RESUMO
Pituitary adenylate cyclase-activating peptide (PACAP) and vasoactive intestinal peptide (VIP) activate two shared receptors, VPAC1 and VPAC2. Activation of VPAC1 has been implicated in elevating glucose output, whereas activation of VPAC2 may be involved in insulin secretion. A hypothesis that a VPAC2-selective agonist would enhance glucose disposal by stimulating insulin secretion without causing increased hepatic glucose production was tested using a novel selective agonist of VPAC2. This agonist, BAY 55-9837, was generated through site-directed mutagenesis based on sequence alignments of PACAP, VIP, and related analogs. The peptide bound to VPAC2 with a dissociation constant (K(d)) of 0.65 nmol/l and displayed >100-fold selectivity over VPAC1. BAY 55-9837 stimulated glucose-dependent insulin secretion in isolated rat and human pancreatic islets, increased insulin synthesis in purified rat islets, and caused a dose-dependent increase in plasma insulin levels in fasted rats, with a half-maximal stimulatory concentration of 3 pmol/kg. Continuous intravenous or subcutaneous infusion of the peptide reduced the glucose area under the curve following an intraperitoneal glucose tolerance test. The peptide had effects on intestinal water retention and mean arterial blood pressure in rats, but only at much higher doses. BAY 55-9837 may be a useful therapy for the treatment of type 2 diabetes.
Assuntos
Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Fragmentos de Peptídeos/farmacologia , Receptores de Peptídeo Intestinal Vasoativo/agonistas , Peptídeo Intestinal Vasoativo/farmacologia , Sequência de Aminoácidos , Animais , Pressão Sanguínea/efeitos dos fármacos , Células Cultivadas , Diarreia/tratamento farmacológico , Diarreia/metabolismo , Glucose/farmacologia , Frequência Cardíaca/efeitos dos fármacos , Hormônios/sangue , Humanos , Injeções Intravenosas , Injeções Subcutâneas , Secreção de Insulina , Ilhotas Pancreáticas/efeitos dos fármacos , Dados de Sequência Molecular , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Ratos , Ratos Wistar , Receptores de Polipeptídeo Hipofisário Ativador de Adenilato Ciclase , Receptores do Hormônio Hipofisário/metabolismo , Receptores de Peptídeo Intestinal Vasoativo/metabolismo , Receptores Tipo II de Peptídeo Intestinal Vasoativo , Receptores Tipo I de Polipeptídeo Intestinal Vasoativo , Peptídeo Intestinal Vasoativo/análogos & derivados , Peptídeo Intestinal Vasoativo/química , Peptídeo Intestinal Vasoativo/metabolismoRESUMO
Exocrine secretions proceed in two phases which can be studied individually in submandibular glands. We have investigated the response to neuropeptides and purinergic agonists of rat submandibular glands. Pituitary Adenylate Cyclase Activating Peptide (PACAP), an analog of VIP increased the intracellular concentration of cyclic AMP in acinar cells. PACAP also stimulated the activity of the Na(+)-K(+)-2Cl(-)-cotransporter. Extracellular ATP increased the [Ca2+]i in ductal cells. Two distinct receptors were involved in this response. A metabotropic purinergic receptor of the P2Y1 type raised the cellular concentration of IP3 after activating a phospholipase C. The second component of the purinergic response involved an ionotropic P2X7 receptor. After binding an agonist, this receptor formed a non-specific cation channel permeant to calcium and manganese, highly sensitive to inhibition by nickel. Two phospholipases A2 were activated following the occupancy of this receptor. The calcium-independent enzyme triggered kallikrein secretion in response to extracellular ATP. In conclusion, neuropeptides and purinergic agonists activate the acinar and ductal phases of the salivary secretion and are therefore promising candidates for the development of new sialagogues for therapeutic use.
Assuntos
Neuropeptídeos/fisiologia , Neurotransmissores/fisiologia , Agonistas Purinérgicos , Glândula Submandibular/efeitos dos fármacos , Glândula Submandibular/metabolismo , Animais , Proteínas de Transporte/fisiologia , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Polipeptídeo Hipofisário Ativador de Adenilato Ciclase , Ratos , Simportadores de Cloreto de Sódio-Potássio , Peptídeo Intestinal Vasoativo/análogos & derivados , Xerostomia/tratamento farmacológicoRESUMO
Melatonin secretion from the mammalian pineal gland is strongly stimulated by noradrenaline and also by vasoactive intestinal polypeptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP). Three types of receptors for VIP and PACAP have been characterized so far: VIP1/PACAP receptors and VIP2/PACAP receptors, which possess similar high affinities for VIP and PACAP, and PACAP1 receptors which exhibit a 100-1000-fold higher affinity for PACAP. The aim of the present study was to characterize the receptor subtype(s) mediating the stimulatory effects of VIP and PACAP on melatonin synthesis in the rat pineal gland. Autoradiographic studies showed that PACAP and VIP were equally potent in displacing binding of radioiodinated PACAP27 from pineal sections. Amplification of pineal complementary DNAs by polymerase chain reaction using specific primers for the different receptor subtypes revealed that all three receptor messenger RNAs are expressed and that VIP1/PACAP receptor messenger RNA was predominant over VIP2/PACAP receptor messenger RNA. In vitro, VIP and PACAP stimulated melatonin synthesis with similar high potency and the effect of the two peptides were not additive. The selective VIP1/PACAP receptor agonists [R16]chicken secretin (1-25) and [K15, R16, L27]VIP(1-7)/growth hormone releasing factor(8-27) were significantly more potent than the selective VIP2/PACAP receptor agonist RO 25-1553 in stimulating melatonin secretion. The stimulatory effects of VIP and PACAP were similarly inhibited by the VIP1/PACAP antagonist [acetyl-His1, D-Phe2, K15, R16, L27]VIP(3-7)/growth hormone releasing factor(8-27). These data strongly suggest that VIP and PACAP exert a stimulatory effect on melatonin synthesis mainly through activation of a pineal VIP1/PACAP receptor subtype.
Assuntos
Adenilil Ciclases/metabolismo , Glândula Pineal/química , Glândula Pineal/enzimologia , Receptores do Hormônio Hipofisário/análise , Receptores de Peptídeo Intestinal Vasoativo/análise , Agonistas Adrenérgicos beta/farmacologia , Animais , Autorradiografia , Broncodilatadores/farmacologia , Ativação Enzimática/efeitos dos fármacos , Expressão Gênica/fisiologia , Radioisótopos do Iodo , Isoproterenol/farmacologia , Masculino , Melatonina/metabolismo , Neuropeptídeos/farmacologia , Neurotransmissores/farmacologia , Fragmentos de Peptídeos/farmacologia , Peptídeos Cíclicos/farmacologia , Polipeptídeo Hipofisário Ativador de Adenilato Ciclase , RNA Mensageiro/análise , Ratos , Ratos Wistar , Receptores de Polipeptídeo Hipofisário Ativador de Adenilato Ciclase , Receptores do Hormônio Hipofisário/genética , Receptores do Hormônio Hipofisário/metabolismo , Receptores de Peptídeo Intestinal Vasoativo/genética , Receptores de Peptídeo Intestinal Vasoativo/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Peptídeo Intestinal Vasoativo/análogos & derivados , Peptídeo Intestinal Vasoativo/farmacologiaRESUMO
UNLABELLED: Because no significant information exists regarding the structure-activity of vasoactive intestinal polypeptide (VIP) to gut smooth muscle, we performed functional studies in vitro on opossum internal anal sphincter (IAS) smooth muscle strips and supplemented them with binding studies to assess the ability of VIP, its fragments and analogs to inhibit [125I]VIP binding to IAS smooth muscle membranes. Binding of radiolabeled VIP to its receptor was specific, saturable and time- and temperature-dependent. Of all the substances tested, VIP was the most potent in causing a fall in the resting tension of the IAS and inhibiting [125I]VIP binding. VIP 2-28, VIP 10-28 and the putative VIP antagonists (1 x 10(-6) M) [4Cl-D-Phe6,Leu17]VIP (VIP analog) and (N-Ac-Tyr1, D-Phe2)-growth hormone-releasing factor [GRF] (1-29)-NH2 (GRF analog) caused significant inhibition of [125I]VIP binding, but had only minimal effect on the resting tension of the IAS. VIP 9-18 and VIP 1-12 had neither any significant effect nor inhibition of receptor binding. The rank order of potencies for inhibition of binding was VIP > VIP analog > VIP 10-28 = VIP 2-28 > GRF analog > peptide histidine isoleucine > VIP 9-18. The IC50 values for VIP, VIP analog, VIP 10-28, VIP 2-28, GRF analog and peptide histidine isoleucine were 9.6 x 10(-9), 1.6 x 10(-7), 5.5 x 10(-7), 6.2 x 10(-7), 1.2 x 10(-6) and 1.2 x 10(-5) M, respectively. CONCLUSION: the full action of VIP is critically dependent upon the integrity of the entire VIP molecule.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Canal Anal/fisiologia , Músculo Liso/fisiologia , Fragmentos de Peptídeos/farmacologia , Receptores dos Hormônios Gastrointestinais/fisiologia , Peptídeo Intestinal Vasoativo/análogos & derivados , Sequência de Aminoácidos , Canal Anal/efeitos dos fármacos , Canal Anal/metabolismo , Animais , Feminino , Técnicas In Vitro , Radioisótopos do Iodo , Cinética , Masculino , Membranas/efeitos dos fármacos , Membranas/metabolismo , Dados de Sequência Molecular , Músculo Liso/efeitos dos fármacos , Músculo Liso/metabolismo , Gambás , Fragmentos de Peptídeos/metabolismo , Receptores dos Hormônios Gastrointestinais/efeitos dos fármacos , Receptores dos Hormônios Gastrointestinais/metabolismo , Receptores de Peptídeo Intestinal Vasoativo , Sermorelina/análogos & derivados , Sermorelina/farmacologia , Relação Estrutura-Atividade , Peptídeo Intestinal Vasoativo/antagonistas & inibidores , Peptídeo Intestinal Vasoativo/metabolismo , Peptídeo Intestinal Vasoativo/farmacologiaRESUMO
We recently reported that acute pharmacologic depression of dopaminergic tone at different times of day unmasks a sex-specific endogenous stimulatory rhythm regulating PRL secretion. The PRL secretory responses of ovariectomized rats to the dopamine antagonist domperidone (DOM) were higher at 0300 and 1700 h than at 1200 h. These are the times during which surges of PRL appear in mated rats. This experimental paradigm was used to investigate the roles of the putative PRL-releasing factors (PRFs) oxytocin (OT), vasoactive intestinal peptide (VIP), and serotonin (5-HT) in this rhythm. The role of OT was studied by infusion of the OT antagonist 1-deamino-2-D-Trp-4-Val-8-Orn-Oxytocin (OT-A, 0.5 microgram/kg min) for 6 h. Two hours after beginning the OT-A infusion DOM was administered, as a single injection of 200 micrograms/kg iv at either 0300, 1200, or 1700 h. Serial blood samples were collected immediately before and 5, 10, 20, 30, 60, 120, 180, and 240 min after DOM administration. Infusion of OT-A attenuated the heightened PRL secretory responses to DOM given at both 0300 and 1700 h but did not affect the response at 1200 h. The role of VIP was studied by infusing the VIP antagonist [D, 4-Cl-Phe6,Leu17] VIP (VIP-A, 0.1 microgram/kg.min) as described above. VIP-A infusion had no effect on the PRL secretory responses to DOM given at 1200 or 1700 h but attenuated the heightened response at 0300 h. In order to study the role of 5-HT in the rhythm, rats were pretreated with p-chlorophenylalanine (250 mg/kg sc) 48 and again 24 h before the experiment. Pretreatment with p-chlorophenylalanine had no effect on the PRL secretory responses to DOM given at 0300 or 1200 h, but it attenuated the augmented PRL secretory response at 1700 h. These data suggest that both VIP and OT act as endogenous PRFs at 0300 h and 5-HT and OT act as PRFs at 1700 h. We propose that VIP and 5-HT are continuously active oscillatory neurotransmitters regulating OT release into pituitary portal blood and that these daily events only eventuate in PRL release when the mating stimulus has release the lactotroph from the inhibitory effects of dopamine.
Assuntos
Hipotálamo/fisiologia , Periodicidade , Prolactina/metabolismo , Animais , Domperidona/farmacologia , Feminino , Fenclonina/farmacologia , Ovariectomia , Ocitocina/fisiologia , Prolactina/sangue , Ratos , Ratos Endogâmicos , Serotonina/fisiologia , Peptídeo Intestinal Vasoativo/análogos & derivados , Peptídeo Intestinal Vasoativo/farmacologia , Peptídeo Intestinal Vasoativo/fisiologiaRESUMO
The ability of centrally administered vasoactive intestinal peptide (VIP) to stimulate PRL secretion when injected intracerebroventricularly could be due to leakage to the pituitary, where it is known to exert direct PRL-releasing activity, or to a hypothalamic action on its own release or that of another possible PRL-releasing factor. When 3 micrograms VIP were injected into the third ventricle of conscious ovariectomized rats, a significant (P less than 0.005) and transient elevation of plasma oxytocin (OT) levels was observed. When OVX rats were injected iv with 1 ml anti-OT serum 30 min before the central administration of 3 micrograms VIP, the PRL surge seen after VIP injection in normal rabbit serum-treated controls was completely absent. The PRL surge seen after central VIP administration was not significantly altered by iv saline infusion (1 ml over 30 min) or by infusion of a VIP antagonist [D-4-Cl-Phe6,Leu17]VIP at a dose of 0.5 microgram/kg.min in 1 ml saline for 30 min before the VIP injection. This was not due to the inability of the VIP antagonist to block the PRL-releasing factor activity of VIP, since it significantly antagonized that action both in vitro and in vivo in the suckling stimulation paradigm. However, the PRL surge was completely absent in ovariectomized rats pretreated by iv infusion of an OT antagonist, [deamino Cys1,D-Trp2,Val4,Orn8]OT, at a similar dose. This recruitment of OT by VIP indicates that it may act at more than one locus within the hypothalamo-pituitary axis to insure the coordinated control of PRL secretion.
Assuntos
Ventrículos Cerebrais/fisiologia , Hipotálamo/fisiologia , Ocitocina/farmacologia , Adeno-Hipófise/metabolismo , Prolactina/metabolismo , Peptídeo Intestinal Vasoativo/farmacologia , Animais , Ventrículos Cerebrais/efeitos dos fármacos , Feminino , Lactação , Ovariectomia , Adeno-Hipófise/efeitos dos fármacos , Gravidez , Ratos , Ratos Endogâmicos , Valores de Referência , Peptídeo Intestinal Vasoativo/análogos & derivadosRESUMO
The thyroid gland is known to harbor cholinergic and VIPergic nerves. In the present study, the influences of cholinergic stimulation by carbachol, cholinergic blockade by methylatropine and stimulation with various VIP sequences on basal, TSH-induced and VIP-induced thyroid hormone section were investigated in vivo in mice. The mice were pretreated with 125I and thyroxine; the subsequent release of 125I is an estimation of thyroid hormone secretion. It was found that basal radioiodine secretion was inhibited by both carbachol and methylatropine. Furthermore, TSH-induced radioiodine secretion was inhibited already by a low dose of carbachol. Moreover, a high dose of carbachol could inhibit VIP-induced radioiodine secretion. Methylatropine did not influence TSH- or VIP-stimulated radioiodine secretion, but counteracted the inhibitory action of carbachol on TSH- and VIP-induced radioiodine release. In addition, contrary to VIP, six various synthesized VIP fragments had no effect on basal or stimulated radioiodine release. It is concluded that basal thyroid hormone secretion is inhibited by both cholinergic activation and blockade. Furthermore, TSH-induced thyroid hormone secretion is more sensitive to inhibition with cholinergic stimulation than is VIP-induced thyroid hormone secretion. In addition, the VIP stimulation of thyroid hormone secretion seems to require the full VIP sequence.