Distribution Of Pituitary Adenylate Cyclase-activating Polypeptide Research Articles

Protective effect of pituitary adenylate cyclase activating polypeptide in diabetic keratopathy

Diabetic keratopathy (DK) is the major complication of the cornea characterizing diabetes-affected patients. This ocular pathology is correlated with the hyperglycemic state leading to delayed corneal wound healing and recurrent corneal ulcers. Pituitary adenylate cyclase-activating polypeptide (PACAP) is a neuropeptide with widespread distribution throughout the body, and exerting cytoprotective effects in the neural and non-neuronal parts of the eye, including the cornea. The purpose of the present study was to investigate whether changes in PACAP expression can concur for delayed epithelial wound healing in diabetic cornea and whether the protective effect of the peptide could be mediated through the activation of the EGFR signaling pathway, which has been reported to be impaired in DK. Expression and distribution of PACAP, PAC1R, and EGFR were investigated through immunohistochemistry analysis in the cornea of normal and diabetic rats. The role of the peptide on wound healing during DK was evaluated in an in vitro model represented by rabbit corneal epithelial cells grown in high glucose conditions. Western blotting and immunofluorescence analysis were used to examine the ability of PACAP to induce the activation of the EGFR/ERK1/2 signaling pathway. Our results showed that in diabetic cornea the expression of PACAP, PAC1R, and EGFR is drastically reduced. The treatment with PACAP via PAC1R activation enhanced cell viability and corneal epithelium wound healing in cells grown under high glucose conditions. Furthermore, both EGFR and ERK1/2 signaling was induced upon the peptide treatment. Overall, our results showed the trophic efficiency of PACAP for enhancing the corneal epithelium re-epithelialization suggesting that the peptide could be beneficially valuable as a treatment for DK.

Distribution of PACAP and PAC1 Receptor in the Human Eye

Pituitary adenylate cyclase–activating polypeptide (PACAP) is a neuropeptide with widespread distribution and diverse biological functions. Several studies show that PACAP has strong cytoprotective effects mediated mostly through its specific PAC1 receptor (PAC1-R) and it plays important roles in several pathological conditions. Its distribution and altered expression are known in various human tissues, but there is no descriptive data about PACAP and its receptors in the human eyebulb. Since PACAP38 is the dominant form of the naturally occurring PACAP, our aim was to investigate the distribution of PACAP38-like immunoreactivity in the human eye and to describe the presence of PAC1-R. Semiquantitative evaluation was performed after routine histology and immunohistochemical labeling on human eye sections. Our results showed high level of immunopositivity in the corneal epithelium and endothelium. Within the vascular layer, the iris and the ciliary body had strong immunopositivity for both PACAP and PAC1-R. Several layers of the retina showed immunoreactivity for PACAP and PAC1-R, but the ganglion cell layer had a special pattern in the immunolabeling. Labeling was observed in the neuropil within the optic nerve in both cases and glial cells displayed immunoreactivity for PAC1-R. In summary, our study indicates the widespread occurrence of PACAP and its specific receptor in the human eye, implying that the results from in vitro and animal studies have translational value and most probably are also present in the human eye.

Localization of CGRP, CGRP receptor, PACAP and glutamate in trigeminal ganglion. Relation to the blood–brain barrier

Calcitonin gene-related peptide (CGRP) receptor antagonists have demonstrated anti-migraine efficacy. One remaining question is where do these blockers act? We hypothesized that the trigeminal ganglion could be one possible site. We examined the binding sites of a CGRP receptor antagonist (MK-3207) and related this to the expression of CGRP and its receptor in rhesus trigeminal ganglion. Pituitary adenylate cyclase-activating polypeptide (PACAP) and glutamate were examined and related to the CGRP system. Furthermore, we examined if the trigeminal ganglion is protected by the blood–brain barrier (BBB). Autoradiography was performed with [3H]MK-3207 to demonstrate receptor binding sites in rhesus trigeminal ganglion (TG). Immunofluorescence was used to correlate binding and the presence of CGRP and its receptor components, calcitonin receptor-like receptor (CLR) and receptor activity-modifying protein 1 (RAMP1), and the distribution of PACAP and glutamate in rhesus and rat TG. Evans blue was used to examine large molecule penetration into the rat TG. High receptor binding densities were found in rhesus TG. Immunofluorescence revealed expression of CGRP, CLR and RAMP1 in trigeminal cells. CGRP positive neurons expressed PACAP but not glutamate. Some neurons expressing CLR and RAMP1 co-localized with glutamate. Evans blue revealed that the TG is not protected by BBB. This study demonstrates CGRP receptor binding sites and expression of the CGRP receptor in rhesus and rat TG. The expression pattern of PACAP and glutamate suggests a possible interaction between the glutamatergic and CGRP system. In rat the TG is outside the BBB, suggesting that molecules do not need to be CNS-penetrant to block these receptors.

Distribution of pituitary adenylate cyclase-activating polypeptide (PACAP) in the human testis and in testicular germ cell tumors

Pituitary adenylate cyclase-activating peptide (PACAP) is a neuropeptide expressed in the central nervous system and peripheral organs. Previous studies revealed the role and distribution of PACAP in the rodent testis, however, its presence in the human testis and in testicular germ cell tumors is not known. We used RT-PCR and immunohistological observations to investigate whether human testicular tissue and testicular germ cell tumors contain PACAP. The mRNAs for PACAP and its receptors were detected in total RNA extracted from human testes. PACAP immunoreactivity was observed in spermatogonia and spermatids from normal testes. In contrast, diffuse PACAP immunopositivity was observed in seminoma tumor cells, while only faint immunoreactivity was observed in embryonal carcinoma cells. Our data suggest that PACAP may play a role in human spermatogenesis and in testicular germ cell tumor development.

Distribution and Molecular Evolution of the Neuropeptide Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) and its Receptors in the Lizard Podarcis sicula (Squamata, Lacertidae)

The presence of the pituitary adenylate cyclase-activating polypeptide (PACAP) and its receptors PAC(1), VPAC(1), and VPAC(2) was studied in the lizard Podarcis sicula gastrointestinal and respiratory tissues. The expression and distribution of this neuropeptide was investigated using RT-PCR, immunohistochemistry, and in situ hybridization techniques. RT-PCR showed that several tissues of this reptile synthesize an mRNA encoding for PACAP. Performing in situ hybridization and immunohistochemistry, we found a wide distribution of PACAP and its mRNA in intestine, stomach, liver, and lung. PACAP receptors possess a specific distribution in both gastrointestinal and respiratory system. Further, we analyzed the conservation of PACAP amino acid sequence demonstrating that this peptide in the lizard is very similar to that of other vertebrates. Our findings suggest that also in reptiles an effective PACAP system is present and that it could be implicated in some essential physiological functions as a result of its high conservation amongst vertebrates.

Localization, characterization and function of pituitary adenylate cyclase-activating polypeptide during brain development

Neural development is controlled by region-specific factors that regulate cell proliferation, migration and differentiation. Pituitary adenylate cyclase-activating polypeptide (PACAP) is a neuropeptide that exerts a wide range of effects on different cell types in the brain as early as the fetal stage. Here we review current knowledge concerning several aspects of PACAP expression in embryonic and neonatal neural tissue: (i) the distribution of PACAP and PACAP receptors mRNA in the developing brain; (ii) the characteristic generation of neurons, astrocytes and oligodendrocytes in brain areas where the PACAP receptor is expressed and (iii) the role of PACAP as a regulator of neural development, inducing differentiation and proliferation in association with other trophic factors or signal transduction molecules.

Expression of pituitary adenylate cyclase activating polypeptide and its type I receptor mRNAs in human placenta

Pituitary adenylate cyclase activating polypeptide (PACAP) was first isolated from ovine hypothalamus and was known to stimulate the release of growth factor in various cells. Recently, we reported the cellular localization of PACAP and its type I (PAC1) receptor in rat placenta during pregnancy. Placenta is a critical organ that synthesizes several growth factors and angiogenic factors for the fetal development and its own growth. However, there is little information regarding the cellular localization of PACAP and its receptor in human placenta at various gestations. The aim of the present study was to define the expression and distribution of PACAP and PAC1 receptor mRNAs in the human placenta during the pregnancy period. PACAP and PAC1 receptor mRNAs were expressed in stroma cells of stem villi and terminal villi. At the early stage, on 7 and 14 weeks, PACAP and PAC1 receptor genes were moderately expressed in stroma cells surrounding the blood vessels within stem villi. These genes were strongly expressed in stroma cells of stem villi and terminal villi on 24 and 38 weeks. The expression of these genes was increased as gestation advanced, and localized in the same areas. Localization of PACAP and PAC1 receptor demonstrate the evidence that PACAP may play an important role, as an autoregulator or pararegulator via its PAC1 receptor. In conclusion, our findings strongly suggest that PACAP may have a critical role in physiological function of the placenta for gestational maintenance and fetal growth.

Pituitary adenylate cyclase-activating polypeptide (PACAP)-like immunoreactivity in the brain of a teleost, Uranoscopus japonicus: immunohistochemical relationship between PACAP and adenohypophysial hormones

The neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP) could play a role in stimulating pituitary hormone release in fish brain. In this study, we used immunochemical techniques to examine the histological and quantitative distribution of PACAP in the central nervous system (CNS) of a teleost, the stargazer, Uranoscopus japonicus. In addition, high performance liquid chromatographic (HPLC) analysis was performed to characterize the form of PACAP present, while the relationship between PACAP and adenohypophysial hormones was also determined immunohistochemically. PACAP-like immunoreactive (LI) neuronal cell bodies and fibers were found not only in the hypothalamo-pituitary region but also in the midbrain and hindbrain regions. PACAP-LI fibers were identified in the neurohypophysis in close proximity to pituitary cells containing immunoreactive hormones such as somatolactin, the N-terminal peptide of proopiomelanocortin, and N-acetyl endorphin. The concentration of immunoreactive PACAP in whole brain tissue was approximately 300 pmol/g wet weight. The average concentrations of immunoreactive PACAP in regions of the telencephalon, diencephalon, tectum, cerebellum, and rhombencephalon were 217.53, 510.26, 83.30, 148.64, and 364.62 pmol/g, respectively. In reverse-phase HPLC experiments, the predominant form of immunoreactive PACAP eluted closely with synthetic stargazer PACAP38, while PACAP27-like immunoreactivity was negligible. These results suggest that PACAP38 is the predominant PACAP form in the stargazer CNS, and that PACAP acts not only as a hypophysiotropic factor for adenohypophysial hormone release but also as a neurotransmitter and neuromodulator in the CNS.

Pituitary adenylate cyclase-activating polypeptide does not colocalize with vasoactive intestinal polypeptide in the hypothalamic magnocellular nuclei and posterior pituitary of cats and rats.

Pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal polypeptide (VIP) immunoreactive cells were demonstrated in the hypothalamic magnocellular nuclei in cats and rats. In cats these immunoreactive cells were stained without any treatment or intervention; however, in rats we had to use the pituitary stalk section to enhance the amount of PACAP and VIP for successful immunostaining. In both species the regions occupied by PACAP and VIP immunoreactive cells partially overlap each other in the paraventricular and supraoptic nuclei. Nevertheless, in either cats or rats PACAP and VIP immunoreactivities do not colocalize in the same cells studied by double labeling immunohistochemistry (IHC) or by the combination of immunohistochemistry and in situ hybridization. As was expected, PACAP and VIP immunoreactive materials were stored in different fibers of the posterior pituitary where the distribution of PACAP and VIP fibers also showed different patterns: PACAP fibers form a dense plexus at the periphery of the posterior lobe, in the vicinity of the intermediate lobe; however, the VIP fibers were evenly distributed mainly in the center of the posterior lobe. In spite of the high sequence homology of PACAP and VIP, the two peptides are synthesized in different subpopulations of hypothalamic neurons. This different distribution correlates well with the different role of the hypothalamic PACAP and VIP in the biologic clock and in the functions of the anterior and posterior pituitary.

Expression patterns of pituitary adenylate cyclase activating polypeptide and its type I receptor mRNAs in the rat placenta.

Pituitary adenylate cyclase activating polypeptide (PACAP) was first isolated from ovine hypothalamus and is known to act as a tropic factor in various cells. Recent report revealed the expression of PACAP and the PACAP type I (PAC(1)) receptor in human and rat placentas at term. Placenta is a critical organ that synthesizes several growth and angiogenic factors for its own growth as well as fetal development. However, there is little information regarding the expression pattern and cellular localization of PACAP and PAC(1) during pregnancy. The aim of this study was to define the expression and distribution of PACAP and PAC(1) receptor mRNAs in the rat placenta during pregnancy. PACAP and PAC(1) receptor mRNAs were expressed in decidual cells, chorionic vessels, and stromal cells of the chorionic villi. Interestingly, the expression of these genes varied with the day of gestation. For example, PACAP and PAC(1) receptor mRNAs expressed in decidual cells on day 13.5 and 15.5, their expression was strong in chorionic vessels and stromal cells of the chorionic villi within the labyrinth zone on day 17.5, 19.5, and 21.5. In fact, as gestation advanced, the expression of PACAP and PAC(1) receptor mRNAs in the decidua cells disappeared, as their high expression became evident in the chorionic vessels and stromal cells of the chorionic villi. Our finding that PACAP and the PAC(1) receptor are co-localized and their genes seemingly co-regulated within specific placental areas, strongly suggest that this peptide may play an important role, as an autoregulator or pararegulator via its PAC(1) receptor, in physiological functioning of the placenta for gestational maintenance.

Distribution of pituitary adenylate cyclase‐activating polypeptide and pituitary adenylate cyclase‐activating polypeptide type I receptor mRNA in the chicken brain

To map in detail the brain areas in which pituitary adenylate cyclase-activating polypeptide (PACAP) may play a significant role in birds, the distribution of PACAP and PACAP type I receptor (PAC(1)-R) mRNA was examined throughout the entire chicken brain by using in situ hybridization histochemistry. Widespread distribution of both PACAP and its receptor mRNA was found. The telencephalic areas where the most intense signals for PACAP mRNA were found included the hyperstriatum accessorium, the hippocampus, and the archistriatum. In the diencephalon, a group of neurons that highly expressed PACAP mRNA was observed from the anterior medial hypothalamic nucleus to the inferior hypothalamic nucleus. Moderate expression was found in the paraventricular nucleus and the preoptic region. A second large group of neurons containing PACAP message was found within the nucleus dorsolateralis anterior thalami and extended caudally to the area around the nucleus ovoidalis and the nucleus paramedianus internus thalami. Furthermore, expression of PACAP message was observed within the bed nucleus of the pallial commissure, nucleus spiriformis medialis, optic tectum, cerebellar cortex, olfactory bulbs, and several nuclei within the brainstem (dorsal vagal and parabrachial complex, reticular formation). The highest expression of PAC(1)-R mRNA was found in the dorsal telencephalon, olfactory bulbs, lateral septum, optic tectum, cerebellum, and throughout the hypothalamus and thalamus. The presence of PACAP and PAC(1)-R mRNA in a variety of brain areas in birds suggests that PACAP mediates several physiologically important processes in addition to regulating the activity of the pituitary gland.

Alteration of PACAP distribution and PACAP receptor binding in the rat sensory nervous system following sciatic nerve transection

Pituitary adenylate cyclase activating polypeptide (PACAP) is a widely expressed neuropeptide that has been involved in nerve regeneration, neurone survival and nociception. In this study, the distribution of PACAP and PACAP-receptors were investigated in rat dorsal root ganglia (DRG), spinal cord and medulla oblongata at 3, 7 or 14 days following unilateral sciatic nerve transection using immunohistochemistry, 125 I -PACAP-binding and in situ hybridisation. In control (contralateral side) DRG, about 30% of the nerve cell bodies (92% being small) were PACAP-immunoreactive (PACAP-IR). In the spinal cord, PACAP-IR fibres were seen in laminae I–II but not in the gracile nuclei. Following sciatic nerve transection, PACAP-IR fibres appeared in the gracile nuclei and occasionally in the deeper laminae of the dorsal horn consistent with the relative increase in larger PACAP-IR DRG neurones. However, the relative number of small PACAR-IR neurones was significantly lower on the transected side as compared to the control side suggesting a dual reaction for PACAP in the DRG following nerve injury. 125 I -PACAP-binding was found in laminae I–II, around the central canal and in the gracile nuclei but not in the DRG. At 14 days after transection, 125 I -PACAP-binding density was significantly reduced in the ipsilateral dorsal horn. PACAP-receptor (PAC 1) mRNA was detected in neurones of the dorsal and ventral horn and in the gracile nuclei with no overt changes observed after transection. Very few DRG nerve cell bodies contained PAC 1 mRNA. The findings are consistent with a role for PACAP both in nociception and regeneration.

LOCALIZATION AND DISTRIBUTION OF PITUITARY ADENYLATE CYCLASE-ACTIVATING POLYPEPTIDE IN THE RAT EPIDIDYMIS

Previous investigation has provided evidence for the control of electrogenic chloride secretion by pituitary adenylate cyclase-activating polypeptide (PACAP) across the rat epididymal epithelium using electrophysiological measurement of transepithelial transport in cultured epididymal system. Hence, it suggests that epididymal and sperm functions are subject to control by a local PACAP system in the rat epididymis. In the present study, localization and distribution of PACAP in the rat epididymal duct was studied by an indirect immunofluorescence technique in conjunction with confocal laser scanning microscopy. Immunoreactivity for PACAP was found in all regions of the epididymal duct. However, the intensity of immunoreactivity for PACAP was stronger in the caput and corpus regions when compared to that of the cauda epididymidis. Much weaker immunostaining for PACAP, as compared to those found in other regions, was observed in the cauda epididymidal tubules which are in close proximity to the vas deferens. No immunoreactivity for PACAP was found in epididymal spermatozoa. Together with the previous finding, the present results suggest that PACAP may exhibit a regional difference in its expression along the epididymal duct and it may act in a paracrine or autocrine fashion in the regulation of epididymal chloride secretion and hence fluid secretion, thus regulating epididymal and sperm functions along the epididymal duct.

Distribution of PACAP (pituitary adenylate cyclase-activating polypeptide)-like and helospectin-like peptides in the teleost gut

Pituitary adenylate cyclase-activating polypeptide (PACAP) and helospectin are two vasoactive intestinal polypeptide (VIP)-related neuropeptides that have recently been demonstrated in the mammalian gut; the aim of this study was to reveal their occurrence and localisation in the gastrointestinal tract, swimbladder, urinary bladder and the vagal innervation of the gut of teleosts, using immunohistochemical methods on whole-mounts and sections of these tissues from the Atlantic cod, Gadus morhua and the rainbow trout, Oncorhynchus mykiss. Both PACAP-like and helospectin-like peptides were present in the gut wall of the two species. Immunoreactive nerve fibres were found in all layers but were most frequent in the myenteric plexus and along the circular muscle fibres. Immunoreactivity was also demonstrated in nerves innervating the swimbladder wall, the urinary bladder and blood vessels to the gut. Immunoreactive nerve cell bodies were found in the myenteric plexus of the gut and in the muscularis mucosae of the swimbladder. In the vagus nerve, non-immunoreactive nerve cells were surrounded by PACAP-immunoreactive fibres. Double staining revealed the coexistence of PACAP-like and helospectin-like peptides with VIP in all visualized nerve fibres and in some endocrine cells. It is concluded that PACAP-like and helospectin-like peptides coexist with VIP in nerves innervating the gut of two teleost species. The distribution suggests that both PACAP and helospectin, like VIP, are involved in the control of gut motility and secretion.

Distribution of PACAP (pituitary adenylate cyclase-activating polypeptide)-like and helospectin-like peptides in the teleost gut

Distribution of PACAP (pituitary adenylate cyclase-activating polypeptide)-like and helospectin-like peptides in the teleost gut

A novel neuropeptide, pituitary adenylate cyclase-activating polypeptide (PACAP), in human intestine: evidence for reduced content in Hirschsprung's disease.

A novel neuropeptide, pituitary adenylate cyclase-activating polypeptide (PACAP), exhibits sequence homology with vasoactive intestinal polypeptide (VIP) and occurs in the mammalian brain, lung and gut. The distribution of PACAP in ganglionic and aganglionic portions of the large intestine of patients with Hirschsprung's disease was examined by immunohistochemistry and radioimmunoassay. PACAP-immunoreactive nerve fibers were distributed in all layers of the ganglionic and aganglionic segments of the intestine, although they were less numerous in the latter, and PACAP-immunoreactive nerve cell bodies were seen in the ganglionic portion of the intestine. The concentration of immunoreactive PACAP was lower in the aganglionic than in the ganglionic segment of the intestinal wall. PACAP and VIP were found to coexist in both ganglionic and aganglionic segments of the intestine. Apparently, PACAP participates in the regulation of gut motility. The scarcer PACAP innervation of the aganglionic segment may contribute to the defect in intestinal relaxation seen in patients with Hirschsprung's disease.