ALBERT

Notes: The development of a virus is programmed by a series of negative and positive controls which determine the timing and the segment on either of the two DNA strands (l or r) to be transcribed into specific messenger RNA's. Bacteriophage λ provides one of the most deeply studied systems for following the development of lysogenic viruses. In the lysogenic repressed state, only 2-4% of the λ genome is expressed. This pc-cI-rex region is transcribed leftward to produce a repressor protein which prevents any further transcription by blocking the oL and oR operators flanking the cI-rex operon (figs. 1, 2). This negative control is relieved by destruction of the repressor, and the result is the “induction” of viral development. The earliest post-induction or postinfection events are the leftward transcription of the pLoL N region from strand l and the rightward transcription mainly of the pRoR-x segment from strand r. The N product acts as a positive control, permitting a leftward transcription beyond gene N and a rightward transcription of genes cII-O-P and also Q. The int-xis system controls the excision of the λ genome, whereas the act of rightward transcription and the products of genes O and P initiate the replication of λ DNA. The product of gene Q, still another positive control, stimulates rightward transcription of the late genes which control the synthesis and assembly of the phage heads and tails as well as cell lysis. Among other types of negative control are the possible competition between the two divergent transcriptions originating in region x, the “antirepressor” effect of the x product, and the interference between the two convergent transcriptions which collide in the central b2 region. The majority of controls are based on protein-DNA interactions and can be modified by mutations. For instance, transcription can be rendered independent of negative repressor control either by constitutive, v, mutations which decrease or abolish the affinity of the o operators for the repressor or by insertion of new promoters-e.g., c17 or ric- on the “downstream” side of the operator. The need for the positive N and Q controls may also be obviated by mutations in the N- or Q-dependent promoter or terminator elements.The specific DNA structure within the controlling sites is not known. However, a remarkable coincidence was observed; namely, the occurrence of pyrimidine-rich clusters in those segments of the individual DNA strands acting as templates for RNA synthesis. This observation, which pertains to all studied DNA's, including those of phages T2, T3, T4, T5, T6, T7, λ, and φ 80, formed the basis for a proposal that implicates pyrimidine-rich clusters in the initiation, control and/or termination of transcription, and also in the determination of the preferred strand and, consequently, the orientation of transcription. General considerations regarding the possible role of the structural singularities, especially those represented by the pyrimidine clusters, in the bipartite structure of the recognition regions in DNA are discussed.

Notes: The cytological effects of 2 mM hydroxyurea upon Chinese hamster cells at various phases of the cell cycle were examined. Cells in the G1, G2, or M phases of the generation cycle treated with hydroxyurea showed no chromosomal aberrations. Cell treated in S phase became moribund and eventually lysed. Some of these moribund S cells reached mitosis much later and were found to have chromatid aberrations. Cells in the log phase of growth, surviving exposure to 2 mM hydroxyurea for six hours, also showed no aberrations. Thus, viable (colony-forming) cells, resulting from synchrony procedures with hydroxyurea are free of chromosomal aberrations.

Notes: Ten adult western gulls (Larus occidentalis) ranging in weight from 761-1,004 gm were studied. The gulls were fed 3% NaCl in their drinking water. They were killed by decapitation, and the salt glands, weighing 0.51-0.78 gm were cooled, homogenized in 0.14 M KCl, centrifuged at 0-4°C at 20,000 × G for 30 minutes and the supernatant used for all enzymes assays. All assays were conducted at 25°C by observing the changes in absorbancy with time using a Gilford Multiple Absorbance Recorder. The enzymes were assayed by measuring either the appearance or disappearance of NADH or NADPH at 340 mμ. The average units of enzyme activity (the amount of enzyme required to form 1 μM of substrate per minute ) per gram of salt gland were as follows: phosphoglucomutase, 0.62; glucose-6-phosphate dehydrogenase, 1.40; aldolase, 2.86; lactic dehydrogenase, 90.1; isocitric dehydrogenase, 5.08; malic enzyme, 0.92; glutamic-oxaloacetic transaminase, 100.5; and glutamic-pyruvic transaminase, 0.50. The protein content of the salt glands varied from 62.5-87.6 mg protein/gm. On the basis of an adjusted calculation of energy yields from the glycolytic scheme and the Krebs cycle, it would appear that only one-third of the energy derived from these pathways would be necessary to maintain the maximum rate of salt secretion, leaving the other two-thirds for other cellular processes. Glutamate metabolism may also be important as an energy source in the salt gland.

Notes: The intact frog (R. pipiens) is both very sensitive to i.v. injection of l-epinephrine (E) in small amounts (a few micrograms), and very tolerant to large doses (several milligrams) of the hormone. A small dose increased the P.D. and the short circuit current (Is) in skin, and strongly increased the strength of auricular contraction of the heart in vivo; ventricular contractions decreased, and heart rate remained normal. Large doses of E depressed P.D. and Is after, occasionally, elevating both for a brief period following treatment. The mucous glands of the skin emptied their content shortly after E, then began to fill again with secretory materials in spite of uninterrupted E infusions. Ventricular and auricular contractions were remarkably little altered and there were no significant changes in heart rate. Changes were seen, however, in the ECG, e.g.: inversion of the P and T waves. No alteration in the QRS complex or in the rhythm was ever noted. Serum glucose remained within normal limits. A transient calorigenic effect, lasting for 10-20 minutes occurred, in which the O2 consumption rose from a normal of 6 to 7 ml/100 g/hr to several times this value. Unanesthetized frogs tolerated without great harm 10 mg of E given s.c. within one hour. Signs of prostration and a moderate increase in respiration were noted for some time following treatment. Great tolerance of frogs to hormones which do play a physiological role is also known for thyroxine and insulin, but is especially notable for epinephrine which is least tolerated by warmblooded animals treated with large doses of E. The reasons for these species differences are unknown.

Notes: The cells of an established mouse fibroblast line, 3T3, have a high plating efficiency and grow rapidly in sparse culture, but stop growing at a very low saturation density in comparison with other lines, because 3T3 is extremely sensitive to contact inhibition of cell division. After each medium change, however, there occurs in a small fraction of the cells in a saturation density culture a series of changes that results in a single rather synchronized division 30 hours later. This is due to a macromolecular substance in the serum which appears to act by reducing the sensitivity of the cells to contact inhibition. The first recognizable event following the addition of serum to a stationary phase culture is a ten fold increase in the rate of RNA synthesis, occurring within 30 minutes. An increase in the rate of protein synthesis follows several hours later. DNA synthesis does not begin before 12 hours, but by two hours after medium change an appreciable fraction of the cells become committed to eventual DNA synthesis and cell division. The sequence of event suggests that regulation of RNA synthesis is the means by which contact inhibition controls cell division.

Notes: Epithelio-mesodermal interaction in mature thyroids of older chick embryos was tested by dissociation, monolayer culture to disrupt cytoplasmic organization, and reaggregation with heterologous and homologous mesoderm. Each combination formed a different histological pattern: with mesentery  -  islands of epithelial cells in a loose connective tissue; with heart  -  follicles at the ends of cords in a rich vascular bed, with perichondrium  -  a ring of follicles surrounding masses of cartilage; with thyroid capsule  -  normal thyroid organization. Electron-microscopic examination of cells incorporated into follicles revealed reorganization of canaliculate endoplasmic reticulum in only the homotypic combination. Therefore, interactions occur in mature organs as well as primordia.

Notes: Neurosecretory activity and fine structure of the supraesophageal and optic ganglia of Daphnia schødleri Sars were studied. The relative amount of paraldehyde fuchsin stainable material present was determined at “daylight” and at one and three hours following for animals maintained under photoperiods of 7.5, 10.5, 13.5, and 16.5 hours. More material was found after one hour in both ganglia and there was a tendency for more in the optic ganglion under 7.5- and 10.5-hour photoperiods.Sections made at two levels in the supraesophageal ganglion and one level in the optic ganglion were examined with an electron microscope. Posterior and anterior parts of the supraesophageal ganglion contain apparent nerve processes at the edge of the ganglion, parallel to the anterior-posterior axis; these have large granules. Neurons in both areas contain patches of presumed polysaccharide granules. In the posterior region are a dorsolateral and a ventrolateral glandular cell, presumably these occur on both sides of the brain. They have very well-developed endoplasmic reticulum and some large granules. The dorsal cell is usually at the tip of a glial attenuation.Concentric lamellar systems are located in dilated nerve processes of the first optic ganglion neuropile. Large whorls (about 3.5 μ) are composed of concentric lamellae. When lamellae do not form complete rings, they end in loops or in dilated tubules that are sometimes constricted as vesicles. Small whorls (1.5 μ) typically have lamellae joined into two or three thick layers. Mitochondria are frequently associated with the whorls. It is proposed that the whorls are active in synthesis, possibly neurohumor production.

Notes: The processes of dissociation and reaggregation were studied in thyroids from 16-day chick embryos by means of electron microscopy. The purpose was to gain specific knowledge of the effects that dissociating agents have upon cells.Fixation after a soak in trypsin showed little effect on secretory cell structure but resulted in marked swelling and detachment of connective tissue elements and disorganization of the basement lamella of the follicles. After an additional exposure to ethylenediaminetetraacetate (EDTA), however, the channels of the endoplasmic reticulum and the Golgi vesicles became swollen and separations appeared between adjacent secretory cells. Since detachment did not occur in the region of the junctional complex, agitation was required to achieve suspension of the cells. Even this treatment resulted in only a small proportion of individual cells, the majority being in the form of small clumps or chains. Transfer of the suspensions to nutrient medium and reaggregation produced pellets which rapidly formed follicles in organ cultures. The individual cells showed the same degree of complexity as in follicles in vivo.The results are related to the known effects of trypsin and EDTA on cells. The significance of morphogenesis in aggregates prepared from cells of older embryonic organs is also discussed.

Notes: Silver stained Cordylophora were examined by light and electron microscopy, which provided a general picture of nerve cell forms and distribution for comparison with electron micrographs of osmium-fixed tissues from the same hydroid. Muscle, nerve and neurosensory components were studied in the nectophore of Nanomia (O. Siphonophora) and in the hydromedusae Sarsia and Euphysa by means of vital staining and optical and electron microscopy of epon sections; particular attention was given to relationships and interconnections between the cellular elements of the two marginal nerve rings. Mitochondrial size, numbers and types of vesicles and the occurrence of neurotubules and of parts of sensory cilia may provide useful ultrastructural clues for recognizing nerve elements, but serial sections are often needed to make identification conclusive.In Cordylophora and Nanomia, some neurites contain massed A vesicles (membrane-bounded dense granules) suggestive of neurosecretion (cf. reports on Hydra). However, a small type of A vesicle also occurs at synapses in Sarsia, indicating a probable role here in junctional transmission. Vesicles occur on both sides of some synapses (as previously reported for Cyanea) but on one side only in others, these being the first examples of polarized junctional ultrastructure in coelenterates.

Notes: The morphology, seasonal variation and histochemistry of the sexual segment of the Indian house lizard, Hemidactylus flaviviridis are described.The sexual segment is the hypertrophied portion of the secondary and the tertiary collecting ducts of the kidney in males. The cells of the sexual segment are columnar and are loaded with secretory granules which are predominantly localized in the apical portion. These granules are either free or occasionally clumped to form an “aggregate” towards the basement membrane and are released into the lumen by apocrine secretion. Development of the sexual segment is synchronous with the spermatogenic activity of the testis and maximum development occurs in March and April when the lizards copulate. The sexual segment is secretory from the beginning of October to the end of May and is regressed during sexual quiescence (June and July). It is not developed in females and young of both sexes in any season of the year.Histochemically, the sexual segment granules are saliva resistant and PAS positive, sudanophilic and are positive to the tests for phospholipid, choline and acid phosphatase. An intense esterase activity was localized in the mucosa of the oviduct and cloaca of the female. Sperms are mixed with sexual segment secretions and are transferred to the female during copulation. A possible role for the lipid-rich sexual segment secretion as a source of energy for sperms in the female reproductive tract is discussed.