dependent and gender-dependent.
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  • 11
    ISSN: 1432-0878
    Keywords: Key words Enteric innervation ; Immunohistochemistry ; Nitric oxide synthase ; Galanin ; Striated muscle ; Esophagus ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The relationship between nitric oxide synthase (NOS)- and galanin-immunoreactive nerve terminals and the origin of NOS-immunoreactive nerve terminals on the motor endplates in the striated muscles of the rat esophagus was investigated. Double immunohistochemical staining revealed a dual innervation of motor endplates by calcitonin gene-related peptide (CGRP)-immunoreactive axons and by axons that were immunoreactive for both NOS and galanin. On average, 91% of NOS terminals were galanin immunoreactive. NOS-immunoreactive fibers were revealed at 67% of endplates, identified by the presence of CGRP terminals. The left vagus and superior laryngeal nerve were cut and 15 days allowed for terminals to degenerate. This caused a significant loss of CGRP fibers, but did not affect the density of innervation of the striated muscle by NOS-immunoreactive fibers. Thus the NOS/galanin fibers are deduced to originate from ganglia in the esophageal wall. This is supported by our observation of numerous NOS-immunoreactive nerve cell bodies in the myenteric ganglia of the esophagus, 74% of which were galanin immunoreactive. There were no CGRP-immunoreactive nerve cell bodies in the wall of the esophagus.
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  • 12
    Electronic Resource
    Electronic Resource
    Springer
    Cell & tissue research 295 (1999), S. 363-367 
    ISSN: 1432-0878
    Keywords: Key words Primary hepatocytes ; Oxidative stress ; Haem oxygenase ; Manganese superoxide dismutase ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract We show that mechanical separation of adherent rat primary hepatocytes after the monolayer-forming stage causes the induction of the oxidative stress genes HO-1 (haem oxygenase) and MnSOD (manganese superoxide dismutase). The procedures for enzymatically breaking up liver tissue structure and isolating hepatocytes do not cause HO-1 and MnSOD activation. Only after a 3-h incubation, during which hepatocytes form a monolayer on culture dishes, does the hydrodynamic shearing away of necrotic cells sticking to the monolayer surface activate these two genes. Analysis of this injury-response pathway shows that oxidative stress and mitochondrial dysfunction play a role, as activation can be repressed by antioxidants and by respiratory inhibitors. Recovery of the cells takes a further 24-h incubation during which HO-1 and MnSOD expression returns to basal levels.
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  • 13
    ISSN: 1432-0878
    Keywords: Key words Coagulating gland ; Apocrine secretion ; Merocrine secretion ; Immunocytochemistry ; Immunoelectron microscopy ; Scanning electron microscopy ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The coagulating gland of the rat synthesizes two prevalent secretory proteins (transglutaminase and 115 K) that are discharched in a different manner, one being secreted in an apocrine fashion (transglutaminase) and the other one in a merocrine way (115 K). Differences in the intra- cellular pathway and the release of either protein were studied using immunofluorescence on semithin sections, immunoelectron microscopy of preembedding-processed chopper sections and postembedding-processed ultrathin sections of rat coagulating gland. Immunohistochemical staining using an anti-transglutaminase antibody resulted in dense labeling of the cytoplasm of secretory cells and their apical blebs, whereas the cisternae of the rough endoplasmic reticulum and the Golgi apparatus were completely unlabeled. When, on the contrary, the anti-115 K antiserum was used, dense labeling of the cisternae of the rough endoplasmic reticulum, the Golgi apparatus, and the secretory granules was seen. Intraluminal secretion was also labeled, but the secretory blebs remained unlabeled. Our findings show that, in the coagulating gland of the male rat, the two secretory proteins studied are processed in parallel, but at completely different intracellular pathways. They are released via different extrusion mechanisms. Transglutaminase is synthesized outside the endoplasmic reticulum, reaches the apical cell pole by free flow in the cytoplasm, and is released via apocrine blebs, the membranes of which appear to be derived from the apical plasma membrane. The protein 115 K, on the other hand, follows the classic route, being synthesized within the cisternae of rough endoplasmic reticulum, subsequently glycosylated in the Golgi apparatus, and released in a merocrine fashion. The mutual exclusion of the two secretory pathways and the regulation of the alternative release mechanism are still unresolved issues.
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  • 14
    ISSN: 1432-0878
    Keywords: Key words Growth-associated protein 43 (GAP-43) ; B50 ; Trigeminal sensory system ; Neuroplasticity ; Skin ; Mucosa ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  In this immunocytochemical study we investigated the distribution of nervous structures in the lower lip of adult rats. The region is characterized by a rich cutaneous and mucosal sensory innervation originating from terminal branches of the trigeminal system. Lower lip innervation was investigated by detection of the general neuronal marker protein gene product 9.5 (PGP 9.5) and the growth-associated protein 43 (GAP-43), a neurochemical marker of neuronal plasticity. The entire neural network of both cutaneous and mucosal aspects was stained by the antibody to PGP 9.5. In particular, nerve fibers were observed in the submucosal and the subepithelial plexuses. Thin immunoreactive fibers were observed within the epithelial layers ending as free fibers or as fibers associated with immunopositive Merkel cells. Well-identified anatomical structures receiving sensory or autonomic innervation were also surrounded by PGP 9.5-ir nerve fibers, in particular, hair follicles, vibrissae, glands, and blood vessels. GAP-43-immunostained nerve fibers were observed in all these structures; however, they were generally less numerous than the PGP 9.5-immunoreactive elements. An equal amount of PGP 9.5 and GAP-43 immunoreactivity occurred, in contrast, in the subepidermal and the submucosal plexuses, or in the epidermis and the mucosal epithelium. The present results show that GAP-43 is normally expressed in the mature trigeminal sensory system of the rat. Skin and oral mucosa are characterized by continuous remodeling that may also involve the sensory nervous apparatus. Continuous neural remodeling, regeneration and sprouting may be the reason for the observed expression of GAP-43.
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  • 15
    Electronic Resource
    Electronic Resource
    Springer
    Cell & tissue research 295 (1999), S. 279-285 
    ISSN: 1432-0878
    Keywords: Key words Spermatids ; Spermatozoa ; Spermatogenesis ; Cell volume reduction ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Cell volume reduction is one of the most distinct morphological changes during spermiogenesis and may be largely attributable to water efflux from the cell. A strong candidate for a water efflux route, aquaporin 7 (AQP7), which is a water channel, was studied immunohistochemically in the rat testis. Immunoreactivity was restricted within the elongated spermatids, testicular spermatozoa, and residual bodies remaining in the seminiferous epithelium. Weak but distinct immunoreactivity was first observed in the cytoplasmic mass of the spermatid at step 8 of spermiogenesis. The Golgi-like apparatus became steadily immunoreactive at step 10. The plasma membrane covering the cytoplasmic mass showed strong immunoreactivity after step 16. At this step, the middle piece of the tail also showed immunoreactivity at the portion protruding into the lumen. The whole head and distal tail, where the elongated spermatid had only a limited amount of cytoplasm, showed no immunoreactivity throughout spermiogenesis. After spermiation, the immunoreactivity of AQP7 remained at the middle piece and in the cytoplasmic droplet in the testicular spermatozoon. The present observations suggest that AQP7 contributes to the volume reduction of spermatids, since this water channel protein is localized on the plasma membrane covering the condensing cytoplasmic mass of the elongated spermatid, and since the seminiferous tubule fluid is hypertonic.
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  • 16
    ISSN: 1432-0878
    Keywords: Key words Water channel protein ; Aquaporin ; AQP5 ; Rat ; Salivary glands ; Immunolocalization ; Secretory stimulation ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Aquaporin-5 (AQP5) is a water channel protein and is considered to play an important role in water movement across the plasma membrane. We raised anti-AQP5 antibody and examined the localization of AQP5 protein in rat salivary and lacrimal glands by immunofluorescence microscopy. AQP5 was found in secretory acinar cells of submandibular, parotid, and sublingual glands, where it was restricted to apical membranes including intercellular secretory canaliculi. In the submandibular gland, abundant AQP5 was also found additionally at the apical membrane of intercalated duct cells. Upon stimulation by isoproterenol, apical staining for AQP5 in parotid acinar cells tended to appear as clusters of dots. These results suggest that AQP5 is one of the candidate molecules responsible for the water movement in the salivary glands.
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  • 17
    ISSN: 1432-0878
    Keywords: Key words TGF-β ; CNS ; Epithelial tissues ; Macrophages ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  We have cloned a novel member of the transforming growth factor-β (TGF-β) superfamily from a human placental cDNA library. The sequence is identical to five very recently published sequences, of which only one (macrophage inhibitory cytokine-1, MIC-1) has been characterized in terms of function. In light of the present data demonstrating the wide distribution of the mRNA and putative multifunctionality, we propose to name this molecule growth/differentiation factor-15/MIC-1 (GDF-15/MIC-1). The deduced amino acid sequence reveals typical features of a secreted molecule. The epithelium of the choroid plexus is the only site in the adult brain expressing detectable levels of GDF-15/MIC-1 mRNA. Many epithelia of non-neural tissues including those of the prostate and intestinal mucosa, bronchi and bronchioli, secretory tubuli of the submandibular gland, and lactating mammary gland are prominent sites of GDF-15/MIC-1 synthesis. GDF-15/MIC-1 is also strongly expressed by macrophages in the adrenal gland. Thus, GDF-15/MIC-1, like many other members of the TGF-β superfamily, is widely distributed in adult tissues, being most strongly expressed in epithelial cells and macrophages.
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  • 18
    ISSN: 1432-0878
    Keywords: Key words Neurokinin-1 receptor ; Substance P ; Junctional epithelium ; Neutrophil ; Endosomes ; Immunoelectron microscopy ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  The distributions of substance P (SP) and the neurokinin-1 receptor (NK1-R), the receptor preferentially activated by SP, were examined in rat gingiva by immunocytochemical methods with light and electron microscopy. SP-immunoreactive nerve fibers were located preferentially in the junctional epithelium (JE) but few in the other oral and oral sulcular epithelia. NK1-R immunoreactivity was found in the endothelial cells (capillaries and postcapillary venules underlying the JE). NK1-R-labeled and -unlabeled unmyelinated nerve fibers were located close to the blood vessels and partially or completely covered by a Schwann cell sheath. In the JE, labeled naked axons without Schwann cell sheaths were observed. Neutrophils and macrophages in the connective tissue underlying the JE and in the JE were also labeled with NK1-R. Furthermore, NK1-R was found in the JE cells. Basically, immunoreaction products for NK1-R were found throughout various cells (endothelial cells, neutrophils, and JE cells) at invaginations of the plasma membrane and in vesicular and granular structures that are probably endosomes and are found close to both the plasma membrane and the nucleus. This is a first report, demonstrating the presence of NK1-R in the gingival tissue in the normal nonstimulated condition. Furthermore, it is thought that SP may modulate the permeability of blood vessels beneath the JE, the production of antimicrobial agents in neutrophils, and the proliferation and endocytotic ability of JE cells through NK1-R.
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  • 19
    ISSN: 1432-0878
    Keywords: Key words Glucocorticoid receptor (GR) ; Immunohistochemistry ; In situ hybridization ; Pituitary ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract By means of double immunohistochemical techniques and a nonradioisotopic in situ hybridization method, we determined the colocalization pattern of glucocorticoid receptor (GR) and pituitary hormones and the GR messenger RNA (mRNA) expression in the pituitaries of Wistar adult male rats. Immunoreactivity for GR was detected in the nuclei of cells in the anterior and posterior pituitary. Double immunohistochemistry revealed that the colocaliza- tion of GR and anterior pituitary hormones occurred in almost 99% of the growth hormone (GH)-producing cells and adrenocorticotropic hormone (ACTH)-producing cells, and in 67% of the thyroid stimulating hormone (TSH)-producing cells. Almost all of the folliculostellate cells (93%), marginal layer cells (94%) in the anterior pituitary, and pituicytes (96%) in the posterior pituitary immunostained for S100 protein antibody were also immunostained with GR. GR mRNA was abundant in the cytoplasm of anterior and intermediate pituitary cells but scattered sparsely in that of the posterior pituitary. These results suggest that glucocorticoids directly influence certain pituitary cells in order to regulate cell function, including the synthesis and/or secretion of hormones.
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  • 20
    ISSN: 1432-0878
    Keywords: Key words Adrenal medulla ; Chromaffin cells ; Neurotrophins ; Neurotrophin receptors ; c-fos ; Neuro-endocrine-immune interactions ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Macrophages are widely distributed in lymphohaemopoietic and many other mammalian tissues, where they are mainly involved in host defence mechanisms, phagocytosis, wound repair, and secretion of growth factors. Increasing evidence suggests that secretory products of macrophages can influence adrenal gland functions. In the present study, we have used specific antibodies to ED1 (cytoplasmic antigen), ED2 (membrane antigen), ED8 (membrane antigen), and OX-6 (MHC class II/membrane antigen) as markers for macrophages to examine their distribution within the adult rat adrenal gland. ED2 and OX-6 recognize distinct subpopulations of adrenal gland macrophages, whereas macrophages immunoreactive (-ir) for ED1 and ED8 could not be detected. OX-6-ir macrophages were most numerous in the cortical reticularis and glomerulosa zones, while only few cells were found in the zona fasciculata and in the adrenal medulla. Macrophages immunoreactive for ED2 were restricted to the adrenal medulla. The majority of these macrophages were associated with vascular sinuses or chromaffin cells. By double-immunolabelling we found that most of ED2-ir medullary macrophages contain neurotrophin-4 (NT-4)-like ir. Attempts to clarify whether macrophages take up NT-4 from NT-4-ir chromaffin cells indicated that medullary macrophages are immunonegative for chromogranin A and neuropeptide Y, two major secretory products of chromaffin cells. In situ hybridizations and immunofluorescence showed expression of the neurotrophin receptor TrkA, but not TrkB in the adrenal medulla. In vitro studies indicated that NT-4, similar to nerve growth factor, can induce c-fos-ir in chromaffin cells. We conclude that chromaffin cells are putative targets for adrenal medullary NT-4, whose functions remain to be clarified.
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  • 21
    ISSN: 1432-0878
    Keywords: Key words Hydroxyindole-O-methyltransferase ; Pineal gland ; In situ hybridisation ; Melatonin ; 5-Methoxyindoles ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Hydroxyindole-O-methyltransferase (HIOMT) is the enzyme involved in the last step of the melatonin synthesis pathway. Recently, a cDNA encoding HIOMT has been isolated from a rat pineal gland library. Using this cDNA, we developed a highly sensitive in situ hybridisation protocol to investigate the distribution of HIOMT mRNA in both the rat brain and dissociated pinealocytes maintained in primary cell culture. In the rat brain, HIOMT mRNA was only detected in the three parts of the pineal complex: the superficial pineal, the stalk and the deep pineal. No extra-pineal hybridisation labelling was observed. These results strongly suggest that melatonin synthesis also occurs in the deep part and the stalk of the pineal gland. HIOMT mRNA was markedly expressed in cultured pinealocytes. No particular subcellular area was observed to express HIOMT mRNA specifically, as the labelling was homogeneously distributed in the cytosol and in the axon-like processes. In conclusion, the use of in situ and in vitro hybridisation with a pineal riboprobe has detected notable HIOMT expression restricted to pinealocytes.
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  • 22
    ISSN: 1432-0878
    Keywords: Key words EDL ; Soleus ; Satellite cells ; Proliferation ; Differentiation ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Following muscle damage, fast- and slow-contracting fibers regenerate, owing to the activation of their satellite cells. In rats, crush-induced regeneration of extensor digitorum longus (EDL, a fast muscle) and soleus (a slow muscle) present different characteristics, suggesting that intrinsic differences exist among their satellite cells. An in vitro comparative study of the proliferation and differentiation capacities of satellite cells isolated from these muscles is presented there. We observed several differences between soleus and EDL satellite cell cultures plated at high density on gelatin-coated dishes. Soleus satellite cells proliferated more actively and fused into myotubes less efficiently than EDL cells. The rate of muscular creatine kinase enzyme appeared slightly lower in soleus than in EDL cultures at day 11 after plating, when many myotubes were formed, although the levels of muscular creatine kinase mRNA were similar in both cultures. In addition, soleus cultures expressed higher levels of MyoD and myogenin mRNA and of MyoD protein than EDL satellite cell cultures at day 12. A clonal analysis was also carried out on both cell populations in order to determine if distinct lineage features could be detected among satellite cells derived from EDL and soleus muscles. When plated on gelatin at clonal density, cells from both muscles yielded clones within 2 weeks, which stemmed from 3–15 mitotic cycles and were classified into three classes according to their sizes. Myotubes resulting from spontaneous fusion of cells from the progeny of one single cell were seen regardless of the clone size in the standard culture medium we used. The proportion of clones showing myotubes in each class depended on the muscle origin of the cells and was greater in EDL- than in soleus-cell cultures. In addition, soleus cells were shown to improve their differentiation capacity upon changes in the culture condition. Indeed, the proportions of clones showing myotubes, or of cells fusing into myotubes in clones, were increased by treatments with a myotube-conditioned medium, with phorbol ester, and by growth on extra-cellular matrix components (Matrigel). These results, showing differences among satellite cells from fast and slow muscles, might be of importance to muscle repair after trauma and in pathological situations.
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  • 23
    ISSN: 1432-0878
    Keywords: Key words Calretinin ; Calcium-binding proteins ; Monosodium glutamate ; Excitotoxicity ; Area postrema ; Circumventricular organs ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  We mapped the distribution of calretinin-immunoreactive neuron populations in a circumventricular organ of the rat, the area postrema, and investigated their sensitivity to excitotoxic stimuli mediated by subcutaneously administered monosodium glutamate. We were specifically interested to ascertain whether the presence of calretinin can, per se, confer an in vivo intrinsic resistance for area postrema neurons to glutamate excitotoxicity. We found that dense populations of calretinin-positive neurons displayed a subregional compartmentation in coronal sections of the area postrema along its rostrocaudal axis. We demonstrated that calretinin-positive neurons differ in their sensitivities to monosodium glutamate depending on their position within the area postrema. Neurons in the caudal area postrema were the most sensitive ones, while those in the rostral area postrema were spared of degeneration. We conclude that calretinin-positive neurons in the area postrema are not uniformly protected against glutamate excitotoxicity. It is possible that differences in the local concentrations of monosodium glutamate due to regional heterogeneities in density and permeability of the capillary bed rather than neuronal expression of calretinin account for the observed effects.
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  • 24
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    Cell & tissue research 294 (1998), S. 57-68 
    ISSN: 1432-0878
    Keywords: Key words Calcium-binding protein ; Nerve terminal ; Motor endplate ; Vagus nerve ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Neurocalcin is a newly identified neuronal calcium-binding protein. We tried here to investigate the immunohistochemical distribution of neurocalcin in the rat esophagus. Nerve cell bodies having neurocalcin immunoreactivity were found throughout the myenteric plexus. In the myenteric ganglia, two types of nerve terminals showed neurocalcin immunoreactivity. One was varicose terminals containing numerous small clear vesicles and forming a synapse with nerve cells. The other terminals were characterized by laminar or pleomorphic structure and many mitochondria. These laminar terminals were supposed to be sensory receptors of the esophageal wall. In the motor endplates of the striated muscles, nerve terminals containing many small clear vesicles and mitochondria also had neurocalcin immunoreactivity. After left vagus nerve cutting under the nodose ganglia, the number of immunopositive thick nerve fibers, laminar endings and nerve terminals on the striated muscles decreased markedly. Retrograde tracing experiments using Fast Blue showed extrinsic innervation of esophagus from ambiguus nucleus, dorsal motor nucleus of vagus, superior cervical ganglia, celiac ganglia, nodose ganglia and dorsal root ganglia. In the celiac ganglia, nodose ganglia and dorsal root ganglia, retrogradely labeled nerve cells were neurocalcin-immunoreactive. Neurons in the celiac ganglia may project varicose terminals, while nodose and dorsal root neurons project laminar terminals. Although cell bodies of motoneurons in the ambiguus nucleus lacked neurocalcin immunoreactivity, these neurons may contain neurocalcin only in the nerve terminals in the motor endplates. Neurocalcin immunoreactivity is distributed in many extrinsic and intrinsic neurons in the esophagus and this protein may play important roles in regulating calcium signaling in the neurons.
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  • 25
    ISSN: 1432-0878
    Keywords: Key words Periodontal ligament ; α-Smooth muscle actin ; Osteopontin ; Bone sialoprotein ; Bone morphogenic protein ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Periodontal ligament width is precisely maintained throughout the lifetime of adult mammals but the biological mechanisms that inhibit ingrowth of bone into this soft connective tissue are unknown. As bone morphogenic proteins strongly stimulate osteogenesis and can induce ectopic bone formation in vivo, we tested the hypothesis that topical application of this powerful osteogenic agent will overwhelm the osteogenic inhibitory mechanisms of periodontal ligament cells and induce ankylosis. Wounds through the alveolar bone and periodontal ligament were created in 45 male Wistar rats. Defects were filled with either a collagen implant or collagen plus bone morphogenic protein (BMP-7), or were left unfilled (controls). Three animals per time period were killed on days 2, 5, 10, 21 and 60 after surgery for each wound type. Cellular proliferation and clonal growth in periodontal tissues were assessed by 3H-thymidine labeling 1 h before death, followed by radioautography. Cellular differentiation of soft and mineralizing connective tissue cell populations was determined by immunohistochemical staining of α-smooth muscle actin, osteopontin and bone sialoprotein. In regenerating periodontium, BMP-7 induced abundant bone formation by 21 days (2.5-fold greater than controls or collagen implant only; P〈0.001), but by day 60 the volume of the newly formed bone had returned to baseline levels and was similar for all groups. Independent of the type of treatment, periodontal ligament width was unchanged throughout the experimental period (P〉0.05). Animals treated with BMP-7 implants showed greatly increased cellular proliferation in the periodontal ligament adjacent to the wound site and in the regenerating alveolar bone at days 5 and 10 after wounding compared to the other treatment groups (P〈0.005). Animals in the BMP-7 group exhibited similar spatial and temporal staining patterns for α-smooth muscle actin, osteopontin and bone sialoprotein as controls. Collectively, these data show that BMP-7 promoted the proliferation of precursor cells in the periodontal ligament but did not induce osteogenic differentiation in this compartment. Consequently a powerful osteogenic stimulus like BMP-7 cannot significantly perturb the mechanisms that regulate periodontal ligament width and maintain periodontal homeostasis.
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  • 26
    ISSN: 1432-0878
    Keywords: Key words: Amperometry  ; Calcium currents ; Catecholamines ; Chromaffin cells ; Differentiation ; Magnetic field ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  The ion Ca2+ has been shown to play an important role in a wide variety of cellular functions, one of them being related to cell differentiation in which nerve growth factor (NGF) is involved. Chromaffin cells obtained from adrenals of 2- to 3-day-old rats were cultured for 7 days. During this time, these cells were subjected to the application of either NGF or extremely low frequency magnetic fields (ELF MF). Since this induced cell differentiation toward neuronal-like cells, the mechanism by which this occurred was studied. When the L-Ca2+ channel blocker nifedipine was applied simultaneously with ELF MF, this differentiation did not take place, but it did when an N-Ca2+ channel blocker was used. In contrast, none of the Ca2+ channel blockers prevented differentiation in the presence of NGF. In addition, Bay K-8644, an L-Ca2+ channel agonist, increased both the percentage of differentiated cells and neurite length in the presence of ELF MF. This effect was much weaker in the presence of NGF. [3H]-noradrenaline release was reduced by nifedipine, suggesting an important role for L-Ca2+ channels in neurotransmitter release. Total high voltage Ca2+ currents were significantly increased in ELF MF-treated cells with NGF, but these currents in ELF MF-treated cells were more sensitive to nifedipine. Amperometric analysis of catecholamine release revealed that the KCl-induced activity of cells stimulated to differentiate by ELF MF is highly sensitive to L-type Ca2+ channel blockers. A possible mechanism to explain the way in which the application of magnetic fields can induce differentation of chromaffin cells into neuronal-like cells is proposed.
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  • 27
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    Cell & tissue research 291 (1998), S. 277-283 
    ISSN: 1432-0878
    Keywords: Key words Diurnal pattern ; Pancreas ; Acinar cell replication ; Cholecystokinin (CCK) ; Proliferating cell nuclear antigen (PCNA) ; 5-Bromodeoxyuridine (BrdU) ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Fully differentiated pancreatic acinar cells can enter the cell cycle under appropriate conditions in the rat. The aim of this study was to analyse the diurnal pattern of acinar cell proliferation as a function of food intake and the release of cholecystokinin (CCK), because the peptide hormone CCK is a major physiological regulator of rat pancreatic acinar cell replication. Pancreatic acinar cell replication was quantitated using an antibody against the S-phase marker proliferating cell nuclear antigen (PCNA). In addition, acinar cells in S-phase were detected after injecting bromodeoxyuridine (BrdU) and subsequent immunohistochemical staining of BrdU-positive nuclei. Rat pancreata were analysed during the day under standard diet conditions, as well as after various schedules of fasting and refeeding and after the application of the CCK receptor antagonist L-364,718. Between 6 a.m. and 6 p.m., the PCNA labeling index was 4.4±0.9%, while between 9 p.m. and 3 a.m. the PCNA labeling index was elevated and reached peak values of 11.4% (mean value: 7.8±2.5%) around midnight. BrdU-positive cells also doubled around midnight, compared to the 9:00 a.m. value. In fasted rats, acinar cell proliferation was completely suppressed and this suppression could be overcome by injection of the CCK analog cerulein. In addition, the CCK antagonist L-364,718 led to the same results as fasting. Here we show for the first time that there is a diurnal pattern of pancreatic acinar cell proliferation in rats, which is dependent on food intake and is mediated by CCK.
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  • 28
    ISSN: 1432-0878
    Keywords: Key words Intergranular bridges ; Granule-granule fusion ; Annexin II ; Anterior pituitary cells ; Multi granular exocytosis ; Clostridium perfringens enterotoxin ; Quick-freeze deep-etch microscopy ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Quick-freeze deep-etch electron microscopy showed the presence of bridge-like structures between adjacent secretory granules in rat anterior pituitary secretory cells. These intergranular bridges were variable in length and thickness. The finest bridges were 7–8 nm in length, while the longest ones were as long as 80 nm. Annexin II, one of the Ca2+-dependent phospholipid-binding proteins, is known to interlink between two membranes and induce aggregation of liposomes and chromaffin granules under the presence of Ca2+. In anterior pituitary cells, annexin II was detected by immunoelectron microscopy at the contact sites of secretory granules with other granules. The anterior pituitary cells treated under the presence of extracellular Ca2+ with Clostridium perfringens enterotoxin which induces Ca2+ influx showed multigranular exocytosis, i.e., multiple fusions of secretory granules with each other and with the plasma membrane. The granule-granule fusion in progress could be captured by the quick-freeze deep-etch technique. The membranes of adjacent secretory granules were partially fused at their contact sites where intergranular strands were no longer seen, while there existed intergranular strands between unfused portions of the granule membranes. From these results, we consider that the intergranular bridges, some of which may be composed of annexin II, are involved in Ca2+-induced granule-granule fusion in anterior pituitary cells.
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  • 29
    ISSN: 1432-0878
    Keywords: Key words Juxtaglomerular cells ; Stereology ; Disector ; Salt diets ; Enalapril ; Renal perfusion pressure ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  In order to study the cellular mode of renin secretion, stereological methods were used to estimate number and volume of rat renin secretory granules during stimulation of the renin system. An acute decrease in renal perfusion pressure to 40 mmHg for 5 min increased plasma renin concentration (PRC) twofold, but did not significantly change the number of renin granules per arteriole or the renin-containing volume of the arteriole. Chronic stimulation was achieved by a combination of low-salt diet and inhibition of angiotensin-converting enzyme (ACE) for 14 days, and resulted in a 36-fold increase in PRC, a 20-fold increase in the number of granules per arteriole, and a 17-fold increase in the arteriolar volume that contained renin. An acute decrease in renal perfusion pressure to 40 mmHg for 5 min in the chronically stimulated rats increased PRC further (1.6-fold), and significantly reduced the number of granules per arteriole by 4000 (45% reduction), but did not change the renin-containing arteriolar volume significantly. The average renin granule size was 0.35 μm3 with no significant differences among the groups. We conclude that recruited granular cells contribute significantly to renin release, and that all granular cells along the arteriole participate in secretory responses. The reduced number of renin granules after acute stimulation is compatible with exocytosis as the dominating mechanism of renin release.
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  • 30
    ISSN: 1432-0878
    Keywords: Key words Pepsinogen C ; Ontogeny ; Mucous neck cell ; Chief cell ; Intermediate mucopeptic cell ; Immunocytochemistry ; In situ hybridization ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  The ontogeny of pepsinogen C-producing cells in rat fundic glands was studied by means of light and electron microscopy using an antiserum raised against a synthetic peptide based on rat pepsinogen C. To confirm the immunocytochemistry results, the expression of rat pepsinogen C messenger RNA (mRNA) in the fundic gland was also examined by in situ hybridization using a digoxigenin-labeled RNA probe. In adult rats, pepsinogen C was produced by chief cells, mucous neck cells, and intermediate mucopeptic cells. Pepsinogen C-producing cells appeared in embryos as early as 18.5 days’ gestation. The development of these cells could be classified into four stages: (1) 18.5 days’ gestation to 0.5 days after birth; (2) 0.5 days to 2 weeks after birth; (3) 3–4 weeks after birth; (4) 4–8 weeks after birth. In embryos and young animals, pepsinogen C-producing cells were mucopeptic cells. By 4 weeks after birth, mucous neck cells could be distinguished morphologically. The maturation stages of the chief cells could be traced by electron microscopy along the longitudinal axis of the rat fundic gland by double-staining with anti-pepsinogen C antibody and periodic acid-thiocarbohydrazide-silver proteinate. Positive reactions for pepsinogen C and pepsinogen C mRNA expression were detected in mucous neck cells. Therefore, we conclude that mucous neck cells are precursor cells of chief cells. Mucous neck cells, intermediate cells, and chief cells are in the same differentiating cell lineage.
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  • 31
    ISSN: 1432-0878
    Keywords: Key words Osteoclast ; Bone ; Bone resorption ; Alveolar bone ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Glutaraldehyde-formaldehyde fixed undecalcified alveolar bone from 7-day-old rats was prepared for light and electron microscopy. Colloidal lanthanum was used as an ultrastructural tracer, and both random and semi-serial sections were examined. Lanthanum penetrated the infoldings of the ruffled border and some nearby vacuoles and vesicles. The majority of vacuoles and vesicles were lanthanum-free. Some osteoclast profiles contained a large vacuole with a cell enclosed in its interior. The enclosed cell exhibited an irregular nucleus containing condensed peripheral chromatin, intact cytoplasmic organelles, conspicuous rough endoplasmic reticulum and large blebs on the cell surface. These features are characteristic of osteoblasts or bone-lining cells or immature osteocytes which may be undergoing apoptosis or necrosis. The observation of remnants of cellular structures within internalized osteoclast vacuoles, together with the above results, suggests that osteoclasts engulf and probably degrade dying osteoblasts/bone-lining cells or immature osteocytes.
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  • 32
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    Cell & tissue research 294 (1998), S. 253-260 
    ISSN: 1432-0878
    Keywords: Key words P2X1 receptor ; Ultrastructure ; Cerebellum ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  The distribution of the P2X1 subtype of purinoceptors associated with the extracellular activities of ATP was studied in the rat cerebellum at the electron-microscope level. Receptors were labelled with peroxidase-antiperoxidase and the avidin-biotin-peroxidase complex for immunocytochemistry. Immunoreactivity to P2X1 receptors was localized in subpopulations of synapses between varicosities of parallel fibres of granule cells and dendritic spines of Purkinje cells. Unlabelled varicosities of parallel fibres formed asymmetric synapses with labelled dendritic spines, whereas labelled varicosities of parallel fibres formed asymmetric synapses with unlabelled dendritic spines. P2X1 immunoreactivity was also localized in some astrocyte processes. The functional significance of these findings is discussed.
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  • 33
    ISSN: 1432-0878
    Keywords: Key words Coagulating gland ; Epididymis ; Gene expression ; Prostate ; Seminal vesicle ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  We studied the expression and distribution of transforming growth factor-β (TGF-β) isoforms in the rat male accessory sex glands and the epididymis. Our data demonstrate the expression of both TGF-β1 and -β3 isoforms in ventral prostate (VP), seminal vesicle (SV), coagulating gland (CG), and epididymis (E) by Northern blot analysis. In addition, there was differential expression of TGF-β3 in the three regions of epididymis, the corpus region being the highest. Immunostaining data showed intense staining for latent TGF-β1 in all the male accessory glands. In contrast, no staining using antibodies specific for active TGF-β1 was observed. No expression of TGF-β2 was evident either by immunohistochemistry or Northern blot analysis. The presence of mature TGF-β3 protein was observed in the secretory epithelium of VP, CG, and corpus E. There was no detectable staining of TGF-β3 in the seminal vesicle and caput and cauda regions of epididymis. These data suggest possible differential regulation of TGF-β isoform expression in the male reproductive system and predict unique roles for individual TGF-β isoforms in sperm maturation and maintenance.
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  • 34
    ISSN: 1432-0878
    Keywords: Key words Inner ear ; Gap junctions ; Connexins ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Gap junction channels which are responsible for direct intercellular communication are composed of connexin proteins. Different connexins are distributed in a tissue-specific manner. Up to now only connexin26 has been identified to be widely expressed in the inner ear. In order to investigate the role of additional gap junction proteins, the expression of connexin30 and 43 was investigated in the rat cochlea. Connexin26 and connexin30 were both expressed in the spiral limbus, the spiral ligament, the stria vascularis and between supporting cells of the organ of Corti. Double-labeling experiments suggest that both connexins are partly colocalized between cells. Weak staining of connexin43 could only be detected in the stria vascularis, the spiral ligament and between organ of Corti supporting cells. The corresponding transcripts for connexin26, 30 and 43 could be detected by Northern blot analysis. The expression of different gap junction channels in the cochlea suggests functional diversity. Gap junctions in the inner ear may control ion concentrations of cochlear fluids or act as conduits through which glucose and other metabolites diffuse.
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  • 35
    ISSN: 1432-0878
    Keywords: Key words Odorant receptors ; Olfactory epithelium ; Regeneration ; Bulbectomy ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The effects of unilateral olfactory bulb ablation upon the odorant receptor expression were studied during the degeneration/regeneration process in the olfactory epithelium of adult rats. Using the in situ hybridization approach, we compared the time course of decay and recovery of expression for three different receptor subtypes (OR14, OR5, OR124). The number of neurons expressing receptor subtypes dramatically decreased in the olfactory epithelium on the lesioned side and reached a minimum at day 5 postsurgery. A progressive recovery was then observed from day 5 to day 15 postlesion, when a plateau was reached. Noticeable differences in the recovery level of receptor expression were observed according to the zonal patterning: the recovery level for neurons located in the lateral zone reached 70% of the control side value while the recovery levels in the dorsal and medial zones represented 35% and 53% of this value, respectively. Axotomy experiments suggest that zone-specific differences in receptor reexpression reported after bulbectomy might be related to the trophic influence of the olfactory bulb.
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  • 36
    ISSN: 1432-0878
    Keywords: Key words Hepatocytes ; Cell polarity ; Cytoskeleton ; Cytokeratin ; Bile canaliculus ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  The usefulness of cultured hepatocytes is limited by the gradual loss of their typical physiological functions that occurs in vitro, mainly due to the absence of microenviromental conditions found in vivo. In this study we describe the effect of retinoic acid on the re-establishment of morphological characteristics and on the reorganization of the cytoskeletal network in cultured rat hepatocytes. Results obtained demonstrate that retinoic acid can influence hepatocyte differentiation, as regards the recovery of cell polarity, polyhedric shape and reformation of bile canaliculi and junctional complexes. The main target of this action appears to be the cytoarchitecture of cytoskeletal components, particularly cytokeratin filaments, which regain the configuration present in intact liver. The reorganization of the intermediate filaments does not seem to be dependent on the induction of higher levels of cytokeratin proteins, but rather appears to be due to post-translational regulation. The effect of retinoic acid on the cytoskeletal organization could determine the stabilization of intercellular contacts by means of junctions, leading to the appearance of morpho-functional characteristics typical of well-differentiated hepatocytes.
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  • 37
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    Cell & tissue research 293 (1998), S. 357-364 
    ISSN: 1432-0878
    Keywords: Key words Thyroxine ; Cartilage ; Differentiation ; Type X collagen ; Organ culture ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Thyroid hormone plays a role in skeletal maturation. The present study was undertaken to investigate the effect of thyroid hormone on growth and maturation of the epiphyseal cartilage in the rat femur organ culture system. The femora increased in both weight and size over time in culture in the absence of serum. The growth rate was suppressed by thyroxine treatment. Thyroxine induced chondrocyte hypertrophy in the area adjacent to the articular surface in the femoral condyle. In this area, the expression of type X collagen, a marker of chondrocyte terminal differentiation, was detected after 21 days in culture by immunohistochemistry and reverse transcription/polymerase chain reaction. These data suggest that thyroxine suppresses cartilage growth by stimulating chondrocyte terminal differentiation.
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  • 38
    ISSN: 1432-0878
    Keywords: Key words Lung ; Neuroendocrine system ; Neuroepithelial bodies ; Innervation ; Neuronal tracing ; DiI ; Calcitonin gene-related peptide ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Our present understanding of the morphology of neuroepithelial bodies (NEBs) in mammalian lungs is comprehensive. Several hypotheses have been put forward regarding their function but none has been proven conclusively. Microscopic data on the innervation that appears to affect the reaction of NEBs to stimuli have given rise to conflicting interpretations. The aim of this study has been to check the validity of the hypothesis that pulmonary NEBs receive an extensive vagal sensory innervation. The fluorescent neuronal tracer DiI was injected into the vagal sensory nodose ganglion and NEBs were visualized in toto by using immunocytochemistry and confocal microscopy on 100-µm-thick frozen sections of the lungs of adult rats. The most striking finding was the extensive intraepithelial terminal arborizations of DiI-labelled vagal afferents in intrapulmonary airways, apparently always co-appearing with calcitonin gene-related peptide (CGRP)-immunoreactive NEBs. Not all NEBs received a traced nerve fibre. Intrapulmonary CGRP-containing nerve fibres, including those innervating NEBs, always appeared to belong to a nerve fibre population different from the DiI-traced fibres and hence did not arise from the nodose ganglion. Therefore, at least some of the pulmonary NEBs in adult rats are supplied with sensory nerve fibres that originate from the vagal nodose ganglion and form beaded ramifications between the NEB cells, thus providing support for the hypothesis of a receptor function for NEBs.
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  • 39
    ISSN: 1432-0878
    Keywords: Key words Sinusoidal endothelial cells ; Kupffer cells ; Vital staining ; Lithium carmine ; Endocytosis ; Reticulo-endothelial system ; Mononuclear phagocyte system ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Sinusoidal cells in the rat liver were studied in vivo and in vitro using the original vital staining with lithium carmine, which has contributed much to the development of the concept of the reticulo-endothelial system. Immunohistochemical and electron-microscopic studies revealed that the dye-incorporating cells were sinusoidal endothelial cells, Kupffer cells, and monocytes. The endothelial cells took up much more dye than did the Kupffer cells and bulged largely into the sinusoidal lumen. Electron microscopy revealed that small particles of lithium carmine were associated with coated vesicles of endothelial cells and ruffled membranes of Kupffer cells. In the endothelial cells, these particles were present in various concentrations within vacuolated structures and condensed in the lysosomes forming large aggregates of lithium carmine lumps. These lumps showed crystalline structures, within which the size of the individual particle was up to 30 nm in width and 50 nm in length. A few endothelial cells containing abundant dye underwent degeneration, and some were taken up by Kupffer cells. Liver endothelial cells isolated from lithium carmine-administered rats endocytosed fluorescence-labeled collagen. Isolated endothelial cells from normal rat liver, when cultured with lithium carmine, did not take up any dye, and their endocytosis of formaldehyde-treated albumin was inhibited dose-dependently. We conclude that in the liver, endothelial cells, but not Kupffer cells, predominantly take up lithium carmine. Furthermore, we propose the existence of a generalized cell system based on its vital staining capacity.
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  • 40
    ISSN: 1432-0878
    Keywords: Key words Nucleus ambiguus ; Enteric nervous system ; Motor endplate ; Vasoactive intestinal peptide (VIP) ; Calcitonin gene-related peptide (CGRP) ; Coinnervation ; Anterograde tracing ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  We investigated the origin of galanin-positive nerve fibers on motor endplates in rat esophagus using anterograde 1,1′-dioleyl-3,3,3′,3′-tetramethylindocarbocyanine methane sulfonate (DiI) tracing from the nucleus ambiguus combined with galanin immunocytochemistry and calcitonin gene-related peptide immunocytochemistry. To demonstrate spatial relationships of galanin-positive nerve fibers to vagal and enteric nerve fibers on motor endplates, we combined galanin immunocytochemistry with calcitonin gene-related peptide immunostaining for labeling of vagal terminals, and vasoactive intestinal peptide immunoreactivity and NADPH-diaphorase histochemistry for demonstration of enteric nerve fibers. Within fine varicose nerve fibers, galanin was colocalized with vasoactive intestinal peptide and NADPH-diaphorase to a high degree and turned out to be completely separated from calcitonin gene-related peptide-positive or anterogradely DiI-labeled vagal motor terminals. These results indicate that the enteric nervous system is the most important and possibly the only source of galanin-positive nerve terminals on motor endplates in rat esophagus. Galanin may be, in addition to nitric oxide and vasoactive intestinal peptide, a mediator of the enteric coinnervation of striated muscle in this organ.
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  • 41
    ISSN: 1432-0878
    Keywords: Key words Dendritic cells ; Maturation ; Intrahepatic translocation ; Immunohistochemistry ; Electron microscopy ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Specific populations of hepatic sinusoidal cells were stained with monoclonal antibodies that recognize monocytes/macrophages (ED1), tissue macrophages (Kupffer cells) (ED2), MHC class II (Ia) antigen (MRC OX6), and dendritic cells/γ,δ T-cells (MRC OX62) and analyzed by light and electron microscopy. The majority of ED1+ and/or ED2+ cells were localized to the hepatic parenchyma, whereas OX6+ and/or OX62+ cells were more densely distributed within Glisson’s sheath than in the hepatic parenchyma. Double-immunoperoxidase staining of normal liver for ED1, ED2, and OX6 identified dendritic cells (DC) of two different phenotypes, ED1+ED2–OX6+ and ED1–ED2–OX6+. DC can be classified into three different types based on ultrastructural characteristics. The first type (type I) is characterized by one or more long cytoplasmic processes and a well-developed lysosomal system. The second type (type II) has an inconspicuous lysosomal system, abundant hyaloplasm, and characteristic short cytoplasmic processes. The third type (type I–II) has cytologic features intermediate between those of type I and type II DC. At the electron-microscopic level, these three cell types are found in the sinusoidal lumen, whereas the majority of type II DC are located in the space of Disse and Glisson’s sheath. Furthermore, some OX6-labeled elongated DC appeared to traverse the lumen of sinusoids through endothelial pores to enter the space of Disse. One hour after intravenous injection of latex particles (0.81 μm in diameter), numerous latex-laden dendritic cells (ED1+OX6+, type I and type I–II) were detected in the lumen of hepatic sinusoids, but not in the space of Disse or Glisson’s sheath. These findings suggest that normal rat liver contains resident dendritic cells which downregulate phagocytic activity and mature into potent accessory cells during migration from the portal vein toward the central vein. These DC then traverse the sinusoidal lumen to the hepatic lymph system via the space of Disse.
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  • 42
    ISSN: 1432-0878
    Keywords: Key words: Acinus ; Guanine nucleotide release factor ; Pancreas ; exocrine ; ras ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The polymerase chain reaction was used to amplify a cDNA encoding the catalytic region of ras-guanine nucleotide release factor (ras-GRF1) from mouse embryonic stem cell mRNA. Antibodies directed against this protein were prepared and affinity purified. Western immunoblotting of rat tissue lysates revealed a 140-kDa protein in brain as expected but, in addition, a strongly immunoreactive 54-kDa protein, p54, was identified in pancreas. Expression of ras-GRF1 in pancreas was confirmed at the RNA level by reverse-transcriptase-coupled polymerase chain reaction analysis; p54 may therefore correspond to a form of ras-GRF1 or a closely related protein. The cellular and subcellular localization of p54 was investigated by enzyme-linked immunocytochemistry and immunogold electron microscopy. In the pancreas, p54 was expressed primarily in acinar cells, where it was localized along the basolateral and apicolateral plasma membranes. Indirect immunofluorescence microscopy of cultured acini further indicated that the plasma membrane localization of p54 was dependent on the maintenance of the acinar histotype and organized acinar structure. When primary acinar cells were permitted to dissipate into monolayer cultures devoid of zymogen granules, ras-GRF1 staining became cytosolic. Our results suggest that ras-GRF1 is involved in the structure and function of the pancreas.
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  • 43
    ISSN: 1432-0878
    Keywords: Key words: Intestines ; Neuropeptide Y ; Nitric oxide synthase ; Vasoactive intestinal polypeptide ; Guinea-pig ; Rat (Wistar) ; Mouse (C3H/HeJ ; gld) ; Ferret ; Man
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The Fas ligand induces apoptosis in activated immunocytes that express the Fas receptor. Fas-ligand transcripts have been found previously in murine intestine but the intestinal tissues that express Fas ligand have not been identified. We used immunohistochemistry to examine the expression of the Fas ligand in the enteric nervous system of rats, mice, guinea-pigs, ferrets and humans. Fas-ligand immunoreactivity was detectable in enteric nerve fibres and neurons in all species tested, representing 25%–50% of the neurons in rats, mice and guinea-pigs. An antigen of approximately 48 kDa was detected by Western blot analysis with Fas-ligand antiserum in the dissected enteric plexuses of duodenum from a C3H/HeJ mouse. In gld mice that harbour a Fas-ligand mutation, Fas-ligand immunoreactivity was slightly more intense in neurons and fibres and was also apparent in submucosal lymphocytes. In the myenteric plexuses of guinea-pig ileum and human colon, Fas-ligand immunoreactivity was not contained in neurons exhibiting nicotinamide-adenine dinucleotide phosphate-diaphorase activity. In the submucosal plexus of guinea-pig ileum, labelled neurons included some neuropeptide-Y-containing neurons but none with vasoactive intestinal polypeptide. We conclude that the Fas ligand is expressed by a large subset of enteric neurons and may provide the basis for cytotoxic neuroimmune interactions in the intestines.
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  • 44
    ISSN: 1432-0878
    Keywords: Key words: Pineal gland ; Laminin ; Cadherin ; Synaptophysin ; BrdU ; Immunohistochemistry ; Embryology ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Epithelial remodeling in the rat pineal during fetal development was immunohistochemically analyzed by using antibodies for laminin and cadherin as molecular markers of basal lamina and intercellular junctions, respectively. The proliferation and differentiation of pinealocytes were also investigated in relation to the advance of epithelial remodeling. The pineal anlage of embryonic day 16 is completely covered by basal lamina immunolabeled for laminin. After embryonic day 17, local dissolution of the basal lamina occurs on the epithelial folds, which develop predominantly in the rostral pineal wall. Some pineal cells migrate through these interruptions and form cellular aggregations outside the basal lamina. Cadherin immunostaining reveals focal dissolution of intercellular junctions in epithelial regions protruding into the pineal lumen. Dissolution of the basal lamina and intercellular junctions accompanied by cellular migration into the stromal tissue or into the pineal lumen continues until birth. The distribution of mitotic cells immunolabeled for BrdU is homogeneous throughout the organ during the fetal period, whereas that of differentiating pinealocytes immunoreactive for synaptophysin shows striking regional heterogeneity in close correlation with the remodeling of the pineal epithelium. The migrating cell populations located either outside the basal lamina or inside the pineal lumen are more liable to become synaptophysin-positive than the rest of the epithelium. These results suggest that epithelial remodeling in the fetal pineal is induced, at least in part, by epithelial infolding and that this remodeling promotes the differentiation of pinealocytes.
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  • 45
    ISSN: 1432-0878
    Keywords: Key words: Arginine ; Analogues ; Immunocytochemistry ; Nitric oxide ; Pituitary ; Retina ; Transport ; Rat (Wistar) ; Rabbit
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Nitric oxide is formed by the action of nitric oxide synthase upon l-arginine. The efficacy of some exogenously applied arginine analogues in inhibiting nitric oxide synthase and thus nitrergic transmission indicates that neurons producing nitric oxide may possess an arginine transport system. To investigate whether arginine analogues are preferentially transported into nitric oxide-utilising cells or into cells making other neurochemicals, we have raised highly specific antisera against a number of arginine analogues including NG-methyl arginine, d-arginine, NGnitro-l-arginine, NG-nitro-l-arginine methyl ester and canavanine. Retinae were incubated in physiological media containing these analogues and rats were given intraperitoneal injections of the analogues to study the pituitary. Immunocytochemistry and NADPH-diaphorase histochemistry revealed that many of these analogues could be transported preferentially, but not exclusively, into nitric oxide-generating cells. However, some nitric oxide-producing cells apparently lacked the ability to take up some arginine analogues. We conclude that nitric oxide-generating cells in the retina and pituitary possess one or more arginine transporters. Other subsets of neurons that use GABA or glutamate as a neurotransmitter may also accumulate arginine analogues, possibly as a substrate for formation of these neurochemicals.
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  • 46
    ISSN: 1432-0878
    Keywords: Key words: Heat-shock protein 60 (hsp60) ; Chaperonin ; Mitochondria ; Gastrointestinal tract ; Epithelium ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Immunohistochemical studies on various parts of the rat gastrointestinal tract by means of an antibody against the mitochondrial chaperonin, heat-shock protein 60 (hsp60), has revealed a cell-specific distribution pattern. The active form of hsp60 is a heptameric complex that is involved in the import and refolding of nuclear-encoded proteins destined for the mitochondrial matrix. This chaperonin is detectable in highly replicating cells, e.g., keratinizing cells of the esophagus and short-living epithelial cells of the intestine. In the stomach, some of the oxyntic cells contain hsp60-positive mitochondria lying near intracellular canaliculi. Neuronal cells of the enteric nervous system present intense positive staining for hsp60 in some areas. All other non-epithelial cells of the digestive tract show weak or no hsp60 immunoreactivity. The presence of hsp60 in mitochondria seems to reflect two different forms of mitochondrial renewal: (1) total reformation of mitochondria and their content after mitotic division and (2) regeneration of these organelles following high activity, e.g., ATP synthesis.
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  • 47
    ISSN: 1432-0878
    Keywords: Key words: Microglia ; Monocytes ; Macrophages ; Astroglia ; Epithelial cells ; Fibroblasts ; Ramification ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Microglial cells are able to switch between an ”active” amoeboid and a ramified ”resting” morphology during development and after experiencing lesions. We have previously shown that in vitro microglial morphology is controlled by their cellular environment, i.e. cells become ramified in astrocyte coculture but amoeboid on monolayers of fibroblasts. In the present study we have extended the analysis of the control of macrophage morphology by maintaining macrophages of different origins in coculture with different epithelial or mesenchymal cells and their conditioned media. Microglia, monocytes and spleen macrophages seeded onto monolayers of astrocytes, kidney epithelia or hepatoma cells developed the ramified morphology but remained amoeboid in fibroblast coculture. Ramification was also induced by media conditioned by these cells as well as by phorbolic esters, i.e. activators of protein kinase C. In double coculture assays, even small numbers of fibroblasts were able to override the ”epithelial” influence. Likewise, microglia remained amoeboid, when incubated on several constituents of the extracellular matrix. These results indicate that macrophage ramification is an active process initiated by diffusible factors secreted by various epithelial cells, possibly acting upon a protein-kinase-C-related receptor. We interprete the modification of macrophage morphology as a functional adaptation to the surrounding type of tissue that is enforced by its constituent cells. Thus, the specific morphologies of microglia, hepatic von Kupffer’s cells or peritubular kidney macrophages could be explained by similar epithelium–macrophage interaction.
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  • 48
    ISSN: 1432-0878
    Keywords: Key words: FGF-2 ; FGF receptor-1 ; Autonomic nervous system ; Sympathetic preganglionic neurons ; Spinal cord ; Development ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The development of the nervous system appears to be under the control of multiple growth factors, neurotrophins and cytokines, which may be expressed either continuously or transiently throughout defined stages of cellular generation, proliferation or differentiation. Fibroblast growth factor (FGF) cytokines and their receptors are abundantly expressed in the embryonic nervous system but their localization at autonomic levels in the fetal spinal cord has not yet been detailed. Immunoreactivity to FGF-2, probably the best characterized member of the FGF family (FGF-1 to FGF-10) and of one of its high affinity receptors, FGFR-1, was found in autonomic neurons at embryonic day E14, the peak day of generation and proliferation in the common ventral motoneuron pool. It was also continuously present throughout the investigated subsequent stages (E15 to postnatal day P30). Immunogold electron microscopy revealed the cytoplasmic localization of FGF-2 and FGFR-1 in intermediolateral neurons, the major group of sympathetic preganglionic neurons in the spinal cord. In these neurons, immunocytochemistry from E14 onwards showed the co-distribution of both markers at the period of axonal outgrowth to peripheral targets, e.g. the adrenal medulla. Our findings suggest autocrine and/or paracrine actions of FGF-2 for sympathetic preganglionic development but do not support its role as a target-derived neurotrophic factor for autonomic neuron development.
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  • 49
    ISSN: 1432-0878
    Keywords: Key words: 2-Acetylaminofluorene ; Selective ductal cytotoxicity ; Submandibular gland ; Regenerative repair ; Immunohistochemistry ; Systemic effects ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Salivary glands of rodents are rarely affected with spontaneous and induced malignancies. This may be linked with low physiologic cell renewal and the infrequency of cytolethal actions by xenobiotics. The genotoxic 2-acetylaminofluorene, carcinogenic for other organs, causes acute damage in salivary ducts. In the submandibular glands the damage is limited to the granulated convoluted tubules. They produce and release regulatory peptides including epidermal growth factor and nerve growth factor. The partial chemical sialoadenectomy is repaired by sequential cell proliferation in the basal cell layer of interlobular ducts and in dilated intralobular ducts (day 4 and 6), in intermediate duct-like structures (day 6 and 8), and lastly in acini (day 8 and 12). This is associated with a transient loss of structural characteristics of striated ducts and acini (up to day 6) and of the immunoreactivity for S-100 protein (up to day 4). Actin immunoreactivity at the acinar base is increased from day 6 to 20. Analogous to the late postnatal differentiation of the granulated convoluted tubules, their structural characteristics and immunoreactivity for epidermal growth factor do not recover within 20 days. The acute lesion of the endocrine ductal segment is suggested to be causally involved with other systemic effects following treatment with 2-acetylaminofluorene. First, hypophagia with loss of body, liver and thymus weight may result from disturbed saliva production. Second, previous studies have shown a mitotic burst of the biliary epithelium and bloodborne lymphocyte-stimulating activities. Either effect could be brought about by regulatory peptides (see above), probably after elevated circulatory release from necrotic granulated convoluted tubules.
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  • 50
    ISSN: 1432-0878
    Keywords: Key words: Liver ; Subcellular fractions ; Immunohistochemistry ; Electron microscopy ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. This study, utilizing rats subjected to two-thirds partial hepatectomy or sham operation, was designed (1) to investigate the content of basic fibroblast growth factor (bFGF) in the subcellular fractions of regenerating and sham-operated rat livers by immunoblot experiments and enzyme-linked immunosorbent assay (ELISA), (2) to show that bFGF immunoreactivity and proliferating cell nuclear antigen (PCNA) immunoreactivity are markers for hepatocellular mitosis before and after partial hepatectomy, and (3) to observe the location and fine structure of the bFGF immunoreaction within the regenerating liver with special attention to bFGF immunoreactivity in the nuclei of regenerating hepatocytes. Immunoblot experiments and ELISA showed a transient increase in high-molecular-weight forms of bFGF in the nuclear subcellular fraction of regenerating liver 48 h after partial hepatectomy. By light microscopy, bFGF and PCNA immunoreactivities were detected in the nuclei of regenerating hepatocytes. Electron microscopy demonstrated bFGF-like immunoreactivity mainly in the nuclear euchromatin and rarely in the heterochromatin or nucleoli of regenerating hepatocytes. The transient increase in high-molecular-weight forms of bFGF in the nuclear euchromatin of regenerating hepatocytes, together with the concomitant expression of PCNA in the regenerating liver, suggests an important role of the high-molecular-weight forms of bFGF in hepatocyte proliferation and/or mitosis, although authentic bFGF with a molecular form of 18 kDa is not considered to be involved in hepatic regeneration.
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  • 51
    ISSN: 1432-0878
    Keywords: Key words: Collagen ; Combined non-radioactive in-situ hybridization and immunohistochemistry ; Gastric ulcer ; Immunohistochemistry ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The restoration of gastric tissue after ulceration involves cellular and matrix components. Our aim was to investigate the kinetics of collagen expression and cellular proliferation in an animal model of gastric ulcer. To demonstrate the expression of type I and IV collagen mRNAs by proliferating cells, a method combining in-situ hybridization and immunohistochemistry was devised. In order to avoid the disadvantages of radioisotopes, digoxigenin-labeled RNA-riboprobes were utilized and combined with single-step immunohistochemistry. This method proved sensitive enough to detect type I and IV procollagen mRNA transcripts in the submucosal area beneath the ulcer crater or adjacent to the ulcer rim. In addition, a subset of cells transcribing either procollagen type I or IV RNA was concomitantly positive for proliferating cell nuclear antigen by immunohistochemistry. Focal proliferation of cells simultaneously expressing extracellular matrix components may therefore occur in the gastric submucosa after ulceration, starting as soon as 3 days after the insult and continuing for several weeks. The devised method of combined in-situ hybridization and immunohistochemistry can be used with standard paraffin-embedded tissues, yields results within 2 days, and avoids radioisotopes.
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  • 52
    ISSN: 1432-0878
    Keywords: Key words: Thalamus ; Intercellular junctions ; Synapse ; Synaptic glomeruli ; Agranular endoplasmic reticulum ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Filamentous contacts are non-synaptic interneuronal junctions characteristic of thalamic relay nuclei. Symmetrical filamentous contacts occur between two dendrites, two somata or a dendrite and a soma; asymmetrical filamentous contacts occur between axon terminals and dendrites, or occasionally somata, chiefly between the large specific afferent axon terminals of the synaptic glomeruli and the shafts of relay cell dendrites. Both are arranged as extensive net-like (reticular) specializations. The strands of the network enclose fenestrae of variable shape and size and, in perpendicular thin sections, appear as stretches of slightly widened intercellular space containing an electron-dense material and bounded by plasma membranes, the cytoplasmic surfaces of which are coated by electron-dense material into which microfilaments appear to insert. The lamina of cytoplasmic material in dendrites and somata is thicker than that in axon terminals and contains distinct electron-dense sub-units. Regular synaptic junctions may be situated like islands within the territory of an asymmetrical filamentous contact, and small spot-like close membrane appositions resembling gap junctions are occasionally seen in the fenestrae adjacent to the strands of both varieties of contact. Bundles of neurofilaments running in different directions, but in a plane parallel to the plasma membrane, are prominent on either side of the symmetrical filamentous contact and on the dendritic side of the asymmetrical variety. The agranular reticulum also exhibits differences between the contact types. Because of their highly specialized ultrastructure and specific distribution, filamentous contacts probably do not serve a purely adhesive function. Their possible role in the establishment and maintenance of orderly connections between cells is discussed but not favoured. Filamentous contacts probably mediate some form of intercellular communication, possibly involving gap junctions.
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  • 53
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    Cell & tissue research 288 (1997), S. 11-23 
    ISSN: 1432-0878
    Keywords: Key words: Nitric oxide synthase (NOS) ; Vasoactive intestinal peptide (VIP) ; Pituitary-adenylyl-cyclase-activating peptide (PACAP) ; α-Internexin ; Peripherin ; Immunohistochemistry ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The presence of nitric oxide synthase (the enzyme which synthesizes nitric oxide), vasoactive intestinal peptide (VIP) and pituitary adenylyl-cyclase-activating peptide (PACAP) in the rat enteric nervous system between term (E18) and 90 days postpartum (P90) was investigated by immunohistochemistry. Neuronal maturity was ascertained by emanining for the presence of two novel neuron intermediate filament proteins: α-internexin, which is transiently expressed in developing neurons, and peripherin, which is expressed in differentiating as well as in mature neurons. The alimentary canal of the foetus at E18, unfed newborn, suckling, weaning and adult rats was studied. Throughout the rat gut the myenteric neurons were recognized by the time the foetal stage was reached and the submucous neurons by the suckling period. α-Internexin was present at E18; its levels increased during ageing and were markedly reduced at P30. Peripherin was first detected at birth; its levels increased with ageing and pratically almost all of the neurons and nerve fibres were labelled from P30. Nitric oxide synthase was present at E18 and the number of labelled neurons gradually increased with ageing, while both VIP and PACAP could be detected by the end of the suckling period in an equal number of neurons to that in adulthood. These data indicate that in the rat gut the onset of the mature neuronal phenotype is not yet achieved at birth and that the neurochemical differentiation is accomplished during the first month of postnatal life.
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  • 54
    ISSN: 1432-0878
    Keywords: Key words: Ontogenesis ; Perineuronal nets ; Extracellular matrix ; Proteoglycans ; Brain ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The extracellular matrix is involved in various morphogenetic processes which are accompanied by changes in its physicochemical properties and spatial organization. In the adult brain it contributes to cellular communication and the regulation of neuronal activity. The present study deals with the postnatal appearance and transformation into adult distribution patterns of extracellular matrix components related to chondroitin-sulphate proteoglycans (CSPGs) in the rat brain. The differential accumulation of these components in neuropil and in perineuronal nets (PNs) enriched in certain regions was examined in 0-, 7-, 14-, 21- and 35-day-old rats and adult animals using the N-acetylgalactosamine-binding Wisteria floribunda agglutinin (WFA) and immunocytochemical detection of CSPGs. The lectin stained the olfactory-bulb glomerular layer and layer Ia of piriform and entorhinal cortex already in newborn animals. On postnatal day 7 diffuse neuropil staining was additionally found in certain subcortical nuclei and in deep neocortical layers. The first sharply contoured PNs were detected at this age in the brain stem, indicating the more advanced maturation of matrix components in subcortical regions. CSPG immunoreactivity yielded staining patterns largely identical to WFA-binding patterns but appeared only between postnatal day 14 and 21. The adult-like stage was revealed with both methods between 21 and 35 days after birth. The results provide further evidence that the accumulation of certain CSPGs in the extracellular space is spatiotemporally related to distinct patterns of neuronal activity at the regional and cellular level.
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  • 55
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    Cell & tissue research 288 (1997), S. 95-99 
    ISSN: 1432-0878
    Keywords: Key words: Distal colon ; Aldosterone adaptation ; Sodium absorption ; Intestinal development ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The distal colon of suckling (14-day-old) and weanling (27-day-old) rats absorbs Na+ via channel-mediated, electrogenic amiloride-sensitive Na+ transport which disappears after weaning. This transport pathway is induced by aldosterone in superficial cells of colonic epithelium. The purpose of the present study was to correlate the changes in distal colon function with changes in the apical and basolateral cell membrane surface areas of superficial enterocytes. The basolateral but not apical membrane surface density (i.e. surface areas of the basolateral and apical membranes of superficial enterocytes per unit volume of superficial enterocytes) was found to increase between postnatal day 14 and 27 (i.e. during the weaning period). The plasma concentrations of aldosterone were very high during this period and electrogenic amiloride-sensitive Na+ transport and Na, K-ATPase activity were increased significantly. High dietary salt intake during the weaning period prevented the developmental increase of basolateral membrane surface density, inhibited electrogenic amiloride-sensitive Na+ transport and significantly depressed plasma aldosterone levels and Na, K-ATPase activity. Apical cell membrane surface density did not change significantly after a sodium load. Thus, high plasma concentrations of aldosterone and/or high colonic Na+ transport during weaning exert an important and selective effect on the basolateral membrane and are responsible for the developmental changes of this cell membrane surface during weaning. Concomitant changes in the morphology of superficial colonic enterocytes and in colonic Na+ transport suggest that the structure of the immature epithelium is related to its function.
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  • 56
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    Cell & tissue research 288 (1997), S. 225-238 
    ISSN: 1432-0878
    Keywords: Key words: Cerebral ischemia ; Plasticity ; GAP-43 ; Hippocampus ; mRNA ; Immunohistochemistry ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Neuroanatomical methods have been used to study selective vulnerability after global brain ischemia. A consistent pattern of ischemic neuronal damage is found in the rodent hippocampus with loss of CA1 neurons and of some cells in the hilus of the dentate gyrus. Very little is known about plastic changes that would be expected in ischemia-resistant areas such as CA3 neurons and granule cells. Neuronal plasticity after lesions may be indicated by changes in labeling with antibodies to the growth-associated protein 43 (GAP-43). Expression of GAP-43 as a marker for neuronal plasticity was studied here in the hippocampus after global brain ischemia. Halothane-anesthetized rats were subjected to 20 min of transient forebrain ischemia using four-vessel occlusion. In situ hybridization was used to study GAP-43 mRNA at 1, 3, 6, and 12 h and at 1, 3, and 7 days after ischemia. Immunostaining was carried out with two different antibodies to GAP-43 in brains which were perfusion-fixed after 1, 2, 4, and 7/8 days. In the control hippocampus, GAP-43 mRNA was localized to CA1–CA3 and the hilus. Moderate increases in cellular signals were seen in hilar cells and granule cells early after ischemia, and some changes occurred in CA3 at late stages. Hybridization was lost in CA1 due to cell death. With immunostaining, GAP-43 was not seen in the cytoplasm of neurons, whereas dense labeling occurred in a differentiated pattern in the axonal and dendritic layers. At 1 day after ischemia, neurons in the hilus of the dentate gyrus and in the stratum pyramidale and lucidum of CA3 showed strong cytoplasmic labeling for GAP-43. Few cells were labeled in these regions at 2 days, and none at later stages. Pyramidal cells in CA1 and CA3 areas and granule cells were never labeled. These studies demonstrate a transient expression of GAP-43 mRNA and protein in a subset of vulnerable neurons after transient brain ischemia. The cytoplasmic localization in hilar neurons could be due to increased synthesis of GAP-43 or to changes in axoplasmic transport. It is suggested that axonal damage occurs in hilar cells which stimulates GAP-43 expression. The increased production of trophic factors after ischemia in granule cells could also cause plastic changes in hilar cells. Since hilar neurons are in a strategic position to control the excitability of the dentate area, increased expression of GAP-43 may indicate an important pathophysiological process. In seizure experiments, strong expression of GAP-43 mRNA in granule cells was associated with abnormal mossy fiber sprouting and development of chronic epilepsy. The relevance of the minor GAP-43 mRNA upregulation after ischemia must be considered. The changes in CA3 neurons at several days after ischemia might represent a plastic response to a loss of CA1 neurons.
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  • 57
    ISSN: 1432-0878
    Keywords: Key words: Microtubules ; MAP2 ; Tau ; Cytoskeleton ; RT-PCR ; Oligodendrocyte ; Brain ; Cell culture ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Oligodendrocytes in culture are characterized by large membranous sheets containing an elaborate network of microtubules. Microtubule-associated proteins (MAPs) participate in microtubule stability and the regulation of the cellular architecture. We have investigated the expression of two major groups of MAPs, MAP2 and tau, in cultured rat brain oligodendrocytes. Alternatively spliced isoforms of mRNAs encoding MAP2 and tau were assessed by means of reverse transcription and polymerase chain reaction using a newly designed set of MAP2- and tau-specific primers. The data were compared with data obtained with cultures of rat brain astrocytes and rat cerebral neurons, and adult rat brain. The results show that oligodendrocytes, similarly to neurons, express mainly MAP2c transcripts containing three microtubule-binding repeats. They also contain small amounts of MAP2b mRNA. Six low molecular weight tau isoforms, namely tau 1–6, have been described in the brain (Goedert et al. 1991). The major isoform of tau mRNA in oligodendrocytes was found to be tau 1, which represents a marker typical for immature neurons. Tau 2 and tau 4 isoforms were also detected, albeit at a very low level. Immunoblot analysis of oligodendroglia cell extracts confirmed the presence of tau protein. It migrates as a single polypeptide with an apparent molecular weight of approximately 55 kDa. In addition, oligodendrocytes express MAP2c protein, which migrates as a close double band with an apparent molecular weight around 70 kDa. Indirect immunofluorescence staining indicated that tau and MAP2 immunoreactivity was expressed in oligodendrocytes of immature and mature morphologies in the cell somata and cellular processes. Tau was particularly found in the end of the cellular extensions, and both proteins exhibited a distribution similar to myelin basic protein. Thus, oligodendroglia, like neuronal cells, contain microtubule-associated proteins, mainly MAP2c and the tau 1 isoform, although at a much lower level. The presence of these MAPs in myelin-forming cells further points to the functional significance of the cytoskeleton during oligodendrocyte differentiation, process outgrowth, and myelin formation.
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  • 58
    ISSN: 1432-0878
    Keywords: Key words: Adrenoceptors ; Diurnal rhythms ; Regulation ; In situ hybridization ; Pineal gland ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The rat pinealocyte is stimulated by norepinephrine, which is released from sympathetic nerve fibers innervating the gland. Norepinephrine binds to β1-adrenoceptors situated on the pinealocyte cell membrane. Ligand binding to these receptors exhibits a diurnal rhythm, with the largest number occurring in the late part of the light phase when the release of norepinephrine is minimal. By using in situ hybridization with a cDNA antisense oligonucleotide probe recognizing mRNA encoding the rat β1-adrenoceptor, we have demonstrated a stronger hybridization signal in the rat pineal gland than in other brain regions. Cells containing β1-mRNA are located in the superficial pineal gland, the deep pineal gland, and the pineal stalk. However, the number of receptors varies considerably between the individual pinealocytes. The β1-mRNA in situ hybridization signal for mRNA encoding the β1-adrenoceptor of the rat pineal has been quantified over a 24-h period; the strongest signal is found at mid-dark and the weakest signal at mid-light, indicating that the transcriptional regulation of β1-mRNA synthesis in the rat pineal is diurnal. In addition, maximal receptor protein expression occurs late in the light phase in the rat pineal and is thus considerably delayed compared with its peak mRNA synthesis. This lag time indicates that the β1-receptor is regulated at the translational or post-translational level. Removal of the sympathetic input to the pineal gland by superior cervical ganglionectomy decreases the β1-mRNA signal in the gland.
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  • 59
    ISSN: 1432-0878
    Keywords: Key words: Kidney (proximal tubule) ; Apical tubule ; Endosome ; Ultrastructure ; Endocytosis ; Membrane recycling ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The apical cytoplasm of several absorbing epithelia contains well-developed apical tubules (AT) which contribute to membrane recycling from endocytic vacuoles to the apical cell membrane. In this study, we examined three-dimensional structures of the AT in rat kidney proximal tubule cells by transmission and scanning electron microscopy. In thin sections, the AT appeared as straight tubules with a rather constant diameter (70–90 nm), but others were curved and, occasionally, branching. No AT were labeled with the marker for the external cell surface (ruthenium red) or exhibited histochemical enzyme activity for lysosomal hydrolase (acid phosphatase). After intravenous injection of horseradish peroxidase, it was absorbed in the kidney proximal tubule cells and the AT were labeled with HRP reaction products. Stereo-viewing of the labeled AT in thick sections revealed that they formed an interconnected tubular network. Scanning electron microscopy allowed a three-dimensional view of the AT, in which a network of branching and anastomosing tubules was revealed. These observations indicate that the AT are intracellular endosomal compartments which form an extensive tubular network in the apical cytoplasm. The possibility that this apical tubular network serves as a large membrane store for membrane recycling is discussed.
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  • 60
    ISSN: 1432-0878
    Keywords: Key words: Endothelins ; Endothelin receptors ; Adrenal gland ; Steroid secretion ; Catecholamine secretion ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Endothelins (ET) are a family of vasoactive peptides that act via two subtypes of receptors, named ETA and ETB. ET-1 binds to both ETA and ETB, whereas the isopeptide ET-3 preferentially binds to ETB. The localization of ETA and ETB receptors in the rat adrenal gland and their involvement in the adrenal secretagogue effect of ETs has been studied in vitro. Autoradiographic assessment of the selective displacement of [125I]ET-1, [125I]ET-3 and [125I]BQ-3020 (an ETB agonist) by BQ-123 or BQ-788 (specific antagonists of ETA and ETB, respectively) indicates that the zona glomerulosa and adrenal medulla possess both ETA and ETB, whereas the zona fasciculata/reticularis is exclusively provided with ETB. ET-1, ET-3 and BQ-3020 enhance aldosterone and corticosterone secretion by dispersed cells of the zona glomerulosa and zona fasciculata/reticularis, respectively. BQ-123 does not affect the secretagogue action of these three agonists, whereas BQ-788 completely annuls it. ET-1 induces a marked rise in catecholamine release by fragments of the adrenal medulla, and both BQ-123 and BQ-788 partially reverse this effect. ET-3 and BQ-3020 elicit a catecholamine release that is less intense than that produced by ET-1; this response is unaffected by BQ-123 and abolished by BQ-788. Thus, in the rat, the corticosteroid secretagogue effect of ETs seems to be exclusively mediated by the ETB receptor subtype, and the catecholamine secretagogue action by both ETA and ETB. The functional relevance of ETA receptors present in the zona glomerulosa remains to be investigated.
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  • 61
    ISSN: 1432-0878
    Keywords: Key words: Calcification ; Concretions ; Collagen ; Pineal gland ; Aging ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The relationship between collagen fibrils and calcified concretions exclusively appearing in the pineal gland of adult/aging rats has been investigated. Deposits of lanthanum, which replace calcium ions are distributed along collagen fibrils with a repeating period of about 70 nm. Calcium has been detected histochemically between collagen bundles surrounding extracellular concretions by means of the pyroantimonate method and by X-ray microanalysis. It is associated with phosphorus. The data presented here suggest that collagen fibrils are involved in the genesis and growth of extracellular concretions located in the connective tissue surrounding the pineal gland of aging rats.
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  • 62
    ISSN: 1432-0878
    Keywords: Key words: Secretoneurin ; Pineal gland ; Neuropeptide immunocytochemistry ; Sympathetic nervous system ; Superior cervical ganglion ; Pinealocytes ; Cell culture ; Rat (Wistar) ; Syrian hamster ; Mesocricetus auratus ; Siberian hamster ; Phodopus sungorus (Rodentia)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The pineal gland of mammals is richly innervated by nerve fibres containing various peptides of central or sympathetic origin. In this study, we have investigated the presence, regulation and effect of secretoneurin (SN), a 33-amino-acid peptide discovered recently in the central and sympathetic nervous systems. The rat pineal gland contains about 34 fmol SN/pineal, although only 20% of the secretoneurin immunoreactivity (SN-IR) is the free peptide. SN-IR is localised in only a few nerve fibres found in the parenchyma of the gland and along the pineal stalk. In the rat pineal, SN-IR shows no midday/midnight differences but is significantly decreased (55%–65%) after superior cervical ganglionectomy or exposure to constant light. These observations suggest that, in the rat pineal, SN-IR originates partly from central areas and partly from the superior cervical ganglia. The pineal of the Syrian hamster contains much more SN-IR: 120 fmol SN-IR/pineal (about 20% being the free peptide). SN-IR is found both in nerve fibres and pineal cells. Some of the fibres run along the pineal stalk, indicating that they are of central origin, although a sympathetic origin cannot be ruled out. Pineal SN-IR is below detectable levels in the Siberian hamster. In cultured rat pinealocytes, SN significantly inhibits serotonin release and, to a lower extent, melatonin release.
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  • 63
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    Cell & tissue research 288 (1997), S. 449-469 
    ISSN: 1432-0878
    Keywords: Key words: Neuronal ultrastructure ; Catecholamines ; Dopamine ; Immunocytochemistry ; Connections ; Quantitation ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Interrelations of tyrosine-hydroxylase-immunoreactive afferent fibres with neuronal elements were studied in central, basal and intercalated nuclei of the rat amygdaloid complex. Comparison with dopamine-β-hydroxylase-immunoreacted and phenylethanolamine-N-methyltransferase-immunoreacted parallel sections indicated that the tyrosine-hydroxylase immunoreaction labelled preferentially dopaminergic axons. At the elec- tron-microscopic level, the majority of tyrosine-hydroxylase-immunoreactive axons possessed small boutons containing small clear vesicles and contacting dendrites, spines or somata of amygdala neurons, forming mostly symmetric synapses. They were often directly apposed to or in the vicinity of unlabelled terminals synapsing on the same structure. Synaptic density was highest in the central lateral part of the central nucleus. In the central and basal nuclei labelled axons synapsed preferentially on small dendrites and dendritic spines, and on somata of a few neurons. A detailed study of the neuronal ultrastructure showed that innervated somata possessed the differential characteristics displayed by the predominant neuron types in the medial and central lateral central nucleus and resembled the typical projection neurons in the basal nuclei. In the paracapsular intercalated cell groups the majority of neurons possessed intense perisomatic innervation by immunoreactive terminals. The results suggest that tyrosine-hydroxylase-immunoreactive, predominantly dopaminergic amygdaloid afferent fibres preferentially modulate the effect of extrinsic inputs into neurons of the central and basal nuclei, while a nonselective regulation is exerted upon the output of paracapsular intercalated neurons. It is suggested that this innervation pattern may be important for the coordinated integration of extrinsic and intraamygdaloid connections and thus for balanced output of the structure.
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  • 64
    ISSN: 1432-0878
    Keywords: Key words: raf ; Dorsal root ganglion ; Neurotrophin ; trk ; Dorsal horn ; Axonal transport ; Protein kinase ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. We have used immunocytochemistry to examine the distribution of raf protein kinases in sensory neurons of the adult rat. In lumbar and trigeminal sensory ganglia, cells of all size ranges appeared to be raf immunoreactive and this was confirmed by double labeling using subpopulation specific markers. Almost all cells labeled with Griffonia simplicifolia IB4 (a small cell marker) or immunostained by using a large cell marker (RT97) showed raf immunoreactivity. These two markers label cells known to differ in their expression of neurotrophin receptors (trk). Thus raf kinases are not confined to cells expressing only certain trk subtypes. In the dorsal horn of the spinal cord, raf immunoreactivity was present in scattered neurons. However, sensory axons identified by IB4 histochemistry were devoid of raf immunostaining. Lectin-labeled nerve fibers in the cornea, lower lip and glans penis were also not immunoreactive. Ligation of the sciatic nerve did not produce any accumulation of raf immunoreactivity, confirming that raf kinases are not axonally transported to the peripheral processes of sensory neurons. Surgical dissection of the sciatic nerve caused the normal homogeneous cytoplasmic raf immunoreactivity to be replaced in some cells by a staining concentrated predominantly under the plasma membrane. One possibility is that this represents activation of raf in these cells.
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  • 65
    ISSN: 1432-0878
    Keywords: Key words: Prolactin cells ; Anterior pituitary (inner and outer zones) ; Secretory activity ; Ultrastructural immunohistochemistry ; Standard and ultrastructural reverse hemolytic plaque assays ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The aim of this study was to compare the ultrastructure of prolactin cells in the inner and outer zones of the male rat anterior pituitary, and to relate their morphological features to their secretory activity, by means of standard and ultrastructural reverse hemolytic plaque assays (RHPA). The immuno-ultrastructural study showed that in the inner pituitary small-granulated cells represented 52% of the prolactin cells, there being only 5% with large granules, whereas the prolactin cells with large granules accounted for 52% in the outer zone, with only 7% being small-granulated. Percentages of cells with intermediate-sized granules were 43% and 41%, respectively. Analysis of RHPA data revealed that, under basal conditions, prolactin cells secreted more actively in the inner zone than in the outer zone. Stimulation with thyrotropin-releasing hormone or KCl treatment increased the percentage of secretors and the sizes of hemolytic plaques in both zones. However, in response to thyrotropin-releasing hormone, the increase in number of secretors was always higher in the outer zone, whereas the enlargement of plaque sizes was greater for the ”inner” cells. These findings are in favor of the small-granulated cells, which predominate in the inner zone, being in a stage of active secretion and responsiveness.
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  • 66
    ISSN: 1432-0878
    Keywords: Key words: CREB (cyclic AMP response element binding protein) ; Melatonin ; Pituitary adenylate cyclase-activating polypetide (PACAP) ; Pinealocytes ; Vasoactive intestinal peptide (VIP) ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. We investigated whether vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP), which stimulate melatonin biosynthesis in the mammalian pineal organ, cause phosphorylation of the cyclic AMP response element binding protein (CREB) in rat pinealocytes. Dispersed cells were treated with varying concentrations of VIP and PACAP for 10 to up to 240 min and then immunocytochemically analyzed with an antibody against phosphorylated CREB (pCREB). The experiments showed a dose- and time-dependent induction of pCREB immunoreactivity in the nuclei of subpopulations of pinealocytes identified by the S-antigen immunoreaction. Stimulation with VIP elicited pCREB immunoreaction in approximately 50–65% of the S-antigen immunoreactive pinealocytes. The number of PACAP-responsive pinealocytes was often smaller and more variable. Maximal responses to both neuropeptides were seen after 30 min. pCREB immunoreaction gradually declined within 2 h and could not be induced again by an additional stimulation. In contrast, norepinephrine (NE) elicited pCREB immunoreaction in more than 95% of the pinealocytes, and this response lasted as long as 300 min. Treatment of pinealocytes with forskolin or KCl induced pCREB immunoreaction in the vast majority of pinealocytes, showing that in principle elevation of the intracellular concentrations of both cAMP and calcium can cause the response. Immunoblotting analyses confirmed that the immunoreaction elicited by VIP, PACAP and NE is largely due to phosphorylation of a 42-kDa protein corresponding to CREB, but reflects to a minor extent also phosphorylation of two smaller proteins presumably related to ATF-1. Immunocytochemical and immunochemical investigations with an antibody against total CREB showed that stimulation with VIP, PACAP and NE did not affect the level of CREB. All findings indicate that the stimulatory effects of VIP and PACAP on rat pinealocytes involve phosphorylation of transcription factors of the CREB family as holds also true for NE. However, VIP and PACAP affected only subpopulations of pinealocytes and the reponses lasted for a shorter period of time than those to NE. This conforms to previous results showing that both neuropeptides are also less effective than NE in stimulating the melatonin biosynthesis in the rat pineal organ.
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  • 67
    ISSN: 1432-0878
    Keywords: Key words: Hepatic stellate cells ; Endothelin-1 ; Collagen gel ; Liver ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Hepatic stellate cells become activated and aquire contractility on being cultured. In order to characterize the morphology of contracted and relaxed stellate cells, we performed light- and electron-microscopic analyses of cultured stellate cells on collagen gels. Incubation of stellate cells with medium alone, 10 nM endothelin (ET)-1, or 1 mM N6,2′dibutyryladenosine 3′:5′-cyclic monophosphate (dBcAMP) for 48 h induced contraction of the underlying collagen gels to 83%, 57%, and 97%, respectively, of their original size. Stellate cells relaxed by dBcAMP exhibited a round cell body and extended several long thin cytoplasmic processes with several varicosities. Culture with ET-1 accelerated spreading of the stellate cells on collagen gels and decreased the number of processes. Each such flattened stellate cell attached itself to the underlying collagen matrix by bending its cell body. Collagen fibers around the cell were pulled toward the cell and stretched. Thus, the present study has revealed that ET-1-stimulated cultured stellate cells adduct associated collagen fibers by the retraction of cytoplasmic processes and the bending of their spread cell bodies.
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  • 68
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    Cell & tissue research 287 (1996), S. 187-192 
    ISSN: 1432-0878
    Keywords: Key words: Spleen ; Sinus endothelial cells ; Adherens junctions ; Tight junctions ; Stress fibers ; Actin filaments ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Junctions between the sinus endothelial cells of rat spleen were examined by electron microscopy, using both freeze-fracture and detergent-extraction techniques. Adherens and tight junctions were observed. Adherens junctions were the predominant junctional structures between endothelial cells and were located on basolateral and lateral surfaces. At the basolateral adherens junctions, actin filaments were associated with the junctional membranes and were continuous with the actin filaments in stress fibers. Cross-bridges were present in the interspaces of the adherens junctions and spacing of the bridges was fairly regular. A form of tight junction, the macula occludens, was also observed between the endothelial cells, but it was not observed at every cellular apposition. Electron-dense material, adjoining the cytoplasmic surfaces of membranes in the tight junctions, separated the junctional membranes from masses of thin filaments. At basolateral tight junctions, the actin filaments were continuous with those in the stress fibers. Based on these observations, the two intercellular junctions were considered to play important roles in sinus functions.
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  • 69
    ISSN: 1432-0878
    Keywords: Key words: Nitric oxide ; Nitric oxide synthase ; Gastric mucosa ; Stomach ; Immunohistochemistry ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Nitric oxide has been implicated in the regulation of blood flow, mucosal integrity and mucus secretion in the gastric mucosa. An antiserum directed against the C-terminal hexadecapeptide of rat brain nitric oxide synthase (NOS) and monoclonal antibodies to the neuronal and endothelial forms of NOS were used to establish the location of isoforms of NOS in rat gastric glandular mucosa. Antibodies to the neuronal form of NOS reacted with a band of 160 kDa on immunoblots of brain and gastric mucosa, and the addition of the hexadecapeptide inhibited recognition by the antipeptide antiserum. The antibody to endothelial NOS detected a band of 140 kDa on protein blots of samples of intestinal mesentery and gastric mucosa. Immunohistochemistry using these antibodies demonstrated that material related to neuronal NOS was present in surface cells of the gastric mucosa, and showed a similar localization to intense NADPH diaphorase activity. The antibody to endothelial NOS did not stain the surface of the gastric mucosa but recognized blood vessels in the lower region of the gastric glands and in the sub-mucosa. This study suggests that nitric oxide might act both as an intra- and inter-cellular messenger to regulate mucus release, and that the NOS present in surface cells is related more closely to the neuronal than to the endothelial isoform.
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  • 70
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    Cell & tissue research 284 (1996), S. 347-354 
    ISSN: 1432-0878
    Keywords: Key words: Olfactory system ; Receptors ; membrane ; Gene expression ; Sensory cells ; Segregation ; laminar ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The laminar segregation of sensory neurons expressing a distinct receptor type was determined in tissue sections through the olfactory epithelium by in situ hybridization employing receptor-specific probes. Reactive cells were restricted to the mid-zone of the epithelium, the location of mature neurons. Detailed analyses revealed that neurons expressing a distinct receptor type were distributed in a characteristic manner throughout the layers of the neuronal zone, i.e. they were preferentially located in a particular laminar zone of the epithelium. Cells expressing different receptor types displayed different distribution patterns. In addition, sets of several reactive neurons within the same laminar zone were found to be arranged in an orderly fashion and were positioned at well-defined intervals. These results indicate that the localization of sensory neurons expressing a distinct receptor type is under stringent control leading to characteristic expression patterns.
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  • 71
    ISSN: 1432-0878
    Keywords: Key words: Retinoids ; Vaginal epithelium ; Differentiation ; Keratin ; Apoptosis ; Estradiol ; Progesterone ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. In rodents, the vaginal epithelium shows cyclic changes with an alternating pattern of keratinization under estrogen control and mucification under progesterone control. Retinoids are powerful regulators of cell differentiation, an excess of retinoids suppressing the keratinizing differentiation of keratinocytes. Here, we have examined the vaginal epithelium during the estrous cycle and compare the effects of retinoids on both types of hormonally induced differentiation, i.e. keratinization and mucification. All-trans retinoic acid was administred either by daily injections during the estrous cycle or by a single injection before the estrogen rise; these two protocols gave similar results. Retinoic acid suppressed estrogen-induced vaginal keratinization and cytokeratin K10 expression (a biochemical marker of terminal differentiation). Progesterone-induced mucification was not impaired; however, retinoic acid impeded mucous cell desquamation, suggesting an effect of retinoic acid on cell adhesiveness. Retinoic acid induced the appearance of apoptotic-like cells, as revealed by immunocytochemical staining of DNA fragmentation.
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  • 72
    ISSN: 1432-0878
    Keywords: Key words: Esophagus ; Nitric oxide ; Vasoactive intestinal peptide ; Vagus ; Enteric nervous system ; Confocal imaging ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Enteric co-innervation of motor endplates in the rat esophagus was studied with confocal laser scanning and electron microscopy. Enteric fibers were demonstrated with immunocytochemistry for nitric oxide synthase, vasoactive intestinal peptide or NADPH-diaphorase histochemistry. Vagal motor terminals were identified with calcitonin gene-related peptide (CGRP) immunocytochemistry. Teloglia was stained with immuno- cytochemistry for S100, and TRITC-tagged α-bungarotoxin was used to delineate endplate areas in immmunofluorescence preparations. Both confocal imaging and electron microscopy revealed intimate relationships between enteric and vagal terminals on the one hand, and enteric terminals and the sarcolemma on the other. In addition, electron microscopy could point out direct apposition of a significant proportion of enteric varicosities to vagal motor terminals without intervening teloglial processes. These morphological data are compatible with pre- and postsynaptic modulatory effects of enteric neurons on vagal neuromuscular transmission in striated esophageal muscle.
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  • 73
    ISSN: 1432-0878
    Keywords: Key words: Olfactory bulb ; Development ; Cholinergic transmission ; Nicotinic acetylcholine receptor ; α4 ; 1 Subunit mRNA ; In situ hybridization ; Digoxigenin ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. In addition to their role in signal transduction, nicotinic acetylcholine receptors have been shown in vi-tro to be involved in neuronal growth cone regulation during development. This idea is supported by recent histochemical findings showing that iso- and archicortical nicotinic α4–1 receptor mRNA expression precedes cholinergic fiber ingrowth. To test whether this also holds true for rhinencephalic parts of the telencephalon, we have studied the olfactory bulb by digoxigenin-mediated in situ hybridization, using an α4–1 isoform-specific riboprobe and an alkaline-phosphatase-based detection system. Development is characterized by early intense α4–1 mRNA expression (embryonic day 14), reaching a peak around postnatal day 2 when all olfactory bulb layers are invested with numerous α4–1 transcript-bearing neurons. Subsequently, the density of labeled neurons decreases to reach adult levels (postnatal day 120), where strongly labeled neurons remain in the mitral cell layer, outer external plexiform layer, and glomerular layer. The unifying pattern of iso-, archicortical, and rhinencephalic α4–1 mRNA expression is its early onset, i.e. preceding cholinergic innervation. This points to a possible role of nicotinic receptors regarding neuronal migration in all three regions.
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  • 74
    ISSN: 1432-0878
    Keywords: Key words: Glucose transporter ; GLUT1 ; Connexin 26 ; Gap junctions ; Placenta ; Syncytiotrophoblast ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Interhemal membrane in the rat placenta is composed of three trophoblastic layers and endothelial cells. GLUT1, an isoform of the facilitated-diffusion glucose transporter, is abundant in the cells of the placental barrier, i.e., syncytiotrophoblastic layers I and II. GLUT1 is localized at the plasma membranes of the maternal-blood side of syncytiotrophoblastic layer I, and of the fetal-blood side of syncytiotrophoblastic layer II. Double-immunofluorescence microscopy has shown that connexin 26 is present between these GLUT1-positive sites, i.e., between syncytiotrophoblastic layers I and II. Immunogold electron microscopy has revealed that connexin 26 is localized in the gap junctions connecting the two layers. Connexin 26 in these layers therefore makes them functionally a single syncytial layer for the transfer of small molecules such as glucose in the rat placental barrier. These results suggest that glucose transfer in the rat placental barrier is carried out as follows: GLUT1 is used for the entry of glucose into the cytoplasm of syncytiotrophoblastic layer I, connexin 26 for the transfer of glucose from syncytiotrophoblastic layer I to syncytiotrophoblastic layer II, and GLUT1 for the exit of glucose to the fetal circulation.
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  • 75
    ISSN: 1432-0878
    Keywords: Key words: Pineal gland ; MHC class II antigen ; Immunohistochemistry ; Electron-microscopic immunocytochemistry ; Dendritic cell ; Development ; Carbon tetrachloride ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Cells expressing major histocompatibility complex (MHC) class II (Ia) antigen have been examined during the development of rat pineals and in the pineal gland of adult rats treated with carbon tetrachloride. Cells positive for MHC class II are first detected in the pineal gland of the 7-day-old rat. These positive cells increase in number gradually during development, MHC class II immunoreactivity reaching adult levels at 4 weeks after birth. The MHC class II antigen is intensely labeled on the cell surface, and labeled cells are distributed throughout the organ, several positive cells being gathered into groups. The positive cells are small (7–12 μm in diameter), irregular in shape, and frequently exhibit one or more processes. At the electron-microscopic level, the cytoplasm of positive cells contains few organelles, variously sized empty vacuoles, and a few electron-dense lysosome-like structures. Pinealocytes with synaptic ribbons have been found adjacent to immunoreactive cells. Double-immunoperoxidase staining for MRC OX6, MRC OX42, and ED1 results in OX6−/ED1+/OX42+, OX6−/ED1−/OX42+, and OX6+/ED1−/OX42−cells. These findings suggest that OX6-positive cells in the pineal can be considered as peripheral dendritic cells. The number of cells expressing MHC class II (Ia) antigen significantly increases in the pineal gland of rats after treatment with carbon tetrachloride (P〈0.005). Our results indicate that at least some of the OX6-positive cells migrate into the gland from the circulation under these conditions.
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  • 76
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    Cell & tissue research 283 (1996), S. 305-311 
    ISSN: 1432-0878
    Keywords: Key words: Vagal afferent neurons ; Vagal ganglia ; Neuronal number and size ; Neuronal clustering ; Capsaicin ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The sensory cells of the nodose and jugular ganglia of the rat have been quantitatively evaluated in longitudinal paraffin sections. The right vagal ganglia contain significantly more neurons than the left, particularly neurons with somata having sectioned areas 200–400 μm2 and longest diameters 15–25 μm. Such neurons appear to be homogenously distributed because sections of the right vagal ganglia did not show specific areas of neuronal density compared with those of the left. Neonatal capsaicin treatment reduced the number of neurons in both the left and right ganglia to about 30% of controls. Capsaicin destroyed neurons with sectioned areas of 100–600 μm2 and longest diameters of 15–35 μm, but had no statistically significant effects on larger neurons.
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  • 77
    ISSN: 1432-0878
    Keywords: Key words: Neurotrophin receptor trkB ; p75NGFR ; Synaptophysin ; GAP-43 ; Cochlea ; Development ; ontogenetic ; Olivocochlear system ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The spatio-temporal distribution of the high-affinity neurotrophin receptor trkB was monitored during postnatal development of the rat cochlea. In addition to expression in presumptive afferent type I collaterals, afferent type II fibers, and efferent fibers, trkB immunoreactivity also transiently appeared in the sensory hair cells themselves, from postnatal days 5–9 in the basal turn, and from postnatal days 9–13 in the apical turn. A comparison of trkB with p75NGFR, which is expressed in afferent and efferent fibers, and GAP-43 and synaptophysin, which are expressed in efferent fibers, revealed a time/space correlation of trkB receptor expression in hair cells with the rearrangement of their innervation. Co-expression of the neurotrophin receptor and its ligand has been proposed to be functionally involved in regulating the survival of neurons independent of target-derived neurotrophin factor. Thus, the presence of trkB in target hair cells, suggests that auto/paracrine mechanisms play a role during this critical period of rearrangement of neural connections.
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  • 78
    ISSN: 1432-0878
    Keywords: Key words: Nitric oxide synthase ; Endothelin ; Pulmonary artery ; Pulmonary vein ; Ageing ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The ultrastructural distribution of nitric oxide synthase (neuronal isoform, type I) and endothelin immunoreactivity was examined in the developing and ageing male Wistar rat pulmonary artery and vein. This study demonstrates that from birth to old age (24 months) nitric oxide synthase and endothelin are localized within subpopulations of endothelial cells in the pulmonary vasculature. In the pulmonary artery and vein of newborn rats, and pulmonary vein of 24-month-old rats, positive labelling for nitric oxide synthase was also observed in the vascular smooth muscle. During development and ageing there were ultrastructural and immunocytochemical alterations in the intima of the pulmonary artery and vein. In older animals, damaged endothelial cells were seen alongside healthy-appearing cells, rich in cytoplasmic vesicles and endoplasmic reticulum. In contrast to damaged cells, the healthy-appearing endothelial cells displayed positive cytoplasmic labelling for nitric oxide synthase or endothelin. These immunopositive cells also appeared in the altered regions of the vessels where substantial enlargement of subendothelial extracellular matrix and the presence of various forms of degenerating macrophages and large bundles of collagen fibres were evident. Damage to the pulmonary artery was particularly evident at the ages of 12 and 24 months; various forms of macrophages, some of which displayed positive labelling for nitric oxide synthase and endothelin, were present in the altered intimal subendothelial zone. In conclusion, this study on the pulmonary vasculature suggests that endothelin and NOS in endothelial cells play a role in the local control of vascular tone throughout the lifespan of rats, even in older animals when there is intimal thickening and some endothelial damage. NOS and endothelin was also seen in smooth muscle and macrophages at certain stages in postnatal development and ageing.
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  • 79
    ISSN: 1432-0878
    Keywords: Key words: Substance P ; Receptors ; membrane ; Heart ; Binding sites ; Aorta ; Valves ; Neuropeptide immunocytochemistry ; Magnesium ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Magnesium(Mg)-deficiency, whether dietary or an effect of a clinical condition such as diabetes, results in a variety of cardiovascular pathologies. Substance P (SP) has been implicated in the induction of cardiac focal inflammatory lesions that occur during Mg-deficiency. Blockade of SP receptors results in a significant reduction in the incidence of lesion formation. In an effort to identify potential endogenous cell populations of the heart, which may play a role in SP-dependent lesion formation, film- and light-microscopic autoradiography were used to map the distribution of specific SP binding sites in frozen sections of the normal rat heart and adjacent great vessels. Binding was assessed with 0.1 nM I-125 Bolton-Hunter labelled SP in the absence (total binding) or presence (non-specific binding) of excess unlabelled SP, prolactin, or L-703,606, a non-peptide antagonist of SP receptors. Film autoradiograms revealed prominent small foci of intense autoradiographic reactions dispersed intermittently around the periphery of the great vessels and coronary arteries, among the interstitial connective tissue of the heart, and along the cusps of the cardiac valves. Excess unlabelled SP caused a significant reduction (97.7% displacement; P〈0.001) in the focal autoradiographic reactions. L-703,606 caused a similar reduction in SP binding (97.3% displacement; P〈0.001), while prolactin had no statistically significant effect on the binding of radiolabelled SP. Light-microscopic autoradiograms revealed that the SP binding sites occurred within clusters of connective tissue cells or in rarely observed parasympathetic ganglia. No evidence was found to suggest the presence of SP receptors on endothelial cells, cardiac muscle fibers, or smooth muscle fibers. The connective tissue cells which bound SP within the heart will likely include types that are susceptible to SP activation and thus may play a role in initiation of the focal inflammation characteristic of Mg-deficiency.
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  • 80
    ISSN: 1432-0878
    Keywords: Key words: Periodontium ; Osteopontin ; Bone sialoprotein ; Cell proliferation ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Cellular repopulation was studied in a model in which adjacent mineralising and soft connective tissue matrices are regenerated. Window wounds were created through alveolar bone, with either preservation or removal of periodontal ligament, in 30 male Wistar rats. Three animals per time period were killed on days 1, 3, 7, 14, and 28 after surgery for each wound type. Cellular proliferation in alveolar bone and periodontal ligament was assessed by 3H-thymidine labelling 1 h before death, followed by radioautographic analysis. Cellular differentiation was determined by the temporal expression of the bone-related markers osteopontin and bone sialoprotein, using immunohistochemical methods. In regenerating periodontium, osteopontin was expressed earlier than bone sialoprotein, which was restricted to alveolar bone. After wounding, transient expression of osteopontin was detected in the periodontal ligament at days 1 and 3. In general, wounding induced fivefold higher proliferation and clonal growth of periodontal ligament cells compared to the unwounded (control) side. Combined immunostaining and radioautography demonstrated colocalisation of osteopontin in sites with high numbers of labelled cells in both nascent periodontal ligament and regenerating alveolar bone at days 3 and 7. In contrast, bone sialoprotein, which appeared in regenerating alveolar bone on days 14–28 after wounding, was expressed much later than the peak of cellular proliferation. We conclude that (1) the intact periodontal ligament influences cell proliferation and osteopontin expression; (2) osteopontin is an early marker of periodontal tissue regeneration that is temporally and spatially associated with intensive cell proliferation and migration in osteogenic and periodontal ligament cell populations; and (3) bone sialoprotein is expressed after proliferation at sites of mineralising bone formation.
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  • 81
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    Cell & tissue research 286 (1996), S. 137-144 
    ISSN: 1432-0878
    Keywords: Key words: Muscle satellite cells ; Hypothyroid ; Proliferation ; Differentiation ; Muscle ; Triiodothyronine ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The effect of hypothyroidism, induced by 4-propyl-2-thiouracil, on muscle satellite cells in vivo and in vitro, and on postnatal muscle fiber development in the soleus muscle of rats during the first 40 days of postnatal life was analyzed. The proliferative activity of satellite cells was determined by means of bromodeoxyuridine incorporation. Creatine kinase activity was used as a marker for differentiation. In vivo, hypothyroidism resulted in smaller fibers in which the amount of sarcoplasm remained in balance with the number of myonuclei. The in vivo labeling data of satellite cells did not indicate a decreased proliferative activity, but the in vitro experiments showed that the hypothyroid rat muscles contained fewer satellite cells that were less active in proliferation and differentiation at the start of culture. Despite this, the bromodeoxyuridine signal increased in time at a similar rate as that in control cultures. From this and because the cells resembled control cells in their response to bFGF, we conclude that hypothyroid satellite cells remain responsive to proliferation stimuli. However, in hypothyroid cultures, the activity of creatine kinase is lower, even at longer culture times. We therefore conclude that hypothyroid status affects muscle precursor cells mainly by depressing their ability to differentiate and fuse with existing myofibers.
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  • 82
    ISSN: 1432-0878
    Keywords: Dopamine neurons ; Acetylcholinesterase ; Cholinesterase inhibitors ; Neurite outgrowth ; Neuron survival ; Organotypic culture ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract This study has investigated the possibility that acetylcholinesterase could play a non-classical role as an adhesion factor or growth factor in the development of dopaminergic neurons in organotypic slice culture of postnatal day 1 rats. When the culture medium was supplemented with acetylcholinesterase (3 U/ml), outgrowth of tyrosine hydroxylase-immunoreactive neurites was significantly enhanced. Addition of a specific inhibitor of acetylcholinesterase, BW284c51, caused a decrease in the number of tyrosine hydroxylase neurons and a reduction in the cell body size and extent of neurite outgrowth of remaining neurons. However, echothiophate which also inhibits AChE activity, did not produce these effects. Therefore acetylcholinesterase could act as a growth enhancing factor for dopaminergic neurons, and disruption of an as yet unidentified site on the acetylcholinesterase molecule by BW284c51 could decrease the survival and outgrowth of these neurons.
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  • 83
    ISSN: 1432-0878
    Keywords: Placenta ; Trophoblast ; Glucose transport ; GLUT 1-Man ; Marmoset monkey, Callithrix jacchus ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract In the present study, the facilitative D-glucose transporter protein GLUT 1 was localised by immunohistochemistry in the placenta of human, marmoset (Callithrix jacchus) and rat at different developmental stages. A polyclonal antiserum agains a 13-amino-acid peptide of the GLUT 1 carboxy terminus was used. It identified a protein of around 50 kDa molecular weight in immunoblotting of the placental tissues. GLUT 1 was located in the syncytiotrophoblast, in cytotrophoblast cells and in fetal endothelium. Similar staining patterns, except in human extravillous cytotrophoblast cells, were observed at all differentiation stages, despite differences in the internal placental architecture of the species. In the marmoset placenta, GLUT 1 was undetectable in endothelial cells of maternal vessels. In rat placentae, trophoblastic giant cells, epithelial cells of both visceral and parietal yolk sac, yolk sac vessels and the stratum spongiosum were stained. Reichert's membrane did not immunoreact. Preadsorption of the antiserum with a 13-amino-acid peptide resulted in the loss of immunoreactivity. The results suggest that GLUT 1 is a prominent isoform of glucose transporters in mammalian placentae. It is generally abundant in placental cell populations bordering on the maternal and fetal circulations and may therefore facilitate an effective glucose supply to the fetus and placenta.
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  • 84
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    Cell & tissue research 281 (1995), S. 197-206 
    ISSN: 1432-0878
    Keywords: Renin-angiotensin system ; Morphology — Renal tubules ; Ontogeny ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Recent evidence suggests that a local reninangiotensin system is operational in the kidney and that it mediates some of the actions of angiotensin II on renal tubules. In this study the ontogeny and renal distribution of the unique precursor to angiotensin II formation, angiotensinogen, was investigated in rats by use of immunohistochemistry, immuno-electron microscopy and non-isotopic hybridization histochemistry. At the light-microscopic level, intense staining for angiotensinogen was found in the proximal convoluted tubules of the cortex, with lighter staining in the straight proximal tubules of the outer stripe. The strongest immunostaining was found in the kidneys of neonatal rats, where glomerular mesangial cells and medullary vascular bundles were also immunopositive. The angiotensinogen content of the kidneys in late gestation embryos and neonates showed the presence of angiotensinogen by day E18 and a peak content in the neonate. Non-isotopic hybridization histochemistry with biotinylated oligodeoxynucleotide probes confirmed the presence of angiotensinogen mRNA expression in the proximal convoluted tubules of the renal cortex. Electron-microscopic immunohisto-chemistry showed staining of relatively few electron-dense structures close to the apical membrane of proximal convoluted tubule cells in the adult kidney. In the neonatal rat kidney, angiotensinogen immunostaining at the electron-microscopic level was found throughout the proximal tubule cells and was markedly stronger than that seen in adult kidney. The presence of angiotensinogen, from embryonic day 18, in the proximal tubules, mesangial cells and vasculature of the kidney suggests multiple potential sites of intrarenal angiotensin II generation with an ontogeny in late gestation.
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  • 85
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    Cell & tissue research 282 (1995), S. 155-161 
    ISSN: 1432-0878
    Keywords: Neurofilament ; Primary afferent fibres ; Skin ; Capsaicin ; Immunohistochemistry ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The mouse monoclonal antibody RT97, which recognises the 200-kDa neurofilament subunit in its phosphorylated form, selectively labels the somata of sensory A-fibres (large light cells) in the dorsal root ganglion of the rat. We have tested the hypothesis that this antibody also visualises large diameter sensory fibres and their end structures in peripheral tissue, in particular in the skin. RT97 immunoreactivity is found in endings that are known to be served by myelinated afferent fibres, including Meissner-like endings, Merkel discs, hair follicle receptors, Pacinian corpuscles and free nerve endings. RT97 immunoreactivity has not, however, been observed in endings of presumably unmyelinated sensory fibres (intraepidermal fibres immunoreactive for substance P and calcitonin gene-related peptide) or in sympathetic fibres innervating sweat glands and blood vessels. In addition, neither systemic (100–150 mg/kg as adults) nor perineural capsaicin pre-treatment affects RT97 immunoreactivity in the skin. The data indicate that RT97 is a useful marker in the study of the capsaicin-insensitive sensory innervation of the skin and possibly other peripheral organs.
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  • 86
    ISSN: 1432-0878
    Keywords: Key words: Esophagus ; Epithelial cells ; Intestinal lectin ; L-36 ; RI-H fragment ; Immunocytochemistry ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Using an affinity purified antibody raised against the RI-H fragment of rat intestinal lectin L-36, the latter protein has been identified within the esophageal epithelium by means of ultracryotomy followed by immunogold labeling. The epithelium consists of 4 morphologically distinct cell-types, namely, the basal, spiny, granular and squamous cells, and each of these exhibits a different immunolabeling pattern. The basal cells form a layer on the basal lamina, and in these a diffuse cytoplasmic staining is observed. This basal cell layer is overlaid by spiny cells that extend many cell processes into wide intercellular spaces. In these cells, immunogold particles are found only on small granular inclusions consisting of an electron-lucent homogeneous substance. The granular cells form a third layer over the spiny cells, and are characterized by a number of large granular inclusions with an electron-dense core rimmed by a less electron-dense substance. Immunogold labeling is found on these granules, both on the core and peripheral region. Squamous cell-types constitute the most superficial layer of the epithelium. They are without granular inclusions, and immunogold labeling is confined to the cytoplasmic surface of the thickened plasma membrane. These findings suggest that L-36 is produced in the basal cells as free cytosolic protein, then becomes progressively aggregated into the granular inclusions of the spiny and granular cells, and is eventually transferred onto the cytoplasmic surface of the squamous cell plasma membrane where it may interact with complementary glycoconjugate(s) located at this site. The membrane lining substance thus formed may play a role in stabilizing the squamous cell membranes, thereby maintaining the structural integrity of the epithelium against mechanical stress coming from the esophageal lumen.
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  • 87
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    Cell & tissue research 279 (1995), S. 565-573 
    ISSN: 1432-0878
    Keywords: Pineal gland ; Aging ; X-ray microanalysis ; Calcium concretions ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The genesis of calcium concretions in aged rats was studied by means of transmission and scanning electron microscopy. The potassium pyroantimonate method, combined with X-ray microanalysis, allowed us to study the distribution of cations and calcium. Notable accumulations of calcium (associated with phosphorus) were localized in vesicles, vacuoles, lipid droplets, lipopigments, and mitochondria of dark pinealocytes. The results obtained in the present investigation suggest that these organelles are involved in the genesis of the concretions. The presence of sulfur indicates the existence of an organic matrix. We propose that genesis takes place in dark pinealocytes, which contain more calcium than light pinealocytes. Mineralization foci are some-times associated with cellular debris and enlarge by further apposition of material. Two types of concretions, as determined by electron microscopy and confirmed by electron diffraction, could be observed: the “amorphous” type with concentric layers and the crystalline type with needle-shaped crystals. Once formed, the concretions reach the extracellular space and the cell breaks down. Possible extracellular calcification is suggested in the extracellular calcium-rich floculent material. The mineralization process is interpreted as being an age-related phenomenon and mainly a consequence of the degeneration of pinealocytes.
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  • 88
    ISSN: 1432-0878
    Keywords: Key words: Major pelvic ganglion ; Tyrosine hydroxylase ; Vasoactive intestinal polypeptide ; Neuropeptide Y ; Synaptophysin ; Colon ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The pelvic ganglia are mixed ganglia containing both sympathetic and parasympathetic neurons that receive spinal input via the hypogastric (lumbar cord) and pelvic nerves (sacral cord), respectively. A recent study has utilised immunohistochemistry against synaptophysin (a protein associated with small vesicles) to visualise the preganglionic terminals in these ganglia. By selectively cutting the hypogastric or pelvic nerves and allowing subsequent terminal degeneration, the populations of parasympathetic and sympathetic preganglionic terminals, respectively, can be visualised. The present study has used this method in conjunction with retrograde labelling of pelvic neurons from the distal colon and double label immunofluorescence against tyrosine hydroxylase and vasoactive intestinal polypeptide (VIP) to identify and characterise the sympathetic and parasympathetic neurons projecting to the distal colon from the major pelvic ganglia of the male rat. Approximately equal numbers of distal colonic-projecting pelvic neurons are sympathetic and parasympathetic. Almost all noradrenergic neurons are sympathetic. Of the VIP neurons that project to the distal colon approximately one third are sympathetic, one third parasympathetic and the remaining third are possibly innervated by both the lumbar and sacral cord. Extrapolation from our results also suggests that the majority of non-noradrenergic neuropeptide Y neurons (which are known to comprise the remainder of the neurons) are parasympathetic. These studies have demonstrated that the pelvic ganglia are a major source of sympathetic innervation to the distal bowel and have further shown that the distal colon is another target for the non-noradrenergic sympathetic neurons of the pelvic ganglia.
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  • 89
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    Cell & tissue research 280 (1995), S. 11-19 
    ISSN: 1432-0878
    Keywords: Key words: c-Fos ; c-Jun ; Hybridization ; in situ ; Myogenin ; Muscle regeneration ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. It has been suggested that myogenin is an important factor for the differentiation of myoblasts and that its function in myogenesis is regulated by proto-oncogenes in in vitro experiments. We have characterized the spatial and temporal expression patterns of myogenin, c-fos, c-jun, and muscle creatine kinase mRNAs during the skeletal muscle regeneration process using in situ hybridization histochemistry. Myogenin transcripts are first detected in the myonuclei/nuclei of satel lite cells at 6 h after induction of regeneration. Myogenin mRNA is expressed in desmin-positive myoblasts, yet no muscle creatine kinase mRNA is detected in this cell type. Both the muscle creatine kinase and myogenin mRNAs are expressed in the newly formed myotubes, but not at earlier stages. Transcripts for c-fos and c-jun mRNAs are expressed first in the myonuclei/nuclei of satellite cells at 3 h post-trauma. c-jun mRNA is expressed in both myoblasts and myotubes, while c-fos mRNA was not detected in these cells. These results suggest that myogenin plays important roles in the regeneration of injured muscle and that c-jun and c-fos may have different roles in this process.
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  • 90
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    Cell & tissue research 282 (1995), S. 277-289 
    ISSN: 1432-0878
    Keywords: Key words: Endolymphatic duct ; Endolymphatic sac ; Vascular supply ; Innervation ; Protein-gene product 9.5 (PGP 9.5) ; Peptides ; Dopamine-β-hydroxylase ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. A study of the ultrastructure, vascularization, and innervation of the endolymphatic duct and sac of the rat has been performed by means of light- and electron-microscopic and immunocytochemical methods. Two different types of epithelial cells have been identified: the ribosome-rich cell and the mitochondria-rich cell. These two cell types make up the epithelium of the complete endolymphatic duct and sac, although differences in their quantitative distribution exist. The morphology of the ribosome-rich cells varies between the different parts of the endolymphatic duct and sac; the morphology of the mitochondria-rich cells remains constant. According to the epithelial composition, vascularization, and structural organization of the lamina propria, both duct and sac are subdivided into three different parts. A graphic reconstruction of the vascular network supplying the endolymphatic duct and sac shows that the vascular pattern varies among the different parts. In addition, the capillaries of the duct are of the continuous type, whereas those supplying the sac are of the fenestrated type. Nerve fibers do not occur within the epithelium of the endolymphatic duct and sac. A few nerve fibers regularly occur in the subepithelial compartment close to the blood vessels; these fibers have been demonstrated in whole-mount preparations by the application of the neuronal marker protein gene product 9.5. Single beaded fibers immunoreactive to substance P and calcitonin-gene related peptide are observed within the same compartment. Dopamine-β-hydroxylase-immunoreactive axons are restricted to the walls of arterioles. Morphological differences between the different portions of the endolymphatic duct and sac are discussed with regard to possible roles in fluid absorption and immunocompetence.
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  • 91
    ISSN: 1432-0878
    Keywords: Macrophage ; Peritoneal cavity ; Omentum ; Depletion ; Repopulation ; Freund's adjuvant ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The purpose of this study was to develop a method for the depletion of macrophages from the peritoneal cavity and the omentum of the rat. Rats received two intraperitoneal injections (at days 0 and 3) with liposome-encapsulated clodronate (dichloromethylene bisphosphonate: Cl2MBP-liposomes). This treatment resulted in complete elimination of mature tissue macrophages (ED2-positive macrophages) from the peritoneal cavity and the omentum within 2 days. The elimination included the strongly ED2-positive spindle-shaped cells of the omental membrane. Repopulation of the omental ED2-positive macrophages was not seen within the next 23 days. Whereas ED2-positive macrophages were completely depleted, few ED1-positive cells remained and repopulation of ED1-positive cells was faster. The treatment further depleted macrophages from the spleen, especially from the red pulp, parathymic lymph nodes and liver. Freund's incomplete adjuvant administered one day after the last injection of Cl2MBP-liposomes considerably accelerated repopulation in the omentum. The protocol described might be used to investigate the contribution of mature tissue macrophages to the induction of immune responses, drug metabolism and the elimination of intestinal tumours.
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  • 92
    ISSN: 1432-0878
    Keywords: Key words: Thyroid gland ; C-cells ; Postnatal development ; Calcitonin ; Stereology ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The heterogeneous distribution of thyroid C-cells has until now hindered an objective evaluation of changes caused by age or experimental stimuli. To overcome this, a rigorous methodology has been designed to detect variations in shape, size, and number of C-cells throughout development. Using this methodology, we have demonstrated that C-cells do not significantly alter their shape with age. However, their volume increases gradually from 472 μm3 in newborn rats to 1653 μm3 in 120-day-old animals. Over the same time period, the mean number of C-cells within the thyroid gland increased 9-fold (from 1.6×104 to 1.5×105), and the number of C-cells per unit area decreased (from 6.15×104/mm3 to 2.6×104/mm3). We conclude that there are marked variations in size, total number, and number of C-cells per unit area in the rat thyroid gland after birth.
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  • 93
    ISSN: 1432-0878
    Keywords: Key words: Fibronectin ; Laminin ; Ribosomes ; p58 Membrane protein ; Immunoconfocal microscopy ; Immuno-electron microscopy ; Microtubules ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Polyclonal and monoclonal antibodies were used to examine the effects of the synthetic microtubule disruptive drug nocodazole on the subcellular expression of fibronectin, laminin, and ribosomes in primary cultures of neonatal cardiac ventricular cells. Non-invasive serial optical sectioning was carried out by immunolaser scanning confocal microscopy. In addition, fibronectin and laminin were immunolabelled with peroxidase or gold conjugates for electron-microscopic examination. Immunolabelling for the large 60S ribosome subunit in fibroblast-like non-myocytes showed that punctate ribosome structures with a multi-subunit composition were present in perinuclear region. Double immunostaining with antibodies directed against ribosomes and cellular fibronectin indicated that the punctate structures were cisternae of the rough endoplasmic reticulum. No clear effects of nocodazole treatment were detected on the distribution of cytoskeleton-bound ribosomes. Following immunolabelling for both glycoproteins and double immunolabelling for cellular fibronectin and the 60 S ribosome subunit, fibronectin and laminin were found in the perinuclear cisternae of the rough endoplasmic reticulum and in pleomorphic secretory vesicles. The cisternal stacks of the Golgi complex appeared either unstained or were only weakly labelled. When these cells were exposed to nocodazole, fibronectin and laminin accumulated in peripheral parts of the cytoplasm, including cellular processes. These peripheral accumulations of immunostaining for fibronectin and laminin did not reflect Golgi staining, as shown by double labelling experiments versus wheat-germ-agglutinin staining, and, by exposing cultures to a high dose of brefeldin A.
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  • 94
    ISSN: 1432-0878
    Keywords: Esophagus ; Epithelial cells ; Intestinal lectin, L-36 ; RI-H fragment ; Immunocytochemistry ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Using an affinity purified antibody raised against the RI-H fragment of rat intestinal lectin L-36, the latter protein has been identified within the esophageal epithelium by means of ultracryotomy followed by immunogold labeling. The epithelium consists of 4 morphologically distinct cell-types, namely, the basal, spiny, granular and squamous cells, and each of these exhibits a different immunolabeling pattern. The basal cells form a layer on the basal lamina, and in these a diffuse cytoplasmic staining is observed. This basal cell layer is overlaid by spiny cells that extend many cell processes into wide intercellular spaces. In these cells, immunogold particles are found only on small granular inclusions consisting of an electron-lucent homogeneous substance. The granular cells from a third layer over the spiny cells, and are characterized by a number of large granular inclusions with an electron-dense core rimmed by a less electron-dense substance. Immunogold labeling is found on these granules, both on the core and peripheral region. Squamous cell-types constitute the most superficial layer of the epithelium. They are without granular inclusions, and immunogold labeling is confined to the cytoplasmic surface of the thickened plasma membrane. These findings suggest that L-36 is produced in the basal cells as free cytosolic protein, then becomes progressively aggregated into the granular inclusions of the spiny and granular cells, and is eventually transferred onto the cytoplasmic surface of the squamous cell plasma membrane where it may interact with complementary glycoconjugate(s) located at this site. The membrane lining substance thus formed may play a role in stabilizing the squamous cell membranes, thereby maintaining the structural integrity of the epithelium against mechanical stress coming from the esophageal lumen.
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  • 95
    ISSN: 1432-0878
    Keywords: Fibronectin ; Laminin ; Ribosomes ; p58 Membrane protein ; Immunoconfocal microscopy ; Immuno-electron microscopy ; Microtubules ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Polyclonal and monoclonal antibodies were used to examine the effects of the synthetic microtubule disruptive drug nocodazole on the subcellular expression of fibronectin, laminin, and ribosomes in primary cultures of neonatal cardiac ventricular cells. Non-invasive serial optical sectioning was carried out by immunolaser scanning confocal microscopy. In addition, fibronectin and laminin were immunolabelled with peroxidase or gold conjugates for electron-microscopic examination. Immunolabelling for the large 60S ribosome subunit in fibroblast-like non-myocytes showed that punctate ribosome structures with a multi-subunit composition were present in perinuclear region. Double immunostaining with antibodies directed against ribosomes and cellular fibronectin indicated that the punctate structures were cisternae of the rough endoplasmic reticulum. No clear effects of nocodazole treatment were detected on the distribution of cytoskeleton-bound ribosomes. Following immunolabelling for both glycoproteins and double immunolabelling for cellular fibronectin and the 60 S ribosome subunit, fibronectin and laminin were found in the perinuclear cisternae of the rough endoplasmic reticulum and in pleomorphic secretory vesicles. The cisternal stacks of the Golgi complex appeared either unstained or were only weakly labelled. When these cells were exposed to nocodazole, fibronectin and laminin accumulated in peripheral parts of the cytoplasm, including cellular processes. These peripheral accumulations of immunostaining for fibronectin and laminin did not reflect Golgi staining, as shown by double labelling experiments versus wheat-germ-agglutinin staining, and, by exposing cultures to a high dose of brefeldin A.
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  • 96
    ISSN: 1432-0878
    Keywords: Key words: Nitric oxide synthase ; Immunohistochemistry ; Retrograde tracing ; Celiac-superior mesenteric ganglion ; Sensory ganglion ; Spinal cord ; Intestine ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. The origin of nitric oxide synthase-containing nerve fibers in rat celiac-superior mesenteric ganglion was examined using retrograde tracing techniques combined with the immunofluorescence method. Fluoro-Gold was injected into the celiac-superior mesenteric ganglion. Neuronal cell bodies retrogradely labeled with Fluoro-Gold in the thoracic spinal cord, the dorsal root ganglia at the thoracic level, the nodose ganglion, and the intestine from the duodenum to the proximal colon were examined for nitric oxide synthase immunoreactivity. About 60% of sympathetic preganglionic neurons in the intermediolateral nucleus projecting to the celiac-superior mesenteric ganglion were immunoreactive for nitric oxide synthase, as were approximately 27% of nodose ganglion neurons and about 65% of dorsal root ganglion neurons projecting to the celiac-superior mesenteric ganglion. Neurons projecting to the celiac-superior mesenteric ganglion were found in the myenteric plexus of the small and large intestine. In the proximal colon, about 23% of such neurons were immunoreactive for nitric oxide synthase. However, in the small intestine, no immunoreactivity was found in these neurons.
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  • 97
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    Cell & tissue research 281 (1995), S. 533-542 
    ISSN: 1432-0878
    Keywords: Pelvic plexus ; Neuropeptides ; Tyrosine hydroxylase ; Reproductive tract, male ; Synaptophysin ; FluoroGold ; Retrograde tracing ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Autonomic innervation of the prostate gland supplies the acini, and non-vascular and vascular smooth muscle. The activity of each of these tissues is enhanced by sympathetic outflow, whereas the role of the parasympathetic nervous system in this organ is unclear. In the present study, a range of methods was applied in rats to determine the location of autonomic neurons supplying this gland, the immunohistochemical properties of these neurons, the spinal connections made with the postganglionic pathways and the distribution of various axon types within the gland. Injection of the retrograde tracer, FluoroGold, into the ventral gland visualised neurons within the major pelvic ganglion and sympathetic chain. Fluorescence immunohistochemical studies on the labelled pelvic neurons showed that most were noradrenergic (also containing neuropeptide Y, NPY), the others being non-noradrenergic and containing either vasoactive intestinal peptide (VIP) or NPY. Sympathetic dyelabelled neurons were identified by the presence of varicose nerve terminals stained for synaptophysin on their somata following lesion of sacral inputs. Parasympathetic innervation of dye-labelled neurons was identified by continued innervation after hypogastric nerve lesion. Most noradrenergic prostate-projecting neurons were sympathetic, as were many of the non-noradrenergic VIP neurons. Parasympathetic prostate-projecting neurons were largely non-noradrenergic and contained either VIP or NPY. All substances found in retrogradely labelled somata were located in axons within the prostate gland but had slightly different patterns of distribution. The studies have shown that there are a significant number of non-noradrenergic sympathetic prostate-projecting neurons, which contain VIP.
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  • 98
    ISSN: 1432-0878
    Keywords: CD44, adhesion molecule ; Bone ; Osteoclasts ; Osteocytes ; Immunohistochemistry ; Confocal laser scanning microscopy ; Electron microscopy ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract CD44 is a multifunctional adhesion molecule that binds to hyaluronic acid, type I collagen, and fibronectin. We have studied the immunohistochemical localization of CD44 in bone cells by confocal laser scanning microscopy and transmission electron microscopy in order to clarify its role in the cell-cell and/or cell-matrix interaction of bone cells. In round osteoblasts attached to bone surfaces, immunoreactivity is restricted to their cytoplasmic processes. On the other hand, osteocytes in bone matrices show intense immunoreactivity on their plasma membrane. Intense immunoreactivity for CD44 can be detected on the basolateral plasma membranes of osteoclasts. There is considerably less reactivity observed in the area of the plasma membrane that is in direct contact with bone. The pre-embedding electron-microscopical method has revealed that CD44 is mainly localized on the basolateral plasma membrane of osteoclasts. However, the ruffled border and clear zone show little immunoreactivity. A CD44-positive reaction can be detected on both plasma membranes in the contact region between osteoclasts and osteocytes. These findings suggest that: 1) cells of the osteoblast lineage express CD44 in accordance with their morphological changes from osteoblasts into osteocytes; 2) osteoclasts express CD44 on their basolateral plasma membrane; 3) CD44 in osteoclasts and osteocytes may play an important role in cell-cell and/or cell-matrix attachment via extracellular matrices.
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  • 99
    ISSN: 1432-0878
    Keywords: Thyroid gland ; C-cells ; Postnatal development ; Calcitonin ; Stereology ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The heterogeneous distribution of thyroid C-cells has until now hindered an objective evaluation of changes caused by age or experimental stimuli. To overcome this, a rigorous methodology has been designed to detect variations in shape, size, and number of C-cells throughout development. Using this methodology, we have demonstrated that C-cells do not significantly alter their shape with age. However, their volume increases gradually from 472 μm3 in newborn rats to 1653 μm3 in 120-day-old animals. Over the same time period, the mean number of C-cells within the thyroid gland increased 9-fold (from 1.6x104 to 1.5x105), and the number of C-cells per unit area decreased (from 6.15x104/mm3 to 2.6x104/mm3). We conclude that there are marked variations in size, total number, and number of C-cells per unit area in the rat thyroid gland after birth.
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  • 100
    ISSN: 1432-0878
    Keywords: Key words: Dopamine neurons ; Acetylcholinesterase ; Cholinesterase inhibitors ; Neurite outgrowth ; Neuron survival ; Organotypic culture ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. This study has investigated the possibility that acetylcholinesterase could play a non-classical role as an adhesion factor or growth factor in the development of dopaminergic neurons in organotypic slice culture of postnatal day 1 rats. When the culture medium was supplemented with acetylcholinesterase (3 U/ml), outgrowth of tyrosine hydroxylase-immunoreactive neurites was significantly enhanced. Addition of a specific inhibitor of acetylcholinesterase, BW284c51, caused a decrease in the number of tyrosine hydroxylase neurons and a reduction in the cell body size and extent of neurite outgrowth of remaining neurons. However, echothiophate which also inhibits AChE activity, did not produce these effects. Therefore acetylcholinesterase could act as a growth enhancing factor for dopaminergic neurons, and disruption of an as yet unidentified site on the acetylcholinesterase molecule by BW284c51 could decrease the survival and outgrowth of these neurons.
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