Pituitary adenylate cyclase activating polypeptide (PACAP) in the rat mammary gland

Pituitary adenylate cyclase activating polypeptide (PACAP), a member of the vasoactive intestinal polypeptide (VIP) family of peptides, is present in the brain and in neuronal elements of a number of peripheral organs. Since no information on PACAP in the mammary gland exists, we have investigated, by radioimmunoassay and immunohistochemistry, the occurrence and distribution of PACAP immunoreactivity in the mammary gland of lactating and non-lactating rats. A specific monoclonal mouse anti-PACAP antibody'has been used to show that the peptide is located in nerve fibres associated with bundles of circular and longitudinal smooth muscle surrounding the lactiferous duct of the nipple. PACAP-immunoreactive nerve fibres and nerve bundles are present in the subepidermal connective tissue of the nipple and in the mammary parenchyma, some of the fibres being in close contact with blood vessels. Occasionally, a few delicate varicose fibres are associated with secretory alveoli and lactiferous ducts. The majority of PACAP-positive nerve fibres are, however, located in the glabrous skin of the nipple and the hairy skin adjacent to the nipple forming a subepithelial plexus from which delicate varicose nerve fibres enter the overlying epithelium. Double immunostaining for PACAP and a marker for sensory neurons, calcitonin gene-related peptide, has disclosed that the two peptides are almost completely co-localized. A minor population of the PACAP-immunoreactive nerve fibres shows co-existence with VIP. Although no obvious changes at the immunohistochemical level could be observed during pregnancy or lactation, elevated concentrations of immunoreactive PACAP-38 in mammary extracts have been found during lactation. Our data suggest that PACAP is involved in the nervous control of mammary gland function, probably in the transmission of suckling stimuli.     

Pituitary adenylate cyclase activating polypeptide (PACAP) reduces food intake in mice

Pituitary adenylate cyclase activating peptide (PACAP) is a peptide that is present in the hypothalamus and other areas of the rat brain. This study demonstrates that PACAP reduces food intake after intracerebroventricular injection in food-deprived mice. Behavioral analysis suggests that this decrease in food intake is, in part, compensated for by an increase in other behaviors. Pituitary adenylate cyclase activating peptide also was demonstrated to antagonize increased food intake resulting from administration of neuropeptide Y. Thus, PACAP joins a growing list of neuropeptides involved in the central regulation of food intake.     

Pituitary adenylate cyclase activating peptide (PACAP) in guinea-pig lung: distribution and dilatory effects.

The lower airways of guinea-pigs were analyzed for pituitary adenylate cyclase activating peptide (PACAP) using immunocytochemistry. In the trachea a moderate supply of PACAP-immunoreactive nerve fibers occurred around smooth muscle bundles, glands and small blood vessels. In the lung, PACAP-immunoreactive nerve fibers were distributed around small glands and bronchi. A rich supply of PACAP immunoreactive nerve fibers was found around blood vessels in the lungs. PACAP-suppressed smooth muscle responses were analysed using isolated circular segments of trachea, pulmonary arteries and aorta of guinea-pigs. In both airways and arteries PACAP caused a concentration-dependent relaxation of precontracted segments. The maximal relaxation effects were more pronounced in the airways than in the arteries while the order of potency was aorta greater than pulmonary artery greater than trachea. The effect of PACAP was compared to those of acetylcholine (ACh) and vasoactive intestinal peptide (VIP). In the pulmonary artery the vasomotor responses expressed as maximal dilatation had the order: ACh greater than VIP = PACAP while the order of potency was PACAP = VIP greater than ACh. In the trachea, PACAP was slightly more potent than VIP. The relaxatory responses to PACAP in the trachea and the intrapulmonary arteries were unaffected by pretreatment with atropine, prazosin, yohimbine, propranolol, mepyramine, cimetidine and Spantide. Removal of the endothelium abolished PACAP-induced vascular relaxation. Conceivably, PACAP-containing nerve fibers play a role in the regulation of airway resistance and local blood flow.     

Pituitary adenylate cyclase activating polypeptide (PACAP): discovery and current status of research.

For last 2 years since PACAP was first discovered, many important findings on PACAP have been reported. cDNAs encoding the precursor proteins of PACAP in sheep, human and rat were cloned, and the precursor proteins characterized. PACAP was found in a high concentration in the central nervous system, adrenal medulla and testis. Immunohistochemical study indicated that PACAP containing neural fibers are present throughout the brain, including both internal and external zones of the median eminence. In the hypothalamus many PACAP positive cell bodies were demonstrated in the supraoptic nucleus and the paraventricular nucleus in various species. Four types of high affinity PACAP receptor were demonstrated. PACAP receptors in the central nervous system, pituitary, adrenal medulla and germ cells of the testis are highly specific for PACAP, and not shared with VIP. The PACAP receptor was solubilized and cross-linking of 125I-PACAP27 with the binding protein suggest that the molecular weight of the receptor is around 57,000. Various biological actions of PACAP were reported, but the physiological cellular events linked with PACAP-induced activation of adenylate cyclase remain to be investigated.     

Pituitary adenylate cyclase activating polypeptide (PACAP) is present in human and cat gastric glands.

In the present work we have studied the occurrence of pituitary adenylate cyclase activating polypeptide (PACAP) in human and cat stomach mucosa using immunohistochemistry. As seen under a light microscope, there were many large rounded and ovoid cells that were PACAP immunopositive, mainly in the neck of the gastric glands of both species. The immunopositive material was predominant in the perinuclear area. The PACAP immunolabeling was specific because the preincubation of the antiserum with PACAP abolished the immunostaining. In human samples under electron microscope, the PACAP immunoreactive cells have shown the characteristics of parietal cells. In faintly stained cells, the localization of DAB reaction product was associated with the surface of the intracellular canaliculi. Cell labeling could not be observed besides parietal cells.     

Pituitary adenylate cyclase-activating peptide (PACAP) immunolocalization in lymphoid tissues of the rat

Pituitary adenylate cyclase activating peptide (PACAP) is a novel peptide isolated from the ovine hypothalamus. PACAP exists in 2 molecular forms with 27 (PACAP27) or 38 (PACAP38) amino acid residues. PACAP localization was studied by immunohistochemical methods in central (bone marrow and thymus) and peripheral (spleen, lymph nodes and duodenal mucosa) lymphoid tissues with antisera raised against PACAP27 or PACAP38. PACAP-positive cells were found in all lymphoid tissues examined. These cells were highly positive for PACAP38 but were negative for PACAP27. Morphologically, they were small mononuclear cells with relatively scarce cytoplasm and lymphocyte-like features. PACAP38-positive cells were abundant in peripheral lymphoid tissues (i.e., mesenteric lymph nodes). In the duodenal mucosa, PACAP38-positive cells were located either in the lamina propria or epithelium. These results suggest that PACAP38-positive cells are present within lymphoid tissues and may represent a lymphocyte-like cell subpopulation that has a potential role in cell-to-cell interactions in the immune system and in the integrated communication between neuroendocrine and immune systems.     

Pituitary adenylate cyclase activating polypeptide (PACAP) in the gastrointestinal tract of the rat: distribution and effects of capsaicin or denervation

The expression of pituitary adenylate cyclase activating polypeptide (PACAP) was studied in the gastrointestinal tract (GI-tract) of normal rats using radioimmunoassay, chromatography, immunocytochemistry, and in situ hybridization. PACAP-38, PACAP-27, and PACAP-related peptide were demonstrated in all parts of the GI-tract, PACAP-38 being the predominant form confirmed by chromatography. PACAP-immunoreactive nerve fibers and nerve cell bodies were found in the myenteric ganglia throughout the GI-tract. PACAP-containing nerve cell bodies were also demonstrated in the submucous ganglia of the small and large intestine. The synthesis of PACAP in intrinsic neurons was confirmed by in situ hybridization. Double immunostaining showed that PACAP is present in calcitonin gene-related peptide-containing sensory nerve fibers as well as in vasoactive intestinal polypeptide (VIP)- or VIP/gastrin-releasing peptide (GRP)-containing (intramural) nerve fibers in the upper GI-tract and in anally projecting, intrinsic VIP-and VIP/nitric oxide syntase-containing nerve cell bodies and nerve fibers in the small and large intestine. Neonatal treatment with capsaicin significantly reduced the concentration of PACAP-38 in the esophagus, stomach, and colon. Extrinsic denervation decreased the PACAP-38 concentration in the stomach, while no change was observed in the small intestine. These results indicate that PACAP- immunoreactive nerve fibers in the GI-tract originate from both intrinsic (enteric) and extrinsic (presumably sensory) sources suggesting that PACAP may have diverse gastrointestinal functions.     

Investigation of pituitary adenylate cyclase activating polypeptide (PACAP) in human amniotic fluid samples

Pituitary adenylate cyclase activating polypeptide (PACAP) is a neuropeptide acting as a hormone, a neuromodulator, a neurotransmitter, a trophic factor and is involved in a variety of developmental and regenerative processes. PACAP is present in several human tissues and biological fluids. In many pathological conditions, changes in PACAP levels have been described to reflect disease progression, therefore PACAP has diagnostic value as a potential biomarker. Since PACAP has been shown to play an important role in reproductive physiology and development, it was of interest to examine whether this neuropeptide occurs in the human amniotic fluid. Amniotic fluid samples were collected between the 15-19^th weeks of gestation from volunteering pregnant women undergoing amniocentesis as a prenatal diagnostic tool due to maternal age. Pathological cases were excluded after prenatal karyotype analysis. PACAP-like immunoreactivity was measured by radioimmunoassay and could be detected in all samples. The present study provides evidence for the presence of PACAP in human amniotic fluid, but determination of the exact physiological or pathological significance awaits further investigation.     

垂体腺苷酸环化酶激活肽的发现及研究进展

垂体腺苷酸环化酶激活肽是最初在绵羊下丘脑发现的一种新的具有多种生物活性的多肽。它广泛分布于中枢神经系统、周围神经系统以及非神经组织内。此外,它在某些类型细胞的旁分泌和自分泌主财节中也发挥作用。     

Properties and distribution of receptors for pituitary adenylate cyclase activating peptide (PACAP) in rat brain and spinal cord.

A high density (in the pmol/mg protein range) of specific functional receptors for PACAP (pituitary adenylate cyclase activating polypeptide) was observed in membranes from rat brain cortex, olfactory bulb, hypothalamus, hippocampus, striatum, cerebellum, pons and cervico-dorsal spinal cord, using [125I]PACAP-27 (PACAP 1-27). The tracer bound rapidly, specifically and reversibly. Competition binding curves were compatible with the coexistence, in the eight central nervous areas explored, of high and low affinity binding sites for PACAP-27 (Kd of 0.2 nM and 3.0 nM, respectively), and of only one class of binding sites for PACAP-38 (PACAP (1-38), Kd 0.2-0.9 nM). VIP inhibited only partially the binding of [125I]PACAP-27, and PHI, GRF(1-29)NH2 and secretin were ineffective at 1 microM. Chemical [125I]PACAP-27 cross-linking revealed a single specific 64 kDa protein species. In rat brain cortical membranes, saturation and competition experiments, using [125I]PACAP-38 as radioligand, indicated the presence of both high (Kd 0.13 nM) and low (Kd 8-10 nM) affinity binding sites for PACAP-38 and of low affinity (Kd 30 nM) binding sites for PACAP-27. These data taken collectively suggest the coexistence of PACAP-A receptors with a slight preference for PACAP-27 over PACAP-38 and of PACAP-B receptors that recognize PACAP-38 with a high affinity and PACAP-27 with low affinity. Both PACAP-27 and PACAP-38 stimulated adenylate cyclase with similar potency and efficacy. VIP was markedly less potent in this respect and also less efficient, except on cerebellar membranes.     

Pituitary adenylate cyclase-activating peptide (PACAP), a new vasoactive intestinal peptide (VIP)-like peptide in the respiratory tract

Summary Pituitary adenylate cyclase-activating peptide (PACAP) is a vasoactive intestinal peptide (VIP)-like peptide recently isolated from ovine hypothalami. Nerve fibers displaying PACAP immunoreactivity were found in the respiratory tract of rats, guinea pigs, ferrets, pigs, sheep and squirrel monkeys. A moderate supply of PACAP-immunoreactive fibers was seen in the nasal mucosa of guinea pigs. Few to moderate numbers of PACAP-containing fibers occurred in the tracheo-bronchial wall of rats, guinea pigs, ferrets, pigs, sheep and squirrel monkeys. The fibers were distributed beneath the epithelium, around blood vessels and seromucous glands, and among bundles of smooth muscle. In the lungs, the immunoreactive fibers were observed close to small bronchioli. A few PACAP-immunoreactive nerve cell bodies were seen in the sphenopalatine and otic ganglia of guinea pigs. Simultaneous double immunostaining of the respiratory tract of sheep and ferrets revealed that all PACAP-containing nerve fibers stored VIP. We suggest that neuronal PACAP may take part in the regulation of smooth muscle tone and glandular secretion.     

Pituitary adenylate cyclase activating polypeptide is expressed in autonomic neurons

Pituitary adenylate cyclase activating polypeptide (PACAP) is a novel vasoactive intestinal peptide (VIP)-like peptide, which is present in neuronal elements of several peripheral organs, and thus a putative neurotransmitter/modulator. In the present study, the expression of PACAP in two parasympathetic ganglia (otic, sphenopalatine) and one mixed parasympathetic/sensory ganglion (jugular-nodose) in rat was characterized by use of in situ hybridization and immunocytochemistry and compared to that of VIP and calcitonin gene-related peptide (CGRP). PACAP and VIP were expressed in virtually all nerve cell bodies in the otic and sphenopalatine ganglia; PACAP and VIP were also expressed in subpopulations of nerve cell bodies in the jugular-nodose ganglion. CGRP was expressed in numerous nerve cell bodies in the jugular-nodose ganglion and in a few, scattered, nerve cell bodies in the sphenopalatine ganglion. In the otic and sphenopalatine ganglia, PACAP- and VIP-like immunoreactivities were frequently co-localized; in the jugular-nodose ganglion, PACAP-like immunoreactivity was frequently co-localized with CGRP-like immunoreactivity in presumably sensory neurons and to a lesser extent with VIP in parasympathetic neurons. Thus, PACAP is synthesized and stored in autonomic parasympathetic neurons as well as in vagal sensory neurons, which provides an anatomical basis for the diverse effects of PACAP previously described.     

Anti-shock effect of pituitary adenylate cyclase activating polypeptide (PACAP) on experimental endotoxin shock in dogs

Effects of pituitary adenylate cyclase activating polypeptide with 38 amino acid residues (PACAP-38) on both cardiovascular functions and plasma hormone levels during endotoxin shock were studied in anesthesized dogs. When PACAP-38 (a bolus 420 pmol/kg injection or a bolus 420 pmol/kg injection followed by a continuous 30 pmol/kg/min infusion for 60 min) was administered intravenously 5 min after application of endotoxin, both mean arterial pressure and cardiac output were restored at 10 min. The continuous administration of PACAP-38 was more effective in improving the symptoms of shock. Plasma adrenalin and cortisol levels were significantly increased by both regimens. These results clearly indicate that the anti-shock properties of PACAP-38 may be attributed to its abilities to increase plasma cortisol and adrenalin levels and to stimulate cardiac function.     

Demonstration of specific binding sites for pituitary adenylate cyclase activating polypeptide (PACAP) in rat astrocytes

The high and low affinity binding sites for PACAP were identified in rat astrocytes using [125I]PACAP27 as the labeled ligand. Scatchard analysis of displacement of the bound tracer by unlabeled PACAP27 indicated the existence of two classes of binding sites, with the dissociation constant (Kd) = 1.22 +/- 0.4 nM, the binding maximal capacity (Bmax) = 821 +/- 218 fmols/mg protein for the high affinity binding site, and Kd = 0.59 +/- 0.06 microM, Bmax = 563 +/- 12 pmols/mg protein for the low affinity binding site, respectively. The specificity of [125I]PACAP27 binding was tested using PACAP38 and peptides structurally related to PACAP, such as VIP, GHRF, PHI, secretin and glucagon. PACAP38 completely displaced the binding of [125I]PACAP27 and Scatchard analysis also indicated the presence of two classes of binding sites with similar Kd and Bmax to those for PACAP27. VIP and GHRF competed with [125I]PACAP27, but to a much lesser extent than unlabeled PACAP27 in binding. Other peptides tested did not displace the binding of [125I]PACAP27 at 10(-6) M.     

Pituitary adenylate cyclase-activating polypeptide (PACAP) regulates the expression of PACAP in cultured tilapia astrocytes

Neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP) is a pleiotropic hormone that is involved in numerous physiologic functions. The present study examines the presence and the functional significance of PACAP and its receptor in the brain and astrocytes of tilapia (Oreochromis mossambicus). This is the first demonstration of the full-length nucleotide sequence of tPACAP gene in tilapia pituitary, brain, and cultured astrocytes. Two cDNA variants of the growth hormone-releasing hormone (GHRH)-PACAP gene were identified in tilapia pituitary, brain, and cultured astrocytes as a result of exon skipping with a long form (271 bp) encoding both tPACAP(38) and tGHRH and a short form (166 bp) encoding only tPACAP(38). The short form was found to be more abundant in astrocytes. Addition of ovine PACAP(38) (1 nM) to cultured astrocytes significantly stimulated the expression of tPACAP(38) at 4 hrs, but the effect dropped after 8 hrs of treatment. By contrast, the expression of PACAP type I receptor (PAC(1)-R) mRNA in the astrocytes was not responsive to PACAP(38) treatment. The tPACAP(38) expression also was activated by the cAMP analog, dibutyryl-cAMP, in a dose-dependent manner. Adding high salinity (170 mM NaCl, 500 mOsm/kg osmolarity) to cultured medium substantially increased astroglial tPACAP(38) expression over 4 hrs to a level that was maintained for 16 hrs. This observation was not found when mannitol (270 mM) was supplemented as an osmolarity-enhancing agent (500 mOsm/ kg). Taken together, tPACAP expression in tilapia astrocytes was well regulated by exogenous PACAP, cAMP, and salinity and might be involved in the adaptation to high salinity when the fish is in a seawater environment.     

The activation of adenylate cyclase by pituitary adenylate cyclase activating polypeptide (PACAP) via helodermin-preferring VIP receptors in human SUP-T1 lymphoblastic membranes.

Competition binding curves, using [125I-acetyl-His1]PACAP-27 as radioligand and dose-effect curves of adenylate cyclase activation in human SUP-T1 lymphoblastic membranes showed that PACAP-27 and PACAP-38 stimulate the enzyme through a single class of helodermin-preferring VIP receptors with the following order of potency: helodermin = [acetyl-His1]PACAP-27 greater than PACAP-38 greater than PACAP-27 greater than VIP. PACAP (6-27) (Ki 0.5-0.8 microM) and [Des-His1, Asn3]PACAP-27 (Ki 1-2 microM) acted as competitive antagonists. Using a series of 13 PACAP-27 analogues and fragments and three VIP analogues, we identified positions 1, 2, 3, 9 and 13 in PACAP-27 as being of importance for high-affinity binding. Thus, we added further evidence for considering that the present helodermin-preferring VIP receptors, when compared to a majority of VIP receptors and PACAP receptors, exhibit an original specificity pattern.