Immunohistochemical localization of PACAP in the ovine digestive system

doi:10.1016/0196-9781(93)90131-Y

Peptides

Volume 14, Issue 3, May–June 1993, Pages 449-455

https://doi.org/10.1016/0196-9781(93)90131-Y Get rights and content

Abstract

The localization of immunoreactive PACAP (PACAP-IR) in the entire length of the sheep gastrointestinal tract and the pancreas was studied by an immunohistochemical method. PACAP-IR-containing nerve fibers innervated the longitudinal smooth muscle layer of the mucosa in the esophagus, stomach, and small and large intestines, the muscular layer of the stomach and intestine, Brunner's gland of the duodenum, and the walls of small arteries. PACAP-IR fibers also innervated the exocrine acini, islets of Langerhans, and the small arteries in the connective tissue septa of the pancreas. These findings suggest a regulatory role of PACAP in the digestive system.

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Cited by (72)

Long-term treatment with naproxen changes the chemical coding of the porcine intramural duodenum neurons
2020, Annals of Anatomy
Citation Excerpt :
The presence of VIP and PACAP specific receptors was found in the gastrointestinal tract. Neurons and nerve fibers immunoreactive to these substances have also been noted in the stomach and intestines (Gonkowski et al., 2015; Köves et al., 1993; Makowska et al., 2018). VIP-ergic and PACAP-ergic neurons are the most likely component of internal intestinal reflexes that regulate the secretion of bicarbonate.
Due to numerous therapeutic applications and high availability, non-steroidal anti-inflammatory drugs (NSAIDs) are the most widely used drugs worldwide. However, long-term use of these drugs can lead to damage to the gastrointestinal mucosa. The enteric nervous system (ENS), which is part of the autonomic nervous system, controls most aspects of gastrointestinal activity. Enteric neurons are characterized by considerable chemical plasticity and the appearance of a pathological factor results in a change in the synthesis of neurotransmitters.
The purpose of this study was to determine the effects of naproxen on expression of biologically active substances by intramural neurons supplying the porcine duodenum. The study was performed on eight immature pigs of the Pietrain x Duroc race (approximately 20 kg of body weight). The animals were divided into two groups - a control (C group) and an experimental group (N group). Group C (n = 4) consisted of animals which received empty gelatine capsules. Group N (n = 4) was composed of pigs who received naproxen orally for 28 days, approximately one hour before feeding. After this time, animals from both groups were euthanized. Frozen sections (14 μm thickness) were then prepared from the collected duodenum and subjected to double immunofluorescence staining. Antibodies against the neuronal marker PGP 9.5 and against vasoactive intestinal polypeptide (VIP), substance P (SP), neuronal nitric oxide synthase (nNOS), galanin (GAL), pituitary adenylate cyclase-activating polypeptide (PACAP) and cocaine- and amphetamine- regulated transcript peptide (CART) were used as primary antibodies. The polyclonal donkey anti-rabbit, anti-mouse and anti-guinea pig IgG antibodies - Alexa Fluor 488 and 546 - were also used for staining.
Analysis of the results obtained with a fluorescence microscope showed a significant increase in the number of nNOS-, VIP-, GAL-, PACAP- and CART-immunoreactive ganglionated neurons and a decrease in the number of SP-positive neurons in the myenteric and submucosal plexuses of the porcine duodenum. The obtained results indicate the participation of enteric neurotransmitters in the neuronal duodenal response to naproxen-induced inflammation.
Effects of pituitary adenylate cyclase activating polypeptide on small intestinal INT 407 cells
2017, Neuropeptides
Citation Excerpt :
VIP has long been known as an important intestinal peptide with various effects (Padua et al., 2016). Similarly, PACAP is distributed throughout the gastrointestinal system (Ganko and Calka, 2014a, b; Gonkowski et al., 2015; Koves et al., 1993; Nedvig et al., 2012; Szanto et al., 2012; Tamas et al., 2016; Vu et al., 2016). The presence of PACAP and/or its receptors has been shown in neuroendocrine cells, interstitial cells and fibers of the myenteric and submucosal plexus of the entire gastrointestinal tract as well as other organs of the digestive system, such as pancreas, gall bladder and liver (Arciszewski et al., 2015; Ji et al., 2013; Koves et al., 1993; Oh et al., 2005; Vu et al., 2016; Zhang et al., 2006).
Pituitary adenylate cyclase activating polypeptide (PACAP) is an endogenous neuropeptide having a widespread distribution both in the nervous system and peripheral organs including the gastrointestinal tract. It has been shown to exert actions on intestinal functions, mainly affecting glandular secretion and motility. PACAP has several different effects on cell survival depending on the cell type and the applied stimulus. Its influences on small intestinal epithelial cells are not yet elucidated, therefore the aim of the present study was to investigate the effects of PACAP on intestinal epithelial cells having high turnover (INT 407) against different harmful stimuli, such as oxidative stress, in vitro hypoxia and gamma radiation. We tested the effect of PACAP on proliferation and cell survival using MTT assay. Moreover, various cancer-related factors were evaluated by oncology array. PACAP did not influence the proliferation rate of INT 407 cells. Its cell survival-enhancing effect could be detected against oxidative stress, but not against in vitro hypoxia or gamma irradiation. Clonogenic survival assay was performed to analyze the effect of PACAP on clonogenic potential of cells exposed to gamma radiation. Surprisingly, PACAP enhanced the clone-forming ability decrease induced by irradiation. Western blot analysis of ERK1/2 phosphorylation was performed in order to obtain further information on the molecular background. Our data showed phospho-ERK1/2 suppression of PACAP in irradiated cells. Furthermore, the role of endogenous PACAP against oxidative stress was also investigated performing ADCYAP1 small interfering RNA transfection. We found significant difference in the cell vulnerability between cells undergoing silencing and cells without transfection suggesting the protective role of the endogenously present PACAP against oxidative stress in INT 407 cells. In summary, PACAP seems to be able to exert contradictory effects in INT 407 cells depending on the applied stressor, suggesting its regulatory role in the cellular household.
PACAP is transiently expressed in anterior pituitary gland of rats: In situ hybridization and cell immunoblot assay studies
2008, Peptides
In this work the expression of PACAP (pituitary adenylate cyclase activating polypeptide) in rat anterior pituitary was demonstrated for the first time using in situ hybridization. The number of cells showing PACAP signal in intact male rats was negligible similarly to that of diestrous rats. In proestrous rats sacrificed at 10 h there was a moderate increase in the expression and after a decrease at 16 h and 18 h, there was a transient peak at 20 h and then the number of labeled cells was declined again (22 h). In the cell immunoblot assay study it was observed that the number of PACAP blot forming (PACAP releasing) cells in an anterior pituitary cell culture changed according to a similar pattern as the number of PACAP expressing cells. The number of blots was also the highest when the animals were sacrificed in the evening of proestrus at 20 h. The results obtained by in situ hybridization and cell immunoblot assay well correlate with each other. The above-mentioned results support our hypothesis that the enhanced expression and secretion of PACAP in the pituitary gland may be involved in ceasing the LH surge.
Effects Of PACAP In the Local Regulation Of Endocrine Glands
2006, Handbook of Biologically Active Peptides
Pituitary adenylate cyclase–activating polypeptide (PACAP), a peptide of the vasoactive intestinal polypeptide (VlP)-glucagon superfamily, was initially characterized by virtue of its ability to stimulate cAMP formation in cultured rat anterior pituitary cells. Three PACAP receptors have been cloned so far: a PACAP-selective receptor, termed PAC1-R, and two PACAP/VIP common receptors, termed VPAC1-R and VPAC2-R. PACAP and its receptors are widely expressed in the brain and in peripheral organs, notably in the hypothalamus and in endocrine glands. Indeed, there is now clear evidence that PACAP exerts neuroendocrine, paracrine, and autocrine control of the activity of the pituitary, thyroid, testis, ovary, adrenal medulla, adrenal cortex, and endocrine pancreas. These observations suggest that selective PACAP agonists and antagonists could have therapeutic value for the treatment of various endocrine disorders.
Distribution of PACAP-38 in the central nervous system of various species determined by a novel radioimmunoassay
2004, Journal of Biochemical and Biophysical Methods
Pituitary adenylate cyclase activating polypeptide (PACAP) occurs in two molecular forms: PACAP-38 and PACAP-27. Soon after the isolation and chemical characterization of PACAP, the first radioimmunoassay (RIA) methods have been developed, but it is a still rarely used laboratory technique in the field of PACAP research. The aim of the present study was to develop a novel, highly specific PACAP-38 assay to investigate the quantitative distribution of PACAP-38 in the central nervous system of various vertebrate species under the same technical and experimental conditions. Different areas of the brain and the spinal cord were removed from rats, chickens and fishes and the tissue samples were processed for PACAP-38 RIA. Our results indicate that the antiserum used in the RIA is C-terminal specific, without affinity for other members of the vasoactive intestinal polypeptide (VIP)/secretin/glucagon peptide family. The average ID₅₀ value was 48.6±3.4 fmol/ml determined in 10 consecutive assays. Detection limit for PACAP-38 proved to be 2 fmol/ml. PACAP-38 immunoreactivity was present in the examined brain areas of each species studied, with highest concentration in the rat diencephalons. High levels of PACAP-38 were also detected in the rat telencephalon, followed by spinal cord and brainstem. The central nervous system of the fish also contained considerable concentrations of PACAP-38, whereas lowest concentrations were measured in the central nervous system of the chicken.
Interplay between PACAP and NO in mouse ileum
2004, Neuropharmacology
We investigated the possibility that pituitary adenylate cyclase activating peptide (PACAP) has a role in the control of contractility in the mouse ileum. PACAP-(1-27) produced tetrodotoxin (TTX)-insensitive, concentration-dependent reduction of the amplitude of the spontaneous contractions of longitudinal muscle up to their complete disappearance. This effect was inhibited by PACAP-(6-38), PACAP receptor antagonist, and by apamin, blocker of small-conductance Ca²⁺-activated K⁺-channels. N_ω-nitro-l-arginine methyl ester (L-NAME), nitric oxide (NO) synthase inhibitor, reduced the PACAP-inhibitory response, and the joint application of apamin plus L-NAME produced additive effects. 1H-[1,2,4] oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), inhibitor of NO-stimulated soluble guanylate cyclase, significantly reduced the effect of PACAP. Exogenous NO, given as sodium nitroprusside (SNP), induced a concentration-dependent suppression of the phasic contractions, which was unaffected by apamin but reduced by either PACAP-(6-38) or TTX. Neurally evoked muscular relaxation was deeply antagonised by L-NAME. PACAP-(6-38) induced a reduction of the response to EFS only in the absence L-NAME. In conclusion, our results suggest that PACAP controls smooth muscle contractility, acting directly on the muscle cells through PACAP-27 preferring receptors coupled to apamin-sensitive Ca²⁺-dependent K⁺-channels and indirectly through the stimulation of NO production. In turn, NO would stimulate the release of PACAP from inhibitory neurones.

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ArticleImmunohistochemical localization of PACAP in the ovine digestive system

Abstract

FEBS Lett.

Immunohistochemistry

Peptides

Neuroscience

Neuroscience

Peptides

Biochem. Biophys. Res. Commun.

Biochem. Biophys. Res. Commun.

Brain Res.

Life Sci.

VIP- and CCK-like immunoreactive neurons in the hedgehog (Erinaceus europaeus) and sheep (Ovis aries) brain

J. Comp. Neurol.

Characterization and distribution of binding sites for the hypothalamic peptide, pituitary adenylate cyclase activating polypeptide

Endocrinology

Human preprovasoactive intestinal polypeptide contains a novel PHI-like peptide, PHM-27

Nature

Article
Immunohistochemical localization of PACAP in the ovine digestive system