Receptors for vasoactive intestinal peptide and pituitary adenylate cyclase-activating polypeptide in the goose cerebral cortex

Receptors for vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) in the goose cerebral cortex were characterized using two approaches: (1) in vitro radioreceptor binding of [(125)I]-VIP, and (2) effects of peptides from the VIP/PACAP/secretin family on cyclic AMP formation. The binding of [(125)I]-VIP to goose cortical membranes was rapid, stable, and reversible. Saturation analysis resulted in a linear Scatchard plot, suggesting binding to a single class of receptor binding sites with a high affinity (K(d)=0.76 +/- 0.13 nM) and high capacity (B(max)=70 +/- 7 fmol/mg of protein). Various peptides displaced the specific binding of 0.12 nM [(125)I]-VIP to the goose cerebral cortical membranes in a concentration-dependent manner. The relative rank order of potency of the tested peptides to inhibit [(125)I]-VIP binding to the goose cerebrum was: PACAP(38) asymptotically equal to mammalian VIP > or = PACAP(27) asymptotically equal to chicken VIP > PHI (peptide histidine-isoleucine) > secretin (inactive). About 52% of specific [(125)I]-VIP binding sites in the goose cerebral cortex was sensitive to 5'-guanylimidodiphosphate [Gpp(NH)p], a nonhydrolyzable analogue of GTP. PACAP(38) and PACAP(27) potently stimulated cyclic AMP formation in the goose cerebral cortical slices in a concentration-dependent manner, displaying EC(50) values of 45.5 nM and 51.5 nM, respectively. Chicken VIP was markedly less potent than both forms of PACAP, mammalian VIP only weakly affected the nucleotide production, while effects evoked by PHI were negligible. It is concluded that the cerebral cortex of goose contains VPAC type receptors that are labeled with [(125)I]-VIP and are positively linked to cyclic AMP formation. In addition, the observed stronger action of PACAP, when compared to VIP, on cyclic AMP production in this tissue suggests its interaction with both PAC(1) and VPAC receptors.     

Effect of kainic acid injections and other brain lesions on vasoactive intestinal peptide (VIP)-stimulated formation of cAMP in rat brain

Summary In rat striatal slices, both intrastriatal kainic acid injection, which destroys striatal neurones, and intranigral injection of 6-hydroxydopamine (6-OHDA), which leads to a degeneration of dopamine nerve terminals in the striatum, reduced vasoactive intestinal peptide (VIP)-induced cAMP accumulation by approximately 60%. Cortical ablation, which leads to degeneration of cortico-striatal fibres, had no effect on striatal VIP-induced cAMP formation, Knife cut lesions transecting the stria terminalis, which destroy afferent fibres to the amygdala, decreased the VIP-induced increase in cAMP in amygdala slices by 40%, while kainic acid injection into the amygdala had no effect. Kainic acid injection into several other brain regions, including hippocampus, cortex and hypothalamus also failed to affect the VIP-elicited increase in cAMP in slices, despite reductions in choline acetyltransferase, glutamate decarboxylase, cyclic nucleotide phosphodiesterase and basal levels of cAMP. The results of a study of the effects of various VIP fragments on cAMP stimulation in striatal and cortical slices suggests that the entire sequence of VIP is necessary for full activity. The results suggest that VIP may be involved in neuromodulation or neurotransmission in the striatum and/or nigrostriatal pathway and also in the stria terminalis from the bed nucleus to the amygdala.     

Effect of vasoactive intestinal peptide (VIP) and other peptides on cAMP accumulation in rat brain.

The effects of a number of peptides which have recently been demonstrated to be present in brain, were determined on adenylate cyclase activity in homogenates and on cAMP and cGMP levels in rat brain slices. Substance P, luteinizing hormone releasing factor, thyrotropin releasing factor, somatostatin, glucagon, and neurotensin were without effect in any of these tests. When slices from a number of regions of rat brain were incubated in the presence of 0.5 μM vasoactive intestinal peptide (VIP), a significant increase in the accumulation of cAMP over basal values was observed. There were no changes in cGMP levels. VIP also caused an increase in cell-free adenylate cyclase activity of striatal, cortical and hippocampal homogenates, and this response was considerably increased in the presence of guanylyl-imidodiphosphate (GMP-PNP). The phosphodiesterase inhibitor isobutylmethylxanthine caused a 3 to 6-fold increase in basal levels of cAMP in brain slices, but VIP was still able to elicit a further increase, indicating that its effects on cAMP accumulation were probably due to activation of adenylate cyclase. The increase in cAMP in cortical and hypothalamic but not striatal slices was affected by alterations in the calcium concentration of the incubation medium. When tissue slices were incubated in the presence of VIP and a variety of antagonist drugs (propranolol, phenoxybenzamine, α-flupenthixol, naloxone), no alteration in the VIP induced increase in cAMP was observed. Furthermore, when VIP was incubated in the presence of agonists (dopamine, noradrenaline, isoproterenol, prostaglandin E₁, adenosine), the induced increase in cAMP levels was additive to that caused by VIP. These results support a role for VIP as a neuromodulatory or neuro-transmitter compound in the central nervous system, (CNS), mediating its action through the adenylate cyclase/cAMP system.     

藿香正气液对大鼠血管活性肠肽的影响

目的：观察藿香正气液对大鼠血管活性肠肽（VIP）的影响。方法：80只Wistar大鼠随机分为藿香正气液组和时照组，分别给大鼠灌服藿香正气液或生理盐水1，6h后：①观察大鼠胃肠动力的变化；②运用放射免疫分析法测定血浆、胃窦和空肠组织匀浆中VIP含量的变化；③运用免疫组化法显示胃窦、空肠组织中VIP阳性产物的分布情况。结果：①应用藿香正气液后1，6h，大鼠胃肠动力显著增强，以用药后1h更为明显。②藿香正气液明显降低血浆、胃窦及空肠组织匀浆中VIP的含量，减少胃窦和空肠组织中VIP的阳性产物。结论：藿香正气液促胃肠动力作用可能与其对VIP的影响有关。     

Histamine-induced cyclic AMP formation in the chick hypothalamus: interaction with vasoactive intestinal peptide

Effects of histamine (HA) on cyclic AMP production and its action upon the effects evoked by vasoactive intestinal peptide (VIP) were studied in the chick hypothalamus. HA (0.1-1000 microM) potently stimulated cyclic AMP formation in the hypothalamic slices, reaching maximal effect (2.5-3.5-fold increase) at a 100 microM concentration, and displaying an EC50 value of approximately 6.5 microM/ The stimulatory action of HA was mimicked by agonists of HA receptors, with the following rank order of potency: HA>4-methylHA (H2)>or=Nalpha,Nalpha-dimethylHA (H3>H1=H2)>or=2-methylHA (H1)>amthamine (H2)>dimaprit (H2) approximately tele-methylHA. The HA (100 microM)-evoked increase in cyclic AMP production was concentration-dependently antagonized by selective H2-HA receptor blockers (aminopotentidine>cimetidine>or=ranitidine>zolantadine) and was not affected by mepyramine and thioperamide, a selective H1- and H3-HA receptor antagonist, respectively. The pharmacological profile of HA receptors linked to the cyclic AMP-generating system in the chick hypothalamus indicates that they represent either an avian-specific H2-like HA receptor or a novel subtype of HA receptors. Chicken VIP (cVIP; 0.1-3 microM) potently stimulated cyclic AMP synthesis in the chick hypothalamus in a concentration-dependent manner. A combination of cVIP with HA produced cyclic AMP response more than additive, and such a synergistic interaction was antagonized by ranitidine. It is suggested that in the avian brain HA and VIP may play in concert to regulate neuroendocrine processes.     

Effect of vasoactive intestinal peptide on pulmonary surfactants phospholipid synthesis in lung explants

AIM: To investigate the effect of vasoactive intestinal peptide (VIP) on pulmonary surfactants (PS) phospholipidsynthesis in cultured lung explants. METHODS: Lung explants were cultured with serum-free medium, [methyl^3H]choline incorporation, total phospholipid, phosphatidylcholine, activity of choline-phosphate cytidylyltransferase(CCT) and CCTα mRNA level in lung explants were determined. RESULTS: (1) VIP (10^-10-10^-7 mol/L) for 16 hpromoted [methyl-^3H]choline incorporation in dose dependence and VIP (10.8 mol/L) for 2 h-16 h promoted [methyl-^3H]choline incorporation in time dependence. (2) VIP (10-8 mol/L) enhanced the contents of total phospholipidsand phosphatidylcholine in lung explants. (3) VIP (10^-10-10-7 mol/L) elevated microsomal CCT activity of lungexplants in dose dependence. (4) VIP (10.8 mol/L) increased expression of CCTα mRNA in lung explants andalveolar type II cells (ATII). (5) [D-P-C1-Phe(6)-Leu(17)]-VIP (10^-6 mol/L), a VIP receptors antagonist, abolishedthe increase of [^3H]choline incorporation, microsomal CCT activity and CCTα mRNA level induced by VIP (10-8mol/L) in lung explants. CONCLUSION: VIP could enhance synthesis of phosphatidylcholine, the major compo-nent of pulmonary surfactants by enhancing microsomal CCT activity and CCTα mRNA level via VIP receptor-mediated pathway.     

Antidipsogenic action of vasoactive intestinal peptide in the rat

The effect of vasoactive intestinal peptide (VIP) on the intake of water was studied in the rat. Intracerebroventricular administration of vasoactive intestinal peptide strongly inhibited drinking in rats deprived of water, but peripheral administration had no effect, indicating that the site of action was central. Drinking induced by angiotensin II was also markedly blocked by simultaneous administration of vasoactive intestinal peptide. The results indicate that in the rat, vasoactive intestinal peptide may play a role in the control of intake of water as a neuropeptide thirst inhibitor.     

The significance of vasoactive intestinal peptide in immunomodulation

First identified by Said and Mutt some 30 years ago, the vasoactive intestinal peptide (VIP) was originally isolated as a vasodilator peptide. Subsequently, its biochemistry was elucidated, and within the 1st decade, their signature features as a neuropeptide became consolidated. It did not take long for these insights to permeate the field of immunology, out of which surprising new attributes for VIP were found in the last years. VIP is rapidly transforming into something more than a mere hormone. In evolving scientifically from a hormone to a novel agent for modifying immune function and possibly a cytokine-like molecule, VIP research has engaged many physiologists, molecular biologists, biochemists, endocrinologists, and pharmacologists and it is a paradigm to explore mutual interactions between neural and neuroendocrine links in health and disease. The aim of this review is firstly to update our knowledge of the cellular and molecular events relevant to VIP function on the immune system and secondly to gather together recent data that support its role as a type 2 cytokine. Recognition of the central functions VIP plays in cellular processes is focusing our attention on this "very important peptide" as exciting new candidates for therapeutic intervention and drug development.     

Pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal peptide (VIP) activate protein kinase C in chick cerebral cortex

Pituitary adenylate cyclase-activating polypeptide (PACAP38) and vasoactive intestinal peptide (VIP) were tested for their ability to influence protein kinase C (PKC) activity in the chick cerebral cortical slices. Thirty minutes incubation of the chick tissue with PACAP38 (0.1-1 microM) or VIP (0.3-3 microM) produced significant and concentration-dependent changes in PKC activity. Both peptides enhanced the enzyme activity in cell membrane preparation, and decreased it in cytosol preparation obtained from cerebral cortical slices. These changes in PKC activity suggest that PACAP and VIP are capable of activating this enzyme in cerebral cortex of chick.     

Role of vasoactive intestinal peptide in inflammation and autoimmunity

Vasoactive intestinal peptide (VIP), a peptide produced by immune cells, exerts a wide spectrum of immunological functions that control the homeostasis of the immune system. In the last decade, VIP has been clearly identified as a potent anti-inflammatory factor, both in innate and adaptive immunity. In innate immunity, this peptide inhibits the production of inflammatory cytokines and chemokines from macrophages, microglia and dendritic cells. In addition, VIP reduces the expression of co-stimulatory molecules on antigen-presenting cells, and therefore reduces stimulation of antigen-specific CD4 T-cells. In terms of adaptive immunity, VIP promotes T-helper (Th)2-type responses, and reduces inflammatory Th1-type responses. Several of the molecular mechanisms involved in the inhibition of cytokine and chemokine expression, and in the preferential development and/or survival of Th2 effectors are known. Therefore, VIP and its analogs have been proposed as promising alternative candidates to existing therapies for the treatment of acute and chronic inflammatory and autoimmune diseases. The aim of this review is to update knowledge of the cellular and molecular events that are relevant to VIP function in the immune system. The central functions that VIP plays in cellular processes is being recognized and attention is being focused on this important peptide with regard to exciting new candidates for therapeutic intervention and drug development.     

盐酸戊乙奎醚对小鼠血浆血管活性肠肽的影响

目的观察盐酸戊乙奎醚对小鼠胃肠动力及血浆血管活性肠肽的影响。方法健康昆明小鼠30只,体重18～24g,雌雄不限,随机分为对照组、阿托品组和盐酸戊乙奎醚组,每组10只。阿托品组腹腔注射阿托品0.3mg/kg,盐酸戊乙奎醚组腹腔注射盐酸戊乙奎醚0.3mg/kg,对照组给予等容量0.9%氯化钠溶液。3组给药15min后以营养性半固体糊0.8ml/只灌胃,30min后处死小鼠,采用放免法测定血管活性肠肽水平,计算胃内残留率。结果与对照组比较,阿托品组血管活性肠肽水平升高(P<0.05),盐酸戊乙奎醚组血管活性肠肽水平差异无统计学意义(P>0.05);与阿托品组比较,盐酸戊乙奎醚组血管活性肠肽水平下降(P<0.05)。与对照组比较,阿托品组胃内残留率升高,盐酸戊乙奎醚组胃内残留率差异无统计学意义(P>0.05);与阿托品组比较,盐酸戊乙奎醚组胃内残留率降低(P<0.05)。结论与阿托品比较,盐酸戊乙奎醚不抑制胃肠运动,其对血管活性肠肽的分泌无影响。     

血管活性肠肽化学及生物学稳定性研究

目的:考察血管活性肠肽(vasoactive intestinal peptide,VIP)的化学及生物学的稳定性,为VIP的制剂学研究提供依据。方法:考察VIP在不同pH值(2.0,4.0,7.0,9.0,11.0,13.0)、不同离子强度溶液、不同温度以及人工胃液和人工肠液中的稳定性,用HPLC法检测VIP含量变化。结果:VIP的稳定性具有pH依赖性,VIP在酸性及中性条件下稳定,pH≤7时几乎无降解,但VIP在碱性条件下不稳定,pH=13时30min已完全降解;离子强度对其稳定性无影响;VIP在冷冻条件下稳定性良好,在冷藏条件下低浓度存在降解;VIP在人工胃液和人工肠液中降解迅速,0 min即完全降解,无法检测到主药峰。结论:VIP化学及生物学的稳定性差,口服无效。     

Electrical and mechanical effects of vasoactive intestinal peptide and pituitary adenylate cyclase-activating peptide in the rat colon involve different mechanisms

This work aimed to study the effects of pituitary adenylate cyclase-activating peptide (PACAP) and vasoactive intestinal peptide (VIP) on the mechanical and electrical activity of the circular muscle of the rat colon and the mechanisms involved in such effects. Spontaneous mechanical activity was studied in vitro in an organ bath and the membrane potential was recorded using the microelectrode technique. Both VIP and PACAP (0.1 microM) caused an immediate, sustained and tetrodotoxin (1 microM)-resistant inhibition of the cyclic spontaneous mechanical activity and hyperpolarization. The small-conductance Ca(2+)-activated K(+) channel blocker, apamin (1 microM), did not change the VIP- and PACAP-induced relaxation but reduced the hyperpolarization induced by PACAP whereas it did not change that induced by VIP. In contrast, the purinoceptor antagonist, suramin (100 microM), blocked the hyperpolarization caused by PACAP and VIP but failed to change their mechanical inhibitory effects. Moreover, the putative PACAP and VIP receptor antagonists, PACAP-(6-38) and VIP-(10-28), respectively, both 3 microM, failed to change the effects of either peptide and modified neither the inhibitory junction potential nor the relaxation induced by electrical-field stimulation. Thus, these results suggest that the mechanisms mediating relaxation are not strictly coupled to the mechanisms mediating hyperpolarization. This could be due to activation of two distinct mechanisms of action after agonist receptor interaction.     

Effects of penehyclidine hydrochloride on vasoactive intestinal peptide content in mice

目的 观察盐酸戊乙奎醚对小鼠胃肠动力及血浆血管活性肠肽的影响.方法 健康昆明小鼠30只,体重18 ～24 g,雌雄不限,随机分为对照组、阿托品组和盐酸戊乙奎醚组,每组10只.阿托品组腹腔注射阿托品0.3 mg/kg,盐酸戊乙奎醚组腹腔注射盐酸戊乙奎醚0.3 mg/kg,对照组给予等容量0.9％氯化钠溶液.3组给药15 min后以营养性半固体糊0.8 ml/只灌胃,30 min后处死小鼠,采用放免法测定血管活性肠肽水平,计算胃内残留率.结果 与对照组比较,阿托品组血管活性肠肽水平升高(P＜0.05),盐酸戊乙奎醚组血管活性肠肽水平差异无统计学意义(P＞0.05);与阿托品组比较,盐酸戊乙奎醚组血管活性肠肤水平下降(P＜0.05).与对照组比较,阿托品组胃内残留率升高,盐酸戊乙奎醚组胃内残留率差异无统计学意义(P＞0.05);与阿托品组比较,盐酸戊乙奎醚组胃内残留率降低(P＜0.05).结论 与阿托品比较,盐酸戊乙奎醚不抑制胃肠运动,其对血管活性肠肽的分泌无影响.     

脑出血患者血浆和颅内血肿液中血管活性肠肽的含量

作者测定了脑出血患者静脉血浆和颅内血肿液中血管活性肠肽(VIP)含量的变化,发现脑出血患者静脉血浆VIP含量无显著改变,但颅内血肿液VIP含量显著低于静脉血浆VIP含量;不同时期颅内血肿液VIP含量比较,6～72h含量最低,72h后又有增高趋势。本研究结果表明,VIP参与了脑出血后脑循环改变的病理生理过程,尤其是参与了占位初始所致的局部脑损害过程,若能增加脑出血患者中枢神经系统(CNS)中VIP含量可能为脑出血的治疗指出一个新的途径。     

Ontogeny of vasoactive intestinal peptide receptors in rat ventral prostate

• 1.1. The stimulatory effect of VIP on rat prostatic adenylyl cyclase changes during postnatal development. It peaks at 2 months (peripubertal period), remains in a plateau between 3 and 12 months (adult period), and decreases at 24 months (old period).
• 2.2. The stimulation of rat prostatic adenylyl cyclase by the β-adrenergic agonist isoproterenol follows a pattern rather similar to that of VIP.
• 3.3. The values of VIP binding capacity correlate well with those observed for adenylyl cyclase between 1 and 12 months, whereas there appears to exist a great number of uncoupled VIP receptors at 0.5 and 24 months.
• 4.4. The apparent molecular mass (51 kDa) of the rat prostatic VIP-receptor complex remains unaltered during ontogenic development.

     

Spontaneous hydrolysis of vasoactive intestinal peptide in neutral aqueous solution

Hydrolysis of radioiodinated vasoactive intestinal peptide (VIP) was observed in buffered aqueous solution at neutral pH and 38 °C. The reaction displayed apparent first-order kinetics at initial peptide concentrations below 3 nM (K_obs= 1.5 × 10^?5s^?1), but the rate deviated below predicted values at higher peptide concentrations. The rate constant derived from the reaction progress curve over three half lives, starting at a concentration of 82 PM peptide, was also consistent with a first-order process. The reaction results in several products that were isolated and characterized as peptide fragments. Based on the identity of these fragments, we deduced hydrolysis at five different peptide bonds clustered between residues 17–25 of VIP. Control experiments were devised to eliminate trivial explanations for the peptide hydrolysis. Peptides representing the C-terminal segment 15–28 and the internal segment 14–22 assayed by analogous methods and under identical conditions were not degraded at a measurable rate. Sodium dodecyl sulfate and acetonitrile, agents known to influence the secondary structure of VIP, inhibited its spontaneous hydrolysis, as did chloride salts of sodium and calcium, albumin and a peptide unrelated to VIP. The rate and product distribution are inconsistent with known pathways of peptide degradation involving cyclic imide or anhydride formation at asparagine or aspartate residues. We suggest that the breakdown of VIP in dilute solutions represents an autolytic process.     

Conformational study of vasoactive intestinal peptide by computational methods

The conformational profile of vasoactive intestinal peptide (VIP) was characterized using computational methods. The strategy devised included a close examination of the conformational profile of the first 11 residues fragment followed by a study that considered the compatibility of the different conformations found with a continuation of the polypeptide chain in a-helical conformation. Accordingly, a detailed analysis of the conformational preferences of the N-terminal fragment of VIP(1–11) was carried out within the framework of the molecular mechanics approach, using simulated annealing in an iterative fashion as the sampling technique. In a second step, low-energy structures of the fragment were fused to the remainder of the VIP chain in the form of two noninteracting α-helices, according to a model of the structure of the peptide proposed from NMR studies. After investigation for compatibility of each of the low-energy structures of VIP(1–11) with the two helical regions by energy minimization, only 5 of 35 structures were discarded. Analysis of the structures characterized indicates that most of the conformations of VIP(1–11), including the global minimum, can be described as bent conformations. Conformations exhibiting α-turns and ß-turns, previously proposed by NMR studies were also characterized. The conformational analysis also suggests that the common structural features found in VIP(1–11) should also be present in VIP. Finally, because of the sequence homology between VIP and Peptide T, and the fact that both are ligands of the CD4 receptor, both sets of low-energy conformations were compared for similarity. The relevance of these results as guidance of the design of new peptide analogs targeted to the CD4 receptor is also discussed.