The T84 human colonic adenocarcinoma cell line produces mucin in culture and releases it in response to various secretagogues.

The T84 colonic adenocarcinoma cell line, which has been used extensively as a model for studies of epithelial chloride secretion, also produces mucin and secretes it in culture. Electron microscopy of fixed sections of cultured cells, along with Immunogold labelling with an antibody to human small intestine (SI) mucin, revealed the presence of goblet-like cells with mucin-containing secretory granules. The mucin was of high molecular mass, had an amino acid composition similar to that of purified human SI and colonic mucins, and competed effectively with SI mucin for binding to the anti-(SI mucin) antibody. A sensitive solid-phase immunoassay specific for intestinal mucins was developed and used to measure mucin secretion by T84 cells. Cultures were treated for 30 min at 37 degrees C with a number of agents known to cause chloride secretion by T84 cell monolayers and the amount of mucin appearing in the medium was measured. Carbachol (1 mM), A23187 (10 microM), prostaglandin E1 (PGE1) (1 microM) and vasoactive intestinal polypeptide (VIP) (0.1 microM) all stimulated mucin release, but histamine (1 mM) had no effect. Whereas VIP is reported to stimulate chloride secretion more strongly than carbachol, it was less effective than carbachol in stimulating mucin secretion. Phorbol 12-myristate 13-acetate (PMA) (0.1-10 microM) also stimulated mucin release strongly, implicating a responsive protein-kinase C-dependent pathway. Additive secretory responses were obtained with combined stimulation by VIP (10 nM-1 microM) and carbachol (1 mM). Responses to stimulation with A23187 (1-10 microM) together with PMA (10 nM-10 microM) suggest that cytosolic Ca2+ concentration is a modulator of PMA activity.     

Mutual dependence of VIP/PACAP and CCK receptor signaling for a physiological role in duck exocrine pancreatic secretion

Unlike in rodents, CCK has not been established as a physiological regulator in avian exocrine pancreatic secretion. In the isolated duck pancreatic acini, 1 nM CCK was required for stimulation of amylase secretion, maximal effect being achieved at 10 nM; picomolar CCK was without effect. Vasoactive intestinal peptide (VIP)/pituitary adenylate cyclase activating peptide (PACAP) receptor (VPAC) agonists PACAP-38 and PACAP-27 (10(-12)-10(-7) M) alone had no effect, but made picomolar CCK effective. VPAC agonist VIP 10(-10)-10(-7) M stimulated amylase secretion marginally, but made CCK 10(-12)-10(-10) M effective also. PACAP-27 and VIP both shifted the maximal CCK concentration from 10(-8) to 10(-9) M. This sensitizing effect was mimicked by forskolin. CCK dose dependently induced intracellular Ca2+ concentration ([Ca2+]i) oscillations. PACAP-38 (1 nM), PACAP-27 (1 nM), VIP (10 nM), or forskolin (10 microM) alone did not stimulate [Ca2+]i increase, neither did they modulate CCK (1 nM)-induced oscillations; but when they were added to cells simultaneously exposed to subthreshold CCK (10 pM), calcium spikes emerged. Amylase secretion induced by the simultaneous presence of 10 pM CCK and VPAC agonists was completely blocked by removing extracellular calcium, but the protein kinase C inhibitor staurosporine (1 microM) was without effect. CCK (10 nM)-induced secretion was inhibited by CCK1 receptor antagonist FK480 (1 microM). Gastrin from 10(-12) to 10(-6) M did not stimulate amylase secretion nor did it (100 nM) induce [Ca2+]i increase. The above data suggest that duck pancreatic acini possess both CCK1 and VPAC receptors; simultaneous activation of both is required for each to play a physiological role.     

Effect of vasoactive intestinal peptide, peptide histidine isoleucine and growth hormone-releasing factor-40 on bombesin-like immunoreactivity, somatostatin and gastrin release from the perfused rat stomach

Bombesin-like immunoreactivity (BLI) has been demonstrated in neurons of the gastrointestinal tract and gastric BLI secretion can be demonstrated in response to the classical neurotransmitter acetylcholine. Since structurally related peptides VIP, PHI and GRF have to be considered as peptidergic neurotransmitters it was of interest to determine their effect on gastric BLI secretion. Additionally, somatostatin (SLI) and gastrin secretion was examined. The isolated stomach of overnight fasted rats was perfused with Krebs-Ringer buffer via the celiac artery and the effluent was collected via the portal vein. The gastric lumen was perfused with isotonic saline at pH7 or pH2. All four peptides were tested at a dose of 10(-11) M and 10(-8) M at both pH levels and in addition the effect of VIP and PHI was examined at 10(-14) M and 10(-12) M during luminal pH2. At luminal pH7 VIP and PHI stimulated SLI release at 10(-8) M but had no effect on BLI or gastrin secretion. rGRF and hpGRF were both ineffective on SLI and gastrin release while rGRF inhibited and hpGRF stimulated BLI secretion. This effect was not dose related. At luminal pH2 all four peptides stimulated BLI secretion. Stimulation by PHI was already observed at a dose of 10(-14) M while VIP elicited a stimulatory effect at 10(-12) M. PHI at the two lowest concentrations of 10(-14) and 10(-12) M elicited a stimulation of SLI and gastrin release while the same doses of VIP and the higher doses of all four peptides had no effect on SLI and gastrin secretion at an acidic intraluminal pH.(ABSTRACT TRUNCATED AT 250 WORDS)     

Adenylate cyclase stimulation by VIP in rat and human parotid membranes

Adenylate cyclase activity was stimulated by vasoactive intestinal peptide (VIP) in rat parotid membranes, in the presence of 100 microM guanosine triphosphate (GTP). The threshold concentration of VIP was 300 nM and the activity doubled at the maximal VIP concentration tested (30 microM). The relative potency of peptides of the VIP family was: VIP greater than peptide histidine isoleucinamide (PHI) greater than secretin. The beta-adrenergic agent isoproterenol was a more efficient activator of rat parotid adenylate cyclase and its stimulatory effect, like that of VIP, depended on the presence of GTP. The effects of VIP and isoproterenol were both potentiated by 10 microM forskolin. By comparison with rat parotid preparations, membranes from a human parotid gland responded similarly to the VIP family of peptides (VIP greater than PHI greater than secretin). In both rat and human parotid membranes, two proteins (Mr 44 kDa and 53 kDa) of the alpha-subunit of Ns (the guanyl nucleotide-binding stimulatory protein) were labelled by ADP-ribosylation, in the presence of cholera toxin. Taken together, these results indicate that VIP receptors, when coupled to Ns, were able to activate the adenylate cyclase system in rat and human parotid membranes.     

Vasoactive intestinal polypeptide and acetylcholine stimulate exocrine secretion of epidermal growth factor from the rat submandibular gland

The effect of vasoactive intestinal polypeptide (VIP) and acetylcholine on secretion of epidermal growth factor (EGF) from the rat salivary glands was investigated. VIP in doses of 3 X 10(-10) to 3 X 10(-8) mol/kg per h stimulated secretion of saliva and total output of EGF dose-dependently. Acetylcholine also stimulated salivation and output of EGF. VIP in a dose of 3 X 10(-11) to 3 X 10(-10) mol/kg per h enhanced the stimulatory effect of acetylcholine, but this effect disappeared when the dose of VIP was increased. Adrenalectomy decreased acetylcholine stimulated total output of EGF by approximately 50%, but only by 20% when acetylcholine plus VIP was administered. EGF was localized to the convoluted granular tubules in the submandibular gland, whereas EGF could not be detected in the remaining salivary glands. The results suggest that VIP and acetylcholine cooperate in the control of exocrine secretion from the rat salivary glands. The effect of acetylcholine, however, seems to be partly dependent on circulating catecholamines.     

Adenylyl cyclase stimulation by VIP in rat seminal vesicle membranes.

Vasoactive intestinal peptide (VIP) stimulated adenylyl cyclase activity in membranes from rat seminal vesicle. GTP potentiated the stimulatory effect of VIP so that it was routinely included at 10 microM. The stimulation of adenylyl cyclase by VIP was time and temperature dependent. The response was linear with time up to 15 min at 30 degrees C. Half-maximal adenylyl cyclase activation (in the presence of 10 microM GTP) was achieved at 3.0 nM VIP. The enzyme activity increased about 150% with respect to basal values at the maximal VIP concentration tested (1 microM). The relative potency of peptides upon stimulation of adenylyl cyclase activity was: VIP greater than helodermin greater than peptide histidine isoleucinamide greater than rat growth hormone-releasing factor. Other agents like GTP (0.1 mM), GppNHp (0.1 mM), forskolin (0.1 mM) and sodium fluoride (10 mM) increased the adenylyl cyclase activity 1.8-, 4.4-, 6.7- and 2.4-fold, respectively. Taken together, the presence of VIP in nerve terminals innervating the seminal vesicle of rats and the existence of VIP receptors coupled to adenylyl cyclase strongly suggest a physiological role for this neuropeptide in the modulation of seminal vesicle cell function.     

Vasoactive intestinal peptide stimulates mucus secretion, but nitric oxide has no effect on mucus secretion in the ferret trachea.

Vasoactive intestinal peptide (VIP) and nitric oxide (NO) are neurotransmitters involved in the regulation of bronchial and pulmonary vascular tone. Published studies of the effects of VIP on airway mucus secretion have yielded conflicting results. The purpose of this study was to determine the effect of VIP on mucus secretion in the ferret trachea and if this effect was influenced by NO. We used a sandwich enzyme-linked lectin assay to measure mucin secretion and a turbidimetric assay to measure lysozyme (serous cell) secretion from ferret tracheal segments. VIP (10(-7) M) increased mucin secretion over 2 h. VIP (10(-9) to 10(-5) M) stimulated mucin secretion in a dose-dependent fashion. VIP-induced mucin secretion was partially blocked by a VIP receptor antagonist (a chimeric VIP-pituitary adenylate cyclase-activating peptide analog, VIP receptor antagonist) at a 10-fold excess concentration. At all concentrations tested, neither NG-nitro-L-arginine methyl ester, an inhibitor of NO synthase, nor S-nitroso-N-acetyl-penicillamine, an NO donor, had any significant effect on constitutive or VIP-induced mucus secretion. We conclude that VIP-stimulated mucin and lysozyme secretion was both time dependent and dose dependent and that NO neither stimulates nor inhibits mucus secretion in the ferret trachea.     

Role of cyclic adenosine 3',5'-monophosphate-dependent protein kinase in hormone-stimulated beta-endorphin secretion in AtT20 cells.

Secretion of beta-endorphin from mouse pituitary AtT20 cells is stimulated by a variety of compounds that raise intracellular cAMP and Ca2+. To investigate the role of cAMP-dependent protein kinases in secretion, AtT20 cells were transfected with an expression vector coding for a regulatory (R) subunit of cAMP-dependent protein kinase containing mutations in both cAMP-binding sites. Expression of the mutant regulatory subunit in stable transformants (RAB cells) results in a dominant inhibition of cAMP-dependent protein kinase activity. Isoproterenol (1 microM) or analogs of cAMP stimulated beta-endorphin secretion from AtT20 cells, but failed to stimulate secretion in RAB cells expressing the mutant R subunit. Secretion in response to CRF (100 nM) was inhibited by 80% in these mutant clones, whereas the secretory response to vasoactive intestinal peptide (VIP; 100 nM) or phorbol ester (100 nM phorbol myristate acetate) was not inhibited by the R subunit mutation. Intracellular cAMP was elevated in response to CRF (11- to 15-fold), isoproterenol (5- to 10-fold), and VIP (4- to 8-fold) in RAB cells. Similar concentrations of VIP were required to evoke beta-endorphin secretion in either RAB cells or AtT20 cells. As with most secretagogues, VIP-induced secretion was inhibited in the presence of either EGTA or a voltage-sensitive Ca2+ channel antagonist, PN200-110. The secretory response to VIP was unaffected by down-regulation of protein kinase-C. These results suggest that CRF and isoproterenol work via cAMP-dependent protein kinase to activate beta-endorphin secretion, whereas VIP can act by a different mechanism that does not involve cAMP-dependent protein kinase or protein kinase-C.     

Effect of VIP on the respiratory burst in human monocytes ex vivo during prolonged strain and energy deficiency

VIP-stimulated cyclic AMP production and VIP effect on the production of reactive oxygen compounds in human monocytes activated by serum opsonized zymosan (respiratory burst) were studied during a ranger training course lasting for five days with almost continuous physical activity, and deficiency of sleep and energy. Respiratory burst was inhibited and cyclic AMP production was stimulated by VIP on all days. Maximum cyclic AMP production stimulated by VIP (0.1 microM) on the day of control was 148.6% of basal, and 255.3%, 213.8%, 218.9% and 198.7% on Days 1, 2, 3 and 5. Maximum inhibition was observed 20 min after addition of the peptide on the day of control, after 5 min on Days 1, 2 and 3, and after 10 min on Day 5. Inhibition at the 5-min time point was 33.1% on the day of control, and 34.7%, 53.6%, 53.3% and 36.2% on the different days during the training course. The observed increment in VIP effect adds to prior reported data about increased VIP secretion during the training course, and may indicate enhanced physiological significance of VIP during stress.     

Role of the cytoskeleton in secretion of chloride by shark rectal gland

The salt gland of the spiny dogfish, Squalus acanthias, can be stimulated to secrete chloride by two different endogenous peptides: cardiac natriuretic peptide (CNP) and the neurotransmitter, vasoactive intestinal peptide (VIP). We examined the role of the actin cytoskeleton and of myosin light chains in this process by perfusing isolated rectal glands with and without an inhibitor of actin filament organization (cytochalasin D) and an inhibitor of myosin light chain kinase (ML-7). Cytochalasin D, 10(-6) M, reduced secretion stimulated by a 1-min bolus of CNP (5x10(-7) M) by 50-60%. In the presence of 10(-2) M procaine (which blocks neural release of VIP), cytochalasin D completely prevented CNP stimulation. In contrast, cytochalasin D did not inhibit stimulation of the rectal gland by VIP or by forskolin. Similarly, 5x10(-6)M ML-7 almost completely inhibited direct stimulation of rectal gland secretion by CNP, but did not alter chloride secretion induced by VIP or forskolin. Finally, the average time between hormonal injection and activation of secretion was 2 min longer for CNP than for VIP, consistent with the hypothesis that a contractile cellular function involving the cytoskeleton is important in CNP-induced chloride secretion, but less so when secretion is stimulated by VIP.     

GABAB Receptor-Mediated Enhancement of Vasoactive Intestinal Peptide-Stimulated Cyclic AMP Production in Slices of Rat Cerebral Cortex

Basal and vasoactive intestinal peptide (VIP)-stimulated accumulations of cyclic AMP were measured in slices of rat cerebral cortex. Neither gamma-aminobutyric acid (GABA) nor the selective GABAB receptor agonist (-)-baclofen stimulated basal cyclic AMP accumulation, whereas VIP caused a large dose-dependent increase in cyclic AMP levels. However, in the presence of 100 microM (-)-baclofen, the effects of VIP on cyclic AMP accumulation were significantly enhanced, with the responses to 1 microM and 10 microM VIP being approximately doubled. The enhancing effects of (-)-baclofen was dose related (1-1,000 microM), but an enhancing effect was not observed with 100 microM (+)-baclofen. In the presence of the GABA uptake inhibitor nipecotic acid (1 mM), GABA caused a similar dose-related enhancement of the VIP response. The ability of either GABA or (-)-baclofen to augment VIP-stimulated production of cyclic AMP was not mimicked by the GABAA, agonists isoguvacine and 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP) and was not antagonized by the GABAA antagonist bicuculline. The putative GABAB antagonist 5-aminovaleric acid (1 mM) significantly reduced the effect of (-)-baclofen. The ability of (-)-baclofen to enhance VIP-stimulated accumulation of cyclic AMP was observed in slices of rat cerebral cortex, hippocampus, and hypothalamus. These results indicate that GABA and (-)-baclofen can enhance VIP-stimulated accumulation of cyclic AMP in rat brain slices via an interaction with specific GABAB receptors.     

Effect of vasoactive intestinal peptide on ion transport across dog tracheal epithelium

Under short-circuit conditions, vasoactive intestinal peptide (VIP) did not alter net Na+ movement but selectively stimulated net Cl- secretion across dog tracheal epithelium with a high affinity (Km congruent to 10(-8) M). The increase in Cl- secretion was not different from the rise in short-circuit current (Isc). However, stimulation of Cl- secretion was not maximal, because the addition of isoproterenol (10(-6) M) to VIP-treated tissues further increased the Isc by 54%. The effect of exogenous VIP was not blocked by a combination of atropine, phentolamine, propranolol (10(-5) or 10(-6) M), or tetrodotoxin (10(-6) M). Under open-circuit conditions, VIP caused an increase in the net secretion of Cl- and Na+, but the changes did not reach statistical significance. We conclude that VIP acts directly on receptors on the surface of epithelial cells to stimulate active Cl- secretion. The abundance of VIP nerves in the submucosa suggests that VIP may be important in regulation of fluid movement across the epithelium.     

Fetal adrenal VIP: distribution and effect on medullary catecholamine secretion

Vasoactive intestinal peptide (VIP) was found in the adrenal gland of ovine fetuses at 130-135 days gestation and was shown to stimulate catecholamine secretion. VIP was demonstrated by immunocytochemistry using the indirect antibody-enzyme method. VIP-immunoreactive nerve fibers were observed in the capsule, zona glomerulosa and inner layer of the cortex as well as in the medulla; furthermore small clusters of VIP-containing cell bodies were found at the corticomedullary border. To study the direct effect of VIP on catecholamine release, fetal adrenal medulla was dispersed into single cells and incubated in vitro with VIP for 6 hours. Catecholamine release into the medium was measured at 1, 3 and 6 hours. At 6 hours of incubation, VIP stimulated total catecholamine release from fetal adrenomedullary cells in a dose-dependent manner at concentrations ranging from 10(-8) to 10(-4) M. The release of norepinephrine and epinephrine, but not dopamine, was significantly enhanced. The presence of VIP in the fetal adrenal cortex and medulla, and the ability of VIP to stimulate catecholamine release from fetal adrenomedullary cells in vitro suggest that VIP may be an important modulator of medullary catecholamine secretion during fetal life.     

PACAP 1-38 as neurotransmitter in the porcine antrum

The concentration of PACAP 1-38 in porcine antrum amounted to 15.4+/-7.9 and 20.3+/-8 pmol/g tissue in the mucosal and muscular layers. PACAP immunoreactive (IR) fibres innervated the muscular (co-localised with VIP) and submucosal/mucosal layers (some co-storing VIP and CGRP) including myenteric and submucosal plexus and blood vessels. Only myenteric nerve cell bodies contained PACAP-IR (co-storing VIP). In isolated perfused antrum, vagus nerve stimulation (8 Hz) and capsaicin (10(-5) M) increased PACAP 1-38 release. PACAP 1-38 (10(-9) M) increased substance P (SP), gastrin releasing peptide (GRP) and VIP release. PACAP 1-38 (10(-8) M) inhibited gastrin secretion and stimulated somatostatin secretion and motility dose-dependently. PACAP-induced motility was strongly inhibited by the antagonist PACAP 6-38 but also by atropine and substance P-antagonists (CP99994/SR48968) but PACAP 6-38 had no effect on vagus-induced secretion or motility. Conclusion: PACAP 1-38 may be involved in antral motility and secretion by interacting with cholinergic, SP-ergic, GRP-ergic and/or VIP-ergic neurones, and may also be involved in afferent reflex pathways.     

Protein tyrosine phosphorylation in rabbit peritoneal neutrophils.

Protein tyrosine phosphorylation in rabbit peritoneal neutrophils was examined by immunoblotting with antibodies specific for phosphotyrosine. Stimulation of the neutrophils with chemotactic factor fMet-Leu-Phe (10 nM) caused rapid increases of tyrosine phosphorylation of several proteins with apparent molecular masses of (Group A) 54-58 kDa and 100-125 kDa and (Group B) 36-41 kDa. Stimulation of Group A proteins was observed by fMet-Leu-Phe (10 nM, maximum at 20 s) and A23187 (1 microM, 1 min). Stimulation of Group B proteins was observed by fMet-Leu-Phe (ED50 0.15 nM, 1 min), leukotriene B4 (ED50 0.15 nM, 1 min), phorbol 12-myristate 13-acetate (PMA) (ED50 25 ng/ml, 10 min) and partially by ionophore A23187 (1 microM, 1 min). Pretreatment of the cell with the protein kinase inhibitor H-7 (25 microM, 5 min) and PMA (0.1 microgram/ml, 3 min) partially inhibited the fMet-Leu-Phe effect. However, pretreatment of the cells with quin 2/AM (20 microM, 10 min) completely inhibited the fMet-Leu-Phe effect. The results indicate that rapid regulation of tyrosine phosphorylation is an early event occurring in stimulated neutrophils. Furthermore the effect of fMet-Leu-Phe on tyrosine phosphorylation may require Ca2+ mobilization and may partially require the activity of H-7-sensitive protein kinases.     

Vasoactive intestinal polypeptide and muscarine mobilize intracellular Ca2+ through breakdown of phosphoinositides to induce catecholamine secretion. Role of IP3 in exocytosis

We have recently shown that vasoactive intestinal polypeptide (VIP) is as potent as acetylcholine in inducing the secretion of catecholamines from the rat adrenal medulla. In the present study we have investigated the molecular mechanism involved in the exocytotic secretion of catecholamines by VIP and the effects of VIP on Ca45 uptake and phosphoinositide breakdown and compared them with those of the classical cholinergic agonists. We now show that omission of Ca2+ from the perfusion medium had almost no effect on VIP-induced secretion; however, addition of 1 mM EGTA to calcium-free medium abolished the secretion. Stimulation with VIP did not result in a net increase in Ca45 uptake and it was not modified by a protein kinase C activator, phorbol ester. All these effects of VIP were comparable to those of muscarine. VIP (0.3 to 10 microM) and muscarine (30 to 100 microM) produced time-and concentration-dependent increase (up to 700%) in the production of [3H]inositol phosphates. The production of [3H]inositol phosphates by VIP and muscarine occurred in calcium-free and EGTA medium. The effect of VIP on [3H]IP, [3H]IP2, and [3H]IP3 production was reduced by (1 to 30 microM) VIP antagonist (an analogue of growth hormone-releasing factor, Ac-Tyr1hGRF) and 1 to 20 microM naloxone. Although nicotine produced a brisk secretory response, there was no change in [3H]inositol phosphates. We conclude that inositol 1,4,5-trisphosphate generated upon activation of VIP and muscarine receptors is linked to exocytotic secretion of adrenal medullary hormones through release of internal Ca2+ ions.     

The effects of duodenal peptides on glucagon-like peptide-1 secretion from the ileum. A duodeno--ileal loop?

Secretion of the gut hormone glucagon-like peptide-1 (GLP-1) is stimulated by meal ingestion. The response is rapid, suggesting a stimulatory pathway elicited from the upper gastrointestinal area. In pigs, we have been unable to demonstrate a neural stimulatory pathway, but GLP-1 secretion is regulated by local somatostatin secretion. In search for an endocrine pathway, we studied the effect of a range of concentrations of cholecystokinin octapeptide (26-33) (CCK 8), gastric inhibitory peptide 1-42 (GIP), secretin, motilin, calcitonin gene-related peptide (CGRP), and the modified amino acid, 5-hydroxytryptamine (serotonin, 5-HT) on GLP-1 and somatostatin release from isolated perfused segments of porcine ileum.GLP-1 secretion was stimulated by 1 nM CCK 8 and 10 nM GIP, but suppressed by 1 nM motilin and 1 microM 5-HT. Secretin and CGRP had no effect. Somatostatin secretion was stimulated by CCK 8 at 1 and 10 nM, by GIP at 1 and 10 nM and by 10 nM CGRP. Secretin, 5-HT and motilin had no effect on somatostatin secretion.We conclude that CCK 8 and GIP 1-42 stimulated GLP-1 secretion, but only in concentrations greatly exceeding normal postprandial concentrations. Thus, we find it unlikely that endocrine agents from the duodenum regulate GLP-1 secretion in pigs.     

Effect of calcium ions on the processing of pro-opiomelanocortin by bovine intermediate lobe pro-opiomelanocortin-converting enzyme.

The effect of Ca2+ on the extent and pattern of processing of pro-opiomelanocortin and an N-terminal fragment by a purified pituitary secretory vesicle, soluble aspartic endoprotease, was studied. Ca2+ stimulated the first cleavage of pro-opiomelanocortin by pro-opiomelanocortin-converting enzyme to yield 21-23 kDa adrenocorticotropin and beta-lipotropin, but its effect was minimal. The production of adrenocorticotropin from the 21-23 kDa intermediate was stimulated approximately 2.3-fold in the presence of 10 mM Ca2+, and processing of beta-lipotropin to beta-endorphin was stimulated about 1.3-1.4-fold by 5-10 mM Ca2+. The production of gamma-melanotropin-immunoreactive material from bovine N-pro-opiomelanocortin(1-77) was stimulated approximately 1.3-fold at both 100 microM and 1.5-2.0 mM Ca2+. Further characterization of the gamma-melanotropin-immunoreactive material by HPLC demonstrated that the major products were gamma 3-[Lys]melanotropin and gamma 3-melanotropin at both Ca2+ concentrations. These results indicate that pro-opiomelanocortin-converting enzyme is stimulated by Ca2+.     

Effect of pituitary adenylate cyclase activating polypeptide on rat pancreatic exocrine secretion.

A novel neuropeptide, pituitary adenylate cyclase activating polypeptide (PACAP), which has been isolated from ovine hypothalami, shows 68% homology with vasoactive intestinal peptide (VIP). Since VIP stimulates amylase secretion from the pancreas, we investigated the effect of PACAP and VIP on rat pancreatic exocrine secretion after intravenous injections of PACAP-27, PACAP-38, or VIP at doses of 2.5, 5 or 10 nmol/kg. Results showed: 1) Bolus injection of PACAP stimulated pancreatic amylase and protein secretions in a dose-dependent manner; and 2) Stimulation of amylase secretion with 10 nmol/kg of PACAP-27 was greater than that induced with the same dose of VIP or PACAP-38 (p less than 0.05).     

Bombesin stimulates prolactin secretion from cultured rat pituitary tumour cells (GH4C1) via activation of phospholipase C

Bombesin (BBS) stimulated prolactin (PRL) secretion from monolayer cultures of rat pituitary tumour cells (GH4C1) in a dose-dependent manner with half maximal and maximal effect at 2 nM and 100 nM, respectively. No additional stimulatory effect on PRL secretion was seen when BBS was combined with thyroliberin (TRH) used in concentrations known to give maximal effects, while the effects of BBS and vasoactive intestinal peptide (VIP) were additive. Using a parafusion system, BBS (1 microM) was found to increase PRL secretion within 4 s and the secretion profiles elicited by BBS and TRH (1 microM) were similar. Both BBS and TRH increased inositoltrisphosphate (IP3) as well as inositolbisphosphate (IP2) formation within 2 s. BBS also induced the same biphasic changes in the electrical membrane properties of GH4C1 cells as TRH, and both peptides caused a rapid and sustained increase in intracellular [Ca2+]. These results suggest that BBS stimulates PRL secretion from the GH4C1 cells via a mechanism involving the immediate formation of IP3 thus resembling the action of TRH.