Effect of PT-treatment on ANP-mediated inhibition of adenylate cyclase and amylase release in rat parotid gland

Effects of pertussis toxin (PT) treatment on atrial natriuretic peptide (ANP)-mediated inhibition of adenylate cyclase and amylase release were investigated in rat parotid gland. Adenylate cyclase activity stimulated by GTPS in PT-treated membranes was much larger than that in normal membranes. ANP dose-dependently inhibited adenylate cyclase stimulated by GTPS in control rat parotid membranes, however in membranes prepared from PT-injected (in vivo) rat parotid gland, ANP did not inhibit adenylate cyclase. ANP(10^–7M) inhibited cAMP accumulation stimulated by forskolin (10^–6M) in control rat parotid acinar cells by about 34%, however, in PT-treated cells, the inhibitory effect of ANP was attenuated completely. In control cells, amylase release stimulated by isoproterenol (10^–6M) and forskolin (10^–6M) were also depressed by ANP (10^–7M) by 27 and 30%, respectively. The inhibitory response of ANP on amylase release was completely attenuated by PT-treatment. Gi was detected as a ADP-ribosylated 41-KDa protein by incubation of parotid membranes with PT and [-³²P]NAD. In rat parotid gland, these results suggested that ANP mediates adenylate cyclase/cAMP system and consequently reduces amylase release through ANP-C receptor coupled to Gi. (Mol Cell Biochem)139: 53–58, 1994)     

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.     

Effect of beta-antagonists on isoprenaline-induced secretion of fluid, amylase and protein by the parotid gland of the red kangaroo, Macropus rufus

Selective and non-selective beta-adrenoceptor antagonists were used to block the increases in fluid, protein and amylase secretion caused by sympathomimetic stimulation of the parotid gland of red kangaroos during intracarotid infusion of isoprenaline. ICI118551 at antagonist/agonist ratios up to 300:1 caused increasing but incomplete blockade of fluid secretion, and protein/amylase release. Atenolol at antagonist/agonist ratios up to 300:1 was only marginally more potent than ICI118551 at blocking the fluid, protein and amylase responses. Propranolol at antagonist/agonist ratios of 30:1 was as effective at blocking fluid and protein secretion as the highest ratios of either atenolol or ICI118551. Simultaneous administration of atenolol (30:1) with ICI118551 (30:1) was not as potent as propranolol (30:1). Thus, the beta-adrenoceptor/s in the acini of the kangaroo parotid gland appear to have antagonist-binding affinities atypical of those found for eutherian tissues. The data are consistent with the gland possessing either a single anomalous beta-adrenoceptor or functional beta(2)-receptors in addition to the beta(1)-receptors which are characteristic of eutherian salivary glands.     

Vasoactive intestinal peptide and substance P in salivary glands of the rat following denervation or duct ligation

Immunoreactive vasoactive intestinal peptide (VIP) and substance P (SP) were studied in parotid, submaxillary and sublingual glands of the rat. The concentration of VIP was highest in the submaxillary gland and lowest in the parotid gland. The concentration of SP was highest in the parotid gland; it was at, or below the limit of detection in the sublingual gland. In the parotid gland the total amounts of VIP and SP were reduced by 95% after parasympathetic denervation (section of the auriculo-temporal nerve). In the submaxillary gland the total amounts of the peptides were unchanged after parasympathetic decentralization (section of the chorda-lingual nerve). In this gland the total amount of SP was reduced by 92% and that of VIP by 50%, when the chorda tympani nerve fibres were cut deep into the hilum. Cutting the nerve fibres at the hilum left the total amounts of the peptides unchanged in the submaxillary gland, whereas in the sublingual gland the total amount of VIP was reduced by 70%. Sympathetic denervation did not reduce the total amounts of the peptides. Duct ligation caused gland atrophy. In the parotid gland the total amounts of VIP and SP were reduced by 40%. In the submaxillary gland the same percentage reduction occurred with regard to SP; however, the total amount of VIP was reduced by 99%. The VIP- and SP-containing nerve fibres reach the salivary glands by the parasympathetic nerves. In both submaxillary and sublingual glands a certain fraction of VIP originates within the glands.     

PACAP-38, a novel peptide from ovine hypothalamus, is a potent modulator of amylase release from dispersed acini from rat pancreas.

Despite studies indicating the presence of specific pancreatic acinar receptors for PACAP-38, a peptide that was recently isolated from ovine hypothalamus, the actions of the new peptide on pancreatic enzyme secretion have not been examined. The present study demonstrates that in terms of cAMP production and amylase release from dispersed acini from rat pancreatic acini, PACAP-38 and an N-terminal fragment, PACAP-27, have the same potency and efficacy as vasoactive intestinal peptide (VIP). As with VIP, these actions are potentiated by adding an inhibitor of cyclic nucleotide phosphodiesterase, and combination of PACAP-38 with bombesin, CCK-8, carbachol or the calcium ionophore A23187 results in 2-fold augmentation of the secretory actions of these agents. Inhibition of PACAP-38-induced cAMP production and amylase release by two VIP-receptor antagonists indicates that the secretory effects of PACAP-38 are mediated by interaction with VIP receptors. PACAP-38, a new brain-gut peptide, may be a physiological modulator of pancreatic enzyme secretion.     

Exendin-3, a novel peptide from Heloderma horridum venom, interacts with vasoactive intestinal peptide receptors and a newly described receptor on dispersed acini from guinea pig pancreas. Description of exendin-3(9-39) amide, a specific exendin receptor antagonist.

Exendin-3 increased cellular cAMP levels and amylase release from dispersed acini from guinea pig pancreas. Low concentrations (0.1-3 nM) caused a 12-fold increase in cAMP, whereas higher concentrations (0.3-3 microM) caused an additional 24-fold increase in cAMP. Maximal cAMP with the highest concentration tested was the same as the maximal response with secretin, vasoactive intestinal peptide (VIP), peptide histidine isoleucine, helodermin, or helospectin-I. In terms of amylase release, exendin-3 had the same efficacy but was the least potent of these peptides. Exendin-3-induced increases in amylase release were inhibited by VIP receptor antagonists and the new peptide (greater than 0.1 microM) competed with radiolabeled VIP for binding sites on dispersed acini. Increasing concentrations of an exendin-3 fragment, exendin-3(9-39) amide, did not increase cAMP or amylase release but inhibited the increase in cAMP observed with 0.1-3 nM exendin-3. The fragment did not alter the effects of other peptides that are known to increase acinar cAMP. We conclude that exendin-3 interacts with at least two receptors on guinea pig pancreatic acini; at high concentrations (greater than 100 nM) the peptide interacts with VIP receptors, thereby causing a large increase in cAMP and stimulating amylase release; at lower concentrations (0.1-3 nM) the peptide interacts with a putative exendin receptor, thereby causing a smaller increase in cAMP of undetermined function. Exendin-3(9-39) amide is a specific exendin receptor antagonist.     

An in vitro evaluation of adrenergic action on rat pancreatic amylase secretion: lack of stimulatory effect.

To assess direct evidence of adrenergic stimulation in pancreatic amylase secretion, effects of catecholamines on amylase release and intracellular cyclic AMP accumulation were examined with rat dispersed pancreatic acini. We first carried out control studies with CCK-8 and carbamylcholine to evaluate the usefulness of the material for the examination of amylase secretion, and examined VIP-induced cyclic AMP accumulation to assess the agonist evoked intracellular response. As a result, significant effects of CCK-8, carbamylcholine and VIP were observed, which confirmed that dispersed pancreatic acini used in this study were useful in examining exocrine pancreatic secretion. However, catecholamines failed to stimulate amylase release from pancreatic acini, although a significant increase in intracellular cyclic AMP accumulation was observed. Thus the present study strongly suggests that direct involvement of catecholamine is unlikely in pancreatic amylase secretion, in contrast to results reported previously.     

Evidence that helodermin, a newly extracted peptide from Gila monster venom, is a member of the secretin/VIP/PHI family of peptides with an original pattern of biological properties

Helodermin, a newly isolated peptide from the venom of Gila monster (Heloderma suspectum) was shown to stimulate the adenylate cyclase activity of rat pancreatic membranes as efficiently as secretin and VIP. It also increased cyclic AMP levels and inhibited [125I]VIP binding in rat pancreatic acini. Finally, helodermin activated adenylate cyclase in membranes from rat heart, rat brain, and human heart, showing properties analogous yet distinct from those of secretin, VIP and PHI.     

Forskolin potentiates calcium-dependent amylase secretion from rat pancreatic acinar cells

The cellular and molecular effects of forskolin, a direct, nonhormonal activator of adenylate cyclase, were assessed on the enzyme secretory process in dispersed rat pancreatic acinar cells. Forskolin stimulated adenylate cyclase activity in the absence of guanyl nucleotide. It promoted a rapid and marked increase in cellular accumulation of cyclic AMP alone or in combination with vasoactive intestinal peptide (VIP) but was itself a weak pancreatic agonist and did not increase the secretory response to VIP or other cyclic AMP dependent agonists. Somatostatin was a partial antagonist of forskolin stimulated cyclic AMP synthesis and forskolin plus cholecystokinin-octapeptide (CCK-OP) induced amylase release. Forskolin potentiated amylase secretion in response to calcium-dependent agonists such as CCK-OP, carbachol and A-23187, but did not affect the ability of CCK-OP and (or) carbachol to mobilize 45Ca from isotope preloaded cells; forskolin alone did not stimulate 45Ca release. In calcium-poor media, the secretory response to forskolin and CCK-OP was reduced in a both absolute and relative manner. The data suggests that calcium plays the primary role as intracellular mediator of enzyme secretion and that the role of cyclic AMP may be to modulate the efficiency of calcium utilization.     

Sympathetic denervation impairs agonist-stimulated phosphatidylinositol metabolism in rat parotid glands.

Substance P, muscarinic and alpha-adrenoceptor agonists stimulated the incorporation of [3H]inositol into phosphatidylinositol in rat parotid gland slices. Surgical denervation of the sympathetic input to the rat parotid gland by superior cervical ganglionectomy produced marked reductions in these responses. The stimulated incorporation of radiolabelled precursors into phosphatidylinositol is a measure of its resynthesis after receptor-mediated breakdown of inositol phospholipids. We therefore examined the enzymic site of the lesion induced by sympathetic denervation using parotid gland slices labelled with either [3H]inositol or [32P]phosphate and stimulated with substance P. Receptor-activated phospholipase C attack upon [3H]inositol phospholipids was assayed by measuring the formation of [3H]inositol 1-phosphate in the presence of 10 mM-Li+ to inhibit further breakdown. It was not affected by denervation. Substance P elicited a rapid breakdown of phosphatidylinositol 4,5-bisphosphate and this response was reduced in the denervated gland. The second step in stimulated phosphatidylinositol turnover, phosphorylation of diacylglycerol to phosphatidate was not affected by denervation. Sympathetic denervation appears to induce a specific enzymic lesion in the parotid gland that impairs receptor-stimulated resynthesis of phosphatidylinositol from phosphatidate. This change in membrane lipid metabolism may be related to a number of the effects of sympathetic denervation, such as agonist supersensitivity, reduced gland cell proliferation and induction of new surface receptors.     

Evidence that metabolically active synaptosomes lack functional cyclic AMP-dependent protein kinase.

Cyclic adenosine monophosphate (cAMP)-mediated signal transduction was evaluated in synaptosomes prepared from rat brain cortex. Adenylate cyclase was responsive to known adenylate cyclase stimulators including peptides (CRH and VIP), catecholamines (norepinephrine and isoproterenol) and ligands that directly stimulate adenylate cyclase (forskolin). Cyclic AMP accumulation also increased approximately 2 to 3-fold, but none of the agonists was able significantly to activate cyclic AMP-dependent protein kinase (A-kinase) in cortical synaptosomes. However, in parallel studies with slices prepared from rat brain cortex, adenylate cyclase activity, cAMP accumulation and A-kinase activity were all stimulated by CRH, VIP, norepinephrine, isoproterenol and forskolin. These data suggest that, in intact synaptosomes, either the cellular machinery which facilitates binding of cAMP to the regulatory subunit of A-kinase is missing or the cAMP produced by adenylate cyclase is not accessible to A-kinase.     

Effect of ligands that increase cAMP on caerulein-induced zymogen activation in pancreatic acini

The pathological activation of proteases within the pancreatic acinar cell is critical to initiating pancreatitis. Stimulation of acinar cells with supraphysiological concentrations of the CCK analog caerulein (CER) leads to protease activation and pancreatitis. Agents that sensitize the acinar cell to the effects of CCK might contribute to disease. The effects of physiological ligands that increase acinar cell cAMP [secretin, VIP, and pituitary adenylate cyclase activating peptide (PACAP)] on CER-induced responses were examined in isolated rat pancreatic acini. Each ligand sensitized the acinar cell to zymogen activation by physiological concentrations of CER (0.1 nM). VIP and PACAP but not secretin also enhanced activation by supraphysiological concentrations of CER (0.1 muM). A cell-permeable cAMP analog also sensitized the acinar cell to CER-induced activation. The cAMP antagonist Rp-8-Br-cAMP inhibited these sensitizing effects. These findings suggest that ligands that increase acinar cell cAMP levels can sensitize the acinar cell to the effects of CCK-induced zymogen activation.     

Vasoactive Intestinal Polypeptide Receptors Linked to an Adenylate Cyclase, and Their Relationship with Biogenic Amine- and Somatostatin-Sensitive Adenylate Cyclases on Central Neuronal and Glial Cells in Primary Cultures

The presence of vasoactive intestinal polypeptide (VIP) receptors coupled to an adenylate cyclase was demonstrated on membranes of neurons or glial cells grown in primary cultures originating from the cerebral cortex, striatum, and mesencephalon of mouse embryos. A biphasic pattern of activation was observed in all these cell types, involving distinct high- and low-apparent-affinity mechanisms. The absence of additive effects of VIP and 3,4-dihydroxyphenylethylamine (DA, dopamine), isoproterenol (ISO), and 5-hydroxytryptamine (5-HT, serotonin) suggests that the peptide receptors are colocated with each of the corresponding amine receptors on neuronal membranes of the three structures studied. The nonadditivity between the VIP- and ISO-induced responses on cortical and striatal glial membranes reveals as well a colocation of VIP and beta-adrenergic-sensitive adenylate cyclases on the same cells. A subpopulation of mesencephalic glia could possess only one of the two types of receptors, as a partial additivity of the VIP and ISO responses was seen. In addition, VIP modified the characteristics of the somatostatin inhibitory effect on adenylate cyclase activity of neuronal membranes from the cerebral cortex and striatum but not from those of the mesencephalon. On striatal and mesencephalic glial membranes the somatostatin inhibitory effect was observed only in the presence of VIP. However, as previously seen with ISO, the presence of VIP did not allow the appearance of a somatostatin inhibitory response on cortical glial membranes. This suggests that cortical glia are devoid of somatostatin receptors.     

Does cyclic GMP mediate amylase release from mouse parotid acini?

In mouse parotid acini both cholinergic and beta-adrenergic agonists increased intracellular levels of cyclic-GMP (c-GMP) as well as amylase release. The derivative of c-GMP, 8-bromo-c-GMP, mimicked the effects of cholinergic and beta-adrenergic stimulation on amylase release. Nitroprusside (NP), hydroxylamine (HA) and sodium azide (NaA) increased c-GMP levels and also enhanced amylase release in a dose-dependent manner; cyclic-AMP (c-AMP) levels were not affected. The phosphodiesterase inhibitor 3-isobutyl-1-methyl-xanthine (MIX) enhanced the effects of carbachol on both c-GMP accumulation and amylase release. These results suggest that c-GMP may mediate the actions of cholinergic agonists and at least partially mediate the actions of beta-adrenergic agonists on mouse parotid enzyme release.     

Different Natures of Supersensitivity of Adenylate Cyclase Stimulated by Calcitonin Gene-Related Peptide and Isoproterenol in Rat Diaphragm After Denervation and Reserpine Treatment

In skeletal muscles, calcitonin gene-related peptide (CGRP) released from motor nerve terminals and humoral catecholamines stimulate adenylate cyclase (AC) and enhance muscle contraction. The effects of denervation and treatment with reserpine on twitch contraction and the AC system in rat diaphragm were investigated. The basal levels of twitch contraction and AC activity of the diaphragm of rats were both increased 2 weeks after phrenic nerve denervation but were not altered by treatment with reserpine. Reserpine treatment provoked supersensitivity of AC to isoproterenol, without affecting the response to CGRP. On the other hand, denervation decreased the activation of AC and enhancement of twitch contraction by CGRP, without affecting the responses to isoproterenol. These data suggest that denervation causes up-regulation of AC as a result of loss of CGRP release from nerve terminal and that depletion of catecholamines by reserpine treatment supersensitizes the responses at the beta-adrenoceptor level. Thus, nervous and humoral factors regulate the AC system in striated muscle by different mechanisms.     

Membrane transduction pathway in the neuronal control of protein secretion by the albumen gland in Helisoma (Mollusca)

The albumen gland in Helisoma secretes a perivitelline fluid which surrounds each egg and is made up of several 66 kDa protein subunits and polysaccharide complexes. Forskolin, an adenylate cyclase activator, stimulated the secretion and release of the perivitelline fluid. An acidic extract of the central nervous system increased the intracellular concentration of cAMP in the albumen gland and this results in the release of the 66 kDa molecule and other proteins. Digestion of the brain extract with proteases abolished this activity, suggesting that the factor is a peptide. Cyclic AMP analogues and [BMX also stimulated the protein secretion in dose-dependent manner. Forskolin when added with the brain factor had an additive response. SQ22536, a non-competitive inhibitor of adenylate cyclase, inhibited brain extract dependent adenylate cyclase activity whereas aluminum fluoride, a G protein activator, was found to stimulate adenylate cyclase. Dopamine also stimulates protein secretion by the albumen gland and through the application of various agonists and antagonists of dopamine, it was established that the neurotransmitter acts via D1-like receptors by stimulating adenylate cyclase.     

Role of cAMP inhibition of p44/p42 mitogen-activated protein kinase in potentiation of protein secretion in rat lacrimal gland

We previously found that addition of cAMP and a Ca(2+)/PKC-dependent agonist causes synergism or potentiation of protein secretion from rat lacrimal gland acini. In the present study we determined whether cAMP decreases p44/p42 mitogen-activated protein kinase (MAPK) activity in the lacrimal gland. Since we know that activation of MAPK attenuates protein secretion stimulated by Ca(2+)- and PKC-dependent agonists, we also determined whether this activation causes potentiation of secretion. Freshly prepared rat lacrimal gland acinar cells were incubated with dibutyryl cAMP (DBcAMP), carbachol (a cholinergic agonist), phenylephrine (an alpha(1)-adrenergic agonist), or epidermal growth factor (EGF). The latter three agonists are known to activate p44/p42 MAPK. p44/p42 MAPK activity and protein secretion were measured. As measured by Western blot analysis, DBcAMP inhibited both basal and agonist-stimulated p44/p42 MAPK activity. Cellular cAMP levels were increased by 1) using two different cell-permeant cAMP analogs, 2) activating adenylyl cyclase (L-858051), or 3) activation of G(s)-coupled receptors (VIP). The cell-permeant cAMP analogs, L-858051, and VIP inhibited basal p44/p42 MAPK activity by 50, 40, and 40%, respectively. DBcAMP and VIP inhibited carbachol- and EGF-stimulated MAPK activity. cAMP, but not VIP, inhibited phenylephrine-stimulated MAPK activity. Potentiation of secretion was detected when carbachol, phenylephrine, or EGF was simultaneously added with DBcAMP. We conclude that increasing cellular cAMP levels inhibits p44/p42 MAPK activity and that this could account for potentiation of secretion obtained when cAMP was elevated and Ca(2+) and PKC were increased by agonists.     

The sympathetic and parasympathetic nature of neuropeptide Y-immunoreactive nerve fibres in the major salivary glands of the rat

Summary The distribution and origin of neuropeptide Y in the major salivary glands of the rat was studied by indirect immunofluorescence technique. Numerous nerve fibres immunoreactive for the peptide were seen in the parotid and sublingual glands. Most of the fibres were located around blood vessels and salivary acini. In the submandibular gland the number of immunoreactive nerve fibres around the acini was lower in comparison with that in the parotid and sublingual glands. Some immunoreactive nerve fibres were also found around or along intra- and interlobular ducts in all major salivary glands.A large number of the neuropeptide-containing neuronal cell bodies and nerve fibres were detected in the sympathetic superior cervical ganglion. Sympathetic postganglionic nerve trunks of this ganglion contained numerous immunoreactive nerve fibres as well. A subpopulation of the neuronal cell bodies in the submandibular ganglion were immunoreactive to neuropeptide Y.Both uni- and bilateral superior cervical ganglionectomies caused a significant decrease in the number of immunoreactive nerve fibres around the blood vessels in all the major salivary glands. However, these denervations did not affect the density of nerve fibres around the acini and ducts. On the contrary, unilateral parasympathetic denervation by sectioning the auriculotemporal nerve reduced the fibres around the secretory acini in the parotid gland remarkably, while only a minor reduction in the density of immunoreactive fibres associated with the blood vessels of the gland was detected. Unilateral electrocoagulation of the trigeminal nerve branches caused no detectable change in the density of immunoreactive nerve fibres in any of the major salivary glands.On the basis of the present findings it is concluded that neuropeptide Y-reactive nerve fibres present in all major salivary glands around the blood vessels seem to be mainly sympathetic, whereas those around the acini and ducts seems to be of parasympathetic origin.     

Adenylate cyclase in prothoracic glands during the last larval instar of the silkworm, Bombyx mori

We have previously reported that the absence of prothoracicotropic hormone (PTTH) signal transduction during the early last larval instar of Bombyx mori plays a role in leading to very low ecdysteroid levels in the hemolymph, inactivation of the corpora allata, as well as larval-pupal transformation. In the present study, adenylate cyclase was characterized in crude preparations of prothoracic gland cell membranes in an effort to localize the cause of refractoriness to PTTH. It was found that cyclase activity of the prothoracic glands from the day 6 last instar showed activation responses to fluoride, a guanine nucleotide analogue, as well as calmodulin (CaM) in dose-dependent fashions. The additive effects of day 5 prothoracic gland adenylate cyclase stimulation by fluoride and CaM imply that there may exist Gs protein-dependent and CaM-dependent forms of adenylate cyclase. For day 1 last instar prothoracic glands, which showed no response to stimulation by PTTH in either cAMP generation or ecdysteroidogenesis, adenylate cyclase activity exhibited far less responsiveness to Ca(2+)/CaM than did that from day 5 glands. These findings suggest that day 1 prothoracic glands may possess some lesions in the receptor-Ca(2+) influx-adenylate cyclase signal transduction pathway and these impairments in PTTH signal transduction may be, at least in part, responsible for decreased ecdysteroidogenesis.     

Effects of Vasoactive Intestinal Peptide and Other Peptides on Cyclic AMP Accumulation in Intact Pieces and Isolated Horizontal Cells of the Teleost Retina

The effects of vasoactive intestinal peptide (VIP) and several other peptides have been examined on cyclic AMP accumulation in intact pieces and isolated horizontal cells of the teleost (carp) retina. VIP was the most effective peptide examined, inducing a dose-related response, and an approximately fivefold increase in cyclic AMP production when used at a concentration of 10 microM. Porcine histidine isoleucine-containing peptide and secretin, peptides structurally related to VIP, also stimulated cyclic AMP accumulation, but at concentrations of 10 microM induced responses which were only approximately 40% and 10%, respectively, of the response observed with 10 microM VIP. In contrast, several other peptides, including glucagon, neurotensin, somatostatin, luteinizing hormone-releasing hormone, alpha-melanocyte-stimulating hormone, cholecystokinin octapeptide26-33, gastrin-releasing peptide, thyrotropin-releasing hormone, and VIP10-28 were totally inactive. The response to 10 microM VIP was not antagonized by several dopamine antagonists, indicating the presence of a population of specific VIP receptors coupled to adenylate cyclase, distinct from the population of dopamine receptors coupled to adenylate cyclase also known to be present in this tissue. Finally, experiments involving the use of fractions of isolated horizontal cells indicate that these neurons possess a population of VIP receptors coupled to cyclic AMP production which would appear to share a common pool of adenylate cyclase with a population of similarly coupled dopamine receptors.(ABSTRACT TRUNCATED AT 250 WORDS)