PACAP stimulation of maturational gonadotropin secretion in goldfish involves extracellular signal-regulated kinase, but not nitric oxide or guanylate cyclase, signaling

In goldfish, nitric oxide synthase (NOS) immunoreactivity is present in gonadotropes and extracellular signal-regulated protein kinase (ERK) mediates GnRH stimulation of gonadotropin release and synthesis. In this study, we tested the possible involvement of nitric oxide (NO) and ERK in mediating PACAP-stimulated maturational gonadotropin (GTH-II) release from primary cultures of dispersed goldfish pituitary cells. In static incubation experiments, PACAP-induced GTH-II release was unaffected by two inhibitors of NOS synthase, AGH and 1400W; whereas addition of a NO donor, SNAP, elevated GTH-II secretion. In perifusion experiments, neither NOS inhibitors (AGH, 1400W and 7-Ni) nor NO scavengers (PTIO and rutin hydrate) attenuated the GTH-II response to pulse applications of PACAP. In addition, the GTH-II responses to PACAP and the NO donor SNP were additive while PTIO blocked SNP action. Although dibutyryl cGMP increased GTH-II secretion in static incubation, inhibition of guanylate cyclase (GC), a known down-stream target for NO signaling, did not reduce the GTH-II response to pulse application of PACAP. On the other hand, GTH-II responses to PACAP in perifusion were attenuated in the presence of two inhibitors of ERK kinase (MEK), U 0126 and PD 98059. These results suggest that although increased availability of NO and cGMP can lead to increased GTH-II secretion, MEK/ERK signaling, rather than NOS/NO/GC activation, mediates PACAP action on GTH-II release in goldfish.     

Ghrelin-induced growth hormone release from goldfish pituitary cells is nitric oxide dependent

Ghrelin (GRLN) is an important neuroendocrine regulator of growth hormone (GH) release in vertebrates. Previous studies show goldfish (g)GRLN₁₉-induced GH from the goldfish pituitary involves voltage sensitive Ca²⁺ channels, increases in intracellular Ca²⁺ and the PKC signalling pathway. We set out to examine the role of the nitric oxide (NO) pathway in gGLRN₁₉-induced GH release from primary cultures of goldfish pituitary cells using pharmacological regulators in cell column perifusion systems. The NO scavenger PTIO abolished gGRLN₁₉-induced GH release and co-treatment with the NO donor SNP and GRLN did not produce additive GH release responses. Nitric oxide synthase (NOS) inhibitors 1400 W and 7-Ni abolished GRLN-induced GH release while treatment with another NOS inhibitor, AGH, had no significant effect. Taken together, these results demonstrate that the NOS/NO is an integral component of gGRLN₁₉-induced signalling within the goldfish pituitary cells, and given the relative specificity of AGH for inducible NOS and endothelial NOS isoforms, suggests that neuronal NOS is the likely NOS isoform utilized in goldfish somatotropes by this physiological regulator.     

Evidence that nitric oxide is involved in the regulation of growth hormone secretion in goldfish

Whether nitric oxide (NO) plays a role in regulation of growth hormone (GH) secretion from somatotropes in the pituitary of the goldfish Carassius auratus was investigated. Immunocytochemistry with two antibodies against mammalian NO synthase (NOS) revealed the presence of a NOS-like enzyme in primary cultures of dispersed goldfish pituitary cells, including morphologically identified somatotropes. NO donors S-nitroso-N-acetylpenicillamine and sodium nitroprusside (SNP), as well as a cyclic guanosine monophosphate analogue (dibutyryl guanosine 3':5'-cyclic monophosphate), all significantly increased GH secretion from dispersed goldfish pituitary cells in static culture. Somatostatin abolished the response to SNP, and NOS inhibitors aminoguanadine hemisulfate (AGH) and N-(3-aminomethyl)benzylacetamidine, dihydrochloride (1400W) decreased the GH release response to known neuroendocrine factors stimulatory to GH release (gonadotropin-releasing hormone and a dopamine D1 agonist). AGH and 1400W did not alter basal GH secretion. These data suggest that NO plays a role in mediating the GH response to endogenous neuroendocrine factors in goldfish.     

Role of Ca2+ stores in dopamine- and PACAP-evoked growth hormone release in goldfish

Growth hormone (GH) secretion, evoked by either pituitary adenylate cyclase-activating polypeptide (PACAP) or dopamine (DA), is dependent on both voltage-sensitive calcium channels (VSCC) and cAMP signaling in goldfish. We further characterized the involvement of Ca2+ in evoked release by PACAP and DA, by examining the sensitivity of evoked GH release to perturbations of Ca2+ signaling. Both VSCC and calmodulin/calmodulin-dependent kinase are involved in PACAP signaling as had been shown for DA. In spite of this apparent dependence on VSCC, blockade of TMB-8 but not ryanodine-sensitive intracellular Ca2+ stores inhibited both PACAP- and DA-evoked GH release. Using sarcoplasmic/endoplasmic reticulum Ca-ATPases (SERCA) inhibitors, we found BHQ blocked, whereas thapsigargin (Tg) enhanced stimulated GH release, suggesting that Tg-sensitive SERCA may counteract these cAMP-mobilizing neuroendocrine regulators by sequestering [Ca2+]i. As GH secretion stimulated by two endogenous gonadotropin-releasing hormones is not affected by Tg, it appears that distinct multiple Ca2+ stores mediate the hormone releasing response to different neuroendocrine regulators.     

Goldfish brain somatostatin-28 differentially affects dopamine- and pituitary adenylate cyclase-activating polypeptide-induced GH release and Ca(2+) and cAMP signals

Dopamine (DA) and pituitary adenylate cyclase-activating polypeptide (PACAP) stimulate goldfish growth hormone (GH) release via cAMP- and Ca(2+)-dependent pathways while DA also utilizes NO. In this study, identified goldfish somatotropes responded to sequential applications of PACAP and the DA D1 agonist SKF38393 with increased intracellular Ca(2+) levels ([Ca(2+)](i)), indicating that PACAP and DA D1 receptors were present on the same cell. A native goldfish brain somatostatin (gbSS-28) reduced SKF38393-stimulated cAMP production and PACAP- and NO donor-elicited GH and [Ca(2+)](i) increases, but not PACAP-induced cAMP production nor the GH and [Ca(2+)](i) responses to forskolin, 8-bromo-cAMP and SKF38393. gbSS-28 might inhibit PACAP-induced GH release by interfering with PACAP's ability to increase [Ca(2+)](i) in a non-cAMP-dependent manner. However, DA D1 receptor activation bypassed gbSS-28 inhibitory effects on cAMP production and NO actions via unknown mechanisms to maintain a normal [Ca(2+)](i) response leading to unhampered GH release.     

Pituitary adenylate cyclase-activating polypeptide releases 7B2, adrenocorticotrophin, growth hormone and prolactin from the mouse and rat clonal pituitary cell lines AtT-20 and GH3.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a neuropeptide originally isolated from ovine hypothalami and so called because of its ability to stimulate pituitary adenylate cyclase activity. Alternative amidation and proteolytic processing of prepro-PACAP gives rise to two bioactive-amidated forms, PACAP-NH2(1-38) (PACAP-38) and PACAP-NH2(1-27) (PACAP-27). 7B2 is a polypeptide of 185 amino acids which is predominantly found in secretory granules and is widely distributed in rat and human tissues. We investigated the ability of the two forms of PACAP to stimulate GH, prolactin and 7B2 release by the rat pituitary clonal cell line GH3, and ACTH and 7B2 by the mouse pituitary clonal cell line AtT-20. PACAP-38 and PACAP-27 stimulated 7B2 and GH/prolactin or ACTH secretion with a similar efficacy over the 2-h incubation period from GH3 and AtT-20 cells respectively. 7B2 secretion was also stimulated by corticotrophin-releasing factor (CRF-41) and vasoactive intestinal polypeptide (VIP) in AtT-20 cells, and thyrotrophin-releasing hormone (TRH) and VIP in GH3 cells. Addition of PACAP to CRF-41 resulted in an additive effect on ACTH secretion and a synergistic effect on 7B2 secretion in AtT-20 cells. No synergism was observed when PACAP was added together with TRH, either on GH and prolactin secretion or on 7B2 release from GH3 cells. PACAP-mediated 7B2 secretion from both cell lines and PACAP-stimulated ACTH release from AtT-20 cells were reduced by 5 mg octapeptide synthetic somatostatin analogue/l (5 mg SMS 201-995/l).     

Leptin induces growth hormone secretion from peripheral blood mononuclear cells via a protein kinase C- and nitric oxide-dependent mechanism

Leptin is a key mediator of signals regulating food intake and energy expenditure and exerts potent immunomodulatory effects. We investigated the mechanisms mediating the action of leptin on GH secretion from peripheral blood mononuclear cells (PBMCs). Using immunofluorescence microscopy, we demonstrated a polarized expression pattern of leptin receptor protein on the surface of mononuclear cells and constitutive expression of GH in PBMCs. Leptin exhibited a dose-dependent stimulatory effect on GH secretion by PBMCs and also up-regulated the GH receptor gene expression. We did not observe any additive effects of leptin on GH secretion upon activation of cells with the plant mitogen phytohemagglutinin, unlike leptin, phytohemagglutinin exerted no effect on GH receptor mRNA expression. Leptin led to a nitric oxide (NO) synthase (NOS)-specific, dose-dependent increase in NO production from PBMCs because leptin-induced NO release was blocked by the addition of the NOS inhibitor Nomega-Nitro-l-arginine methyl ester and protein kinase C (PKC) inhibitor calphostin C. This leptin-induced GH secretion was dependent on both PKC and NO activation because the addition of PKC and NOS inhibitors inhibited leptin-induced GH production. Although the addition of sodium nitroprusside, a spontaneous liberator of NO, stimulated GH release from PBMCs, leptin had no additive or synergistic effect on sodium nitroprusside-induced GH production. Together, these findings demonstrate a unique action of leptin on immune cells via its ability to stimulate the GH production by blood mononuclear cells via PKC- and NO-dependent pathways. These data also support a probable role for local immune-derived GH in mediating some of the pleiotropic actions of leptin.     

Nitric oxide stimulates growth hormone secretion from human fetal pituitaries and cultured pituitary adenomas

Nitric oxide (NO), a highly reactive free radical, has been identified as a neurotransmitter in the central and peripheral nervous system. NO synthase (NOS) is the enzyme responsible for NO production from L-arginine and plays an important role in regulating the release of several hypothalamic peptides. In the pituitary, NO was found to increase growth hormone (GH) secretion in several in vitro and in vivomodels. However, its role in human GH regulation is unknown. The aim of this study was to investigate the regulatory effects of NO on human GH and prolactin secretion using primary cell cultures of human fetal pituitaries and cultured hormone-secreting adenomas. Incubation of the human fetal pituitaries (21-24 wk gestation) in the presence of sodium nitroprusside (SNP; 1 mM), a NO donor, for 4 h resulted in a 50-75% increase in GH secretion, similar to the stimulatory effect evoked by growth hormone-releasing hormone (GHRH) (10 nM). However, fetal PRL secretion was not affected by SNP. GH release was also stimulated (40-70% increase) by SNP in 60% of the cultured GH-secreting adenomas studied. SNP-induced GH release was inhibited in both fetal and adenomatous cells by PTI0, a NO scavenger. The addition of cGMP (0.1-1 mM), the second messenger of multiple NO actions, enhanced fetal and adenomatous GH secretion by 55-95%. Neuronal NOS (nNOS) was expressed in normal (fetal and adult) human pituitary tissues and in GH-secreting adenomas. Examination of its functional expression using L-arginine (1 microM) yielded a 35% increase in GH release from cultured GH-secreting adenoma. This response was blocked by a NOS inhibitor with high selectivity for the neuronal enzyme and by a guanylyl cyclase inhibitor. In conclusion, NO stimulates human GH in cultured fetal pituitaries and GH-secreting adenomas. Cyclic GMP is probably involved in this hormonal regulation.     

Endogenous nitric oxide inhibits growth hormone secretion through cyclic guanosine monophosphate-dependent mechanisms in GH3 cells

Constitutive nitric oxide synthase (NOS) is expressed in rat adenohypophysis and clonal GH3 cells. The mechanisms of action of nitric oxide (NO) to inhibit hormone secretion and the possible role of (6R)-5, 6, 7, 8-tetrahydro-L-biopterin (THB) in the action of endogenous NO were studied in GH3 cells. Inhibiting NOS with N(G)-nitro-L-arginine or trapping NO with oxyhemoglobin enhanced both the basal and TRH-stimulated rat GH release. Sodium nitroprusside did not further decrease either the basal or the TRH-stimulated GH secretion, suggesting that endogenous NO exerted the maximal inhibitory effect. Inhibition of de novo synthesis of THB increased GH secretion. A cyclic guanosine-monophosphate (cGMP) antagonist did not increase the basal GH secretion but enhanced TRH-induced GH release. These findings suggest that endogenous NO plays an inhibitory role on basal GH release and TRH-stimulated hormone release from GH3 cells in an autocrine or paracrine fashion, at least partly, through a cGMP-dependent pathway. It is also suggested that endogenous THB plays a role in NO production and subsequent inhibition of hormone secretion in GH3 cells.     

The role of nitric oxide in the control of basal and LHRH-stimulated LH secretion.

The gaseous transmitter nitric oxide (NO) appears to be involved in the control of LH secretion and in the modulation of LH responses after stimulation with luteinizing hormone releasing hormone (LHRH), excitatory amino acids (EAAs) and leptin. The regulatory action of NO in the control of LH secretion includes modulation of LHRH release, changes in hypothalamic-pituitary blood flow and direct effects at pituitary level. To determine the net balance of these actions we evaluated (1) the effects of systemic administration of sodium nitroprusside (SNP, a NO donor) and Nw-nitro-L-arginine methyl ester (NAME, a blocker of NO synthase) on basal and LHRH-stimulated LH secretion in intact and ovariectomized females; and (2) the effects of SNP and NAME on LH secreted by dispersed pituitary cells. Finally, since NO is involved in the stimulatory effect of excitatory amino acids (EAAs) on LH secretion, we analyzed the effects of different inhibitors of NO synthase (NOS) in the LH response to kainic acid (KA), an agonist of kainate receptors, in male and female rats, neonatally injected with estradiol that show an increased sensitivity to EAAs. We found that NAME (40 and 60 mg/kg) increases LH secretion in intact and ovariectomized females, while SNP had no effect. The effect of NAME was not mediated through a direct action at pituitary level, since the basal and LHRH-stimulated LH release remained unchanged in presence of NAME. Similarly, basal and LHRH-stimulated LH secretion from dispersed pituitary cells were unaffected by NAME. Finally, the stimulatory effects of KA on LH release were not abolished by NOS inhibitors. In conclusion, our results provide evidence that the global action of NOS inhibitors is an increase in basal LH secretion, through a mechanism that remains to be fully characterized. In addition, our data demonstrate that the KA-stimulated LH secretion is not mediated by an increase in NO generation.     

一氧化氮在L-精氨酸促生长激素分泌中的作用

氨基酸是促进垂体生长激素(GH)分泌的潜在刺激因子.在所有氨基酸中,左旋精氨酸(L-精氨酸)促GH分泌作用最为明显.口服或静脉滴注精氨酸均可刺激垂体GH的分泌.L-精氨酸促GH的分泌可能涉及多种机制,但一氧化氮(NO)依赖的可溶性鸟苷酸环化酶-环鸟苷酸-蛋白激酶G(sGC-cGMP-PKG)信号转导通路在此过程中起着重...     

Nitric oxide regulates the release of somatostatin from cultured gastric rabbit primary D-cells

BACKGROUND & AIMS: Neuronal nitric oxide synthase (nNOS) is present in gastric D-cells. Mucosal somatostatin is diminished in H. pylori gastritis, where production of nitric oxide (NO) is increased. Therefore, we investigated the role of NO in D-cell function and the effects of prolonged exposure of D-cells to NO. METHODS: Rabbit gastric D-cells were cultured. Somatostatin-14 was measured after 2 hours to examine the effects of arginine, nitric oxide sythase (NOS) inhibitors, and NO donors. Some cells were preincubated with a slow releasing NO donor for 12 hours. Results are expressed as percentage of total cell content. Nitrate content was measured by chemiluminescent assay. RESULTS: L-arginine increased somatostatin-14 release in the presence of CCK8 from 4.4% +/- 0.5% to 6.4% +/- 0.4% (P < 0.02), and this was accompanied by NO release from 27 +/- 7 micromol/L to 86 +/- 12 micromol/L (P = 0.001). D-arginine and L-lysine had no effect. NOS inhibitors LNNA, SMT, and 7NI significantly attenuated the stimulatory response to L-arginine. NO donors sodium nitroprusside (SNP), 1 mmol/L, and S-nitroso-N-acetyl-D-L-penicillamine, 0.1 mmol/L, significantly increased basal and cholecystokinin-8 (CCK8) stimulated somatostatin release. Oxyhemoglobin attenuated the effect of SNP but not of L-arginine. Neither cyclic guanosine monophosphate nor guanylate cyclase were involved in the response to NO. However, inhibition of adenosine diphosphate (ADP) ribosyltransferase significantly decreased the response to L-arginine. Preincubation for 12 hours with 150 micromol/L (Z)-1-[(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate; IP3, inositol triphosphate decreased the 2-hour cellular response to CCK8 and SNP. CONCLUSIONS: NO regulates rabbit D-cells. Acute exposure stimulates somatostatin mediated by ADP ribosylation, whereas long-term exposure reduces cellular responses to stimuli. The latter pathway may be responsible for the suppression of somatostatin in H. pylori gastritis.     

Regulation of growth hormone release in common carp pituitary cells by pituitary adenylate cyclase-activating polypeptide: signal transduction involves cAMP- and calcium-dependent mechanisms

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a member of the glucagon/secretin peptide family and its molecular structure is highly conserved among vertebrates. In this study, the role of PACAP in regulating growth hormone (GH) secretion in fish was examined in vitro using common carp pituitary cells under column perifusion. A dose-dependent increase in GH release was observed after exposing pituitary cells to increasing doses of ovine PACAP38 (oPACAP38) and PACAP27 (oPACAP27), but not vasoactive intestinal polypeptide (VIP). A lack of GH response to VIP stimulation is consistent with the pharmacological properties of PAC-1 receptors, suggesting that this receptor subtype may be involved in PACAP-induced GH secretion in carp species. Although the maximal GH responses induced by oPACAP38 and oPACAP27 were similar, the minimal effective dose and ED50 value for oPACAP38 were significantly lower than that for oPACAP27. These results may indicate that common carp PAC-1 receptors are more sensitive to stimulation by oPACAP38 than by oPACAP27. In parallel studies, oPACAP38 and oPACAP27 were also effective in increasing cAMP release, cellular cAMP content, total cAMP production, and intracellular Ca(2+) ([Ca(2+)](i)) levels in common carp pituitary cells. Besides, the rise in [Ca(2+)](i) induced by oPACAP38 was blocked by removing extracellular Ca(2+) ([Ca(2+)](e)) or by treatment with nifedipine, an inhibitor of voltage-sensitive Ca(2+) channels (VSCC). The dose dependence of PACAP-stimulated GH release in common carp pituitary cells was mimicked by activating adenylate cyclase using forskolin, inhibiting cAMP degradation using IBMX, increasing functional levels of intracellular cAMP using CPT-cAMP, or inducing [Ca(2+)](e) entry using the Ca(2+) ionophore A23187. In contrast, the GH-releasing effect of oPACAP38 was suppressed by treatment with the adenylate cyclase inhibitor MDL12330A, protein kinase A inhibitor H89, and VSCC blocker nifedipine, or by perifusion with a Ca(2+)-free culture medium. These results, as a whole, suggest that PACAP functions as a GH-releasing factor in common carp by activating pituitary receptors resembling mammalian PAC-1 receptors. Apparently, the GH-releasing action of PACAP is mediated through the adenylate cyclase/cAMP/protein kinase A pathway and [Ca(2+)](e) influx through VSCC.     

Stimulatory effect of pituitary adenylate-cyclase activating polypeptide (PACAP) and its PACAP type I receptor (PAC1R) on prolactin synthesis in rat pituitary somatolactotroph GH3 cells

In this present study, we investigated the role of pituitary adenylate cyclase-activating polypeptide (PACAP) and its receptor, PACAP type I receptor (PAC1R) on prolactin synthesis in pituitary somatolactotroph GH3 cells. PACAP increased prolactin promoter activity up to 1.3 ± 0.1-fold. This increase, while significant, was less than the increase resulting from thyrotropin-releasing hormone (TRH) stimulation. By transfection of a PAC1R expression vector to the cells, the response to PACAP on prolactin promoter activity was dramatically potentiated to a degree proportional to the amount of PAC1R transfected. In the PAC1R expressing GH3 cells, TRH and PACAP alone increased prolactin promoter up to 3.3 ± 0.3-fold and 4.9 ± 0.2-fold, respectively, and combined treatment with TRH and PACAP further increased prolactin promoters up to 6.8 ± 0.6-fold. PACAP binds both Gs- and Gq-coupled receptors and stimulates adenylate cyclase/cAMP and protein kinase C/extracellular signal-regulated kinase (ERK) signaling pathways. PACAP increased ERK phosphorylation in PAC1R expressing cells to the same degree as TRH. Combined treatment with TRH and PACAP had a synergistic effect on ERK activation. GH3 cells produce both prolactin and growth hormone. Stimulation of GH3 cells with TRH significantly increased the mRNA level of prolactin and attenuated growth hormone mRNA expression. PACAP increased both prolactin and growth hormone mRNA levels, particularly in PAC1R expressing cells. In addition, increasing amount of PAC1R in GH3 cells potentiated the action of TRH on prolactin promoter activity, as well as on ERK phosphorylation. PAC1R was induced by PACAP itself, but not by TRH. Our current study demonstrates that PACAP and its PAC1R, functions as a stimulator of prolactin alone or with TRH in prolactin producing cells.     

一氧化氮在乳鼠心肌细胞缺氧—复氧预处理延迟保护效应中的作用

探讨缺氧—复氧损伤对乳鼠心肌细胞一氧化氮释放和一氧化氮合酶活性的影响以及一氧化氮在心肌细胞延迟缺氧预处理中的作用。在培养乳鼠心肌细胞缺氧预处理的模型上，测定缺氧—复氧损伤对乳鼠心肌细胞一氧化氮释放和一氧化氮合酶活性，观察延迟缺氧预处理以及N-硝基-L-精氨酸、L广精氨酸、硝普钠对心肌细胞延迟缺氧预处理的影响。结果发现，缺氧—复氧后乳鼠心肌细胞一氧化氮释放增加，一氧化氮合酶活性升高。延迟缺氧预处理可以减少缺氧一复氧对心肌细胞的损伤。非选择性一氧化氮合酶抑制剂N-硝基-L-广精氨酸可以阻断延迟缺氧预处理的心肌保护作用，L广精氨酸不能模拟延迟缺氧预处理，硝普钠可以模拟延迟缺氧预处理。结果提示，一氧化氮可以诱导心肌细胞的延迟缺氧预处理。     

Distribution of nitric oxide synthase and secretory role of exogenous nitric oxide in the isolated rat pancreas.

BACKGROUND: Pancreatic production and in vivo effects of nitric oxide (NO) have been shown by several studies. In order to examine the direct actions of the NO donor sodium nitroprusside (SNP), this study used in vitro specimens of the rat pancreas where the distribution of neuronal nitric oxide synthase (NOS) and the secretory effects of SNP and the cyclic GMP (cGMP) analog 8-bromo cyclic GMP (8-Br cGMP) were investigated. METHODS: NO containing pancreatic nerves were visualized by NOS immunohistochemistry. Basal and stimulated amylase output from rat pancreatic segments was measured by an on-line fluorimetric method. Stimulation was achieved by either acetylcholine (ACh) or electrical field stimulation (EFS). Intracellular free calcium concentration ([Ca2+]i) was measured in dispersed pancreatic acinar cells. RESULTS: NOS containing nerves were demonstrated in the vicinity of pancreatic acini and blood vessels. SNP and 8-Br cGMP inhibited both basal and EFS evoked amylase output but failed to inhibit ACh induced amylase output. Basal [Ca2+]i was decreased by both SNP and 8-Br cGMP but neither SNP nor 8-Br cGMP influenced the ACh evoked increase in [Ca2+]i. CONCLUSION: NO is well distributed in the rat exocrine pancreas. Exogenous nitric oxide may have a dual action in the isolated rat pancreas: Inhibition of basal amylase secretion in acinar cells and inhibition of ACh release from intrinsic nerve terminals. Both effects seem to be calcium dependent and possibly mediated by cGMP.     

Pituitary adenylate cyclase-activating polypeptide (PACAP) potentiates the gonadotropin-releasing activity of luteinizing hormone-releasing hormone

In order to determine if the newly discovered neuropeptide, pituitary adenylate cyclase activating polypeptide (PACAP), interacts with the known hypothalamic releasing factors to modulate pituitary hormone secretion, the effect of PACAP, either alone or in combination with either LHRH, TRH, CRF or GHRH, was examined in rat anterior pituitary cell cultures. While PACAP alone weakly stimulated LH and FSH release, PACAP and LHRH, in combination, interacted synergistically to stimulate gonadotropin secretion. No significant changes in the secretion of either TSH, ACTH, or GH were observed in response to PACAP, either alone or in combination with the other releasing factors. Addition of an LHRH antagonist demonstrated that the PACAP effect on gonadotropin release was neither mediated by the LHRH receptor nor the result of LHRH contamination of the PACAP preparation. Because of the sequence homology (68%) between the N-terminal 28 amino acids of PACAP and VIP, the addition of a VIP antagonist was used to demonstrate that the PACAP effect is not mediated through the VIP receptor. The observation that PACAP interacts synergistically with LHRH in stimulating gonadotropin release suggests intriguing possibilities for PACAP in regulating gonadotropin secretion and reproductive function.     

Pituitary growth hormone secretion in the turbot, a phylogenetically recent teleost, is regulated by a species-specific pattern of neuropeptides.

In mammals, growth hormone (GH) is under a dual hypothalamic control exerted by growth hormone-releasing hormone (GHRH) and somatostatin (SRIH). We investigated GH release in a pleuronectiform teleost, the turbot (Psetta maxima), using a serum-free primary culture of dispersed pituitary cells. Cells released GH for up to 12 days in culture, indicating that turbot somatotropes do not require releasing hormone for their regulation. SRIH dose-dependently inhibited GH release up to a maximal inhibitory effect of 95%. None of the potential stimulators tested induced any change in basal GH release. Also, neither forskolin, an activator of adenylate cyclase, nor phorbol ester (TPA), an activator of protein kinase C, were able to modify GH release, suggesting that spontaneous basal release already represents the maximal secretory capacity of turbot somatotropes. In contrast, forskolin and TPA were able to increase GH release in the presence of SRIH. In this condition (coincubation with SRIH), pituitary adenylate cyclase-activating polypeptide (PACAP) stimulated GH release, whereas none of the other neuropeptides tested (GHRHs; sea bream or salmon or chicken II GnRHs; TRH; CRH) had any significant effect. These data indicate that inhibitory control by SRIH may be the basic control of GH production in teleosts and lower vertebrates, while PACAP may represent the ancestral growth hormone-releasing factor in teleosts, a role taken over in higher vertebrates by GHRH.