垂体腺苷酸环化酶激活肽诱导PC12细胞突起生长的作用

目的　研究垂体腺苷酸环化酶激活肽（ＰＡＣＡＰ） ３８与ＰＡＣＡＰ２７ 在ＰＣ１２ 细胞突起生长中的作用，并探讨介导其作用的受体和细胞内第二信使机制。方法　采用ＰＣ１２细胞分散培养法，观察接种７２ ｈ 时ＰＣ１２ 细胞突起生长阳性细胞的百分比。结果　当ＰＡＣＡＰ３８ 和ＰＡＣＡＰ２７ 的浓度为１ ×１０－ ７ ～１ ×１０－ １１ ｍｏｌ·Ｌ－１ 之间时，均能诱导ＰＣ１２细胞突起生长，其中以１×１０ －９ ｍｏｌ·Ｌ－ １ 的浓度作用最明显，量效曲线呈“钟”形。ＰＡＣＡＰ Ⅰ型受体拮抗剂ＰＡＣＡＰ６～３８和ｃＡＭＰ依赖的蛋白激酶抑制剂ＲｐｃＡＭＰＳ 能显著地抑制ＰＡＣＡＰ诱导ＰＣ１２ 细胞突起生长的作用，而蛋白激酶Ｃ 抑制剂Ｈ７ 却没有这个作用。结论　ＰＡＣＡＰ３８ 和ＰＡＣＡＰ２７均能诱导ＰＣ１２ 细胞突起生长，该作用是由ＰＡＣＡＰ Ⅰ型受体介导的，是通过细胞内ｃＡＭＰ 依赖的蛋白激酶系统实现的     

Pituitary adenylate cyclase activating polypeptide: a potential neuroprotective peptide

Pituitary adenylate cyclase activating polypeptide (PACAP) was first isolated from hypothalamic extracts on the basis of its ability to stimulate cAMP formation in pituitary cells. PACAP is widely distributed in the central and peripheral nervous systems and exerts numerous effects. Currently available data indicate that PACAP is a promising neuroprotective peptide. PACAP plays an important role during the development of the nervous system and in regeneration following nervous injuries. It has strong anti-apoptotic effects in several neuronal cultures and in vivo. PACAP protects neurons against various toxic insults in vitro, has anti-inflammatory actions and stimulates the release of neuroprotective substances from astrocytes. In vivo, the protective effects of PACAP have been shown in various models of brain injuries, including cerebral ischemia, Parkinson's disease, trauma and nerve transections. The upregulation of PACAP following several types of nerve injuries indicates that endogenous PACAP plays a role in the post-traumatic recovery of the nervous system. The present report reviews the current knowledge on the neurotrophic and neuroprotective effects of PACAP.     

Pituitary adenylate cyclase activating peptide mediates inhibitory nonadrenergic noncholinergic relaxation

We investigated the contribution of pituitary adenylate cyclase activating peptide (PACAP) to inhibitory nonadrenergic noncholinergic (inhibitory-NANC) relaxation of tracheal smooth muscle in cats. We also investigated the roles of vasoactive intestinal peptide (VIP) and nitric oxide (NO) on this function. Smooth muscle strips prepared from feline trachea were precontracted with 1 microM serotonin, and inhibitory-NANC relaxation was induced by electrical-field stimulation in the presence of atropine and propranolol. PACAP-(6-38) (a selective antagonist of PACAP; 1, 3 and 10 microM), VIP-(10-28) (a selective antagonist of VIP; 1, 3 and 10 microM) and N(omega)-nitro-L-arginine methyl ester (L-NAME, a selective NO synthase inhibitor; 3, 10 and 30 microM) each partially but significantly attenuated the amplitude of inhibitory-NANC relaxation. The effects of PACAP-(6-38) and VIP-(10-28) were additive. Addition of PACAP-(6-38) and/or VIP-(10-28) further attenuated relaxation in the presence of L-NAME. These results suggest that PACAP, VIP and NO contribute to the relaxation induced by inhibitory-NANC in tracheal smooth muscle in cats, and that they mediate this relaxation via different pathways.     

The effects of itopride on oesophageal motility and lower oesophageal sphincter function in man

Aliment Pharmacol Ther 2011; 33: 99–105

Summary

Background Itopride is a new prokinetic agent that combines antidopaminergic and cholinesterase inhibitory actions. Previous studies suggested that itopride improves heartburn in functional dyspepsia, and decreases oesophageal acid exposure in gastro‐oesophageal reflux disease. It remains unclear whether this effect is due to effects of itopride on the lower oesophageal sphincter (LES). Aims To study the effects of itopride on fasting and postprandial LES function in healthy subjects. Methods Twelve healthy volunteers (five men; 32.6 ± 2.0 years) underwent three oesophageal sleeve manometry studies after 3 days premedication with itopride 50 mg, itopride 100 mg or placebo t.d.s. Drug was administered after 30 min and a standardized meal was administered after 90 min, with measurements continuing to 120 min postprandially. Throughout the study, 10 wet swallows were administered at 30‐min intervals, and gastrointestinal symptoms were scored on 100 mm visual analogue scales at 15‐min intervals. Results Lower oesophageal sphincter resting pressures, swallow‐induced relaxations and the amplitude or duration of peristaltic contractions were not altered by both doses of itopride, at all time points. Itopride pre‐treatment inhibited the meal‐induced rise of transient LES relaxations (TLESRs). Conclusions Itopride inhibits TLESRs without significantly affecting oesophageal peristaltic function or LES pressure. These observations support further studies with itopride in gastro‐oesophageal reflux disease.     

VIP stimulates ACTH release and adenylate cyclase in human ACTH-secreting pituitary adenomas

The effect of vasoactive intestinal polypeptide (VIP) on in vitro ACTH release and adenylate cyclase activity was investigated in human ACTH-secreting pituitary adenomas from 4 patients with Cushing's disease and 2 patients with Nelson's syndrome. In all the tumors tested, VIP elicited a dose-dependent stimulation of hormone release from adenoma fragments (90-247% at 10-7 M VIP) and of cAMP formation in membrane preparations (75-140% at 3 X 10-6 M VIP). Therefore a role of VIP in the control of ACTH secretion in human ACTH-secreting adenomas is suggested; a cAMP-dependent mechanism of action can also be hypothesized.     

垂体腺苷酸环化酶激活肽对大鼠局灶性脑缺血/再灌注损伤脑保护作用的研究

目的观察垂体腺苷酸环化酶激活肽(PACAP)对大鼠局灶性脑缺血/再灌注损伤的脑保护作用。方法采用线栓法制备大鼠局灶性脑缺血/再灌注损伤(MCAO)模型,缺血前经侧脑室分别给予不同剂量的PACAP,脑缺血2 h/再灌注24 h,测定脑含水量、脑组织超氧化物歧化酶(SOD)活性和丙二醛(MDA)、一氧化氮(NO)含量。结果与NS组相比,PACAP各组脑含水量、MDA及NO含量均明显降低,SOD活性有不同程度的提高。结论PACAP对大鼠局灶性脑缺血/再灌注损伤有明显的脑保护作用,中、高剂量组效果优于低剂量组,其机制可能与减轻脑水肿、清除自由基、抗脂质过氧化有关。     

Ethnic variation in lower oesophageal sphincter pressure and length

Background  Oesophageal manometry (OM) is used to diagnose oesophageal motor disorders. Normal values of OM among United States ethnic groups are only available for Hispanic Americans (HA).
Aim  To obtain normal values of OM in adult African American (AA) volunteers, compare these with those obtained in HA and non-Hispanic white (nHw) volunteers to determine if ethnic variation in normal oesophageal motor function exists.
Methods  Healthy AA, HA and nHw were recruited from the Jacksonville metropolitan area. Ethnicity was self-reported. Exclusion criteria were symptoms suggestive of oesophageal disease, medication use or concurrent illness affecting OM. All underwent OM using a solid-state system with wet swallows. Resting lower oesophageal sphincter (LOS) pressure and LOS length were measured at mid-expiration, while per cent peristaltic contractions, distal oesophageal contraction velocity, amplitude and duration were measured after 5 cc water swallows.
Results  Fifty-six AA, 20 HA and 48 nHw were enrolled. All completed OM. AA had significantly higher resting LOS pressure, LOS length and distal oesophageal contraction duration than nHw ( P  < 0.05).
Conclusions  Significant ethnic exist in OM findings between AA and nHw. These underscore the need for ethnic specific reference values for OM to allow for correct diagnosis of oesophageal motor disorders in AA.     

Dissociation of cardiac inotropic and adenylate cyclase activating adrenoceptors

1 At higher temperatures, near the physiological range for mammals and nonhibernating frogs, the adrenoceptors for both inotropic responses to adrenaline and noradrenaline and for cyclic 3',5'-adenosine monophosphate (cyclic AMP) production in rat and frog isolated heart preparations, had typical beta characteristics. Phenoxybenzamine potentiated the inotropic response and the accumulation of cyclic AMP; conversely, propranolol inhibited the two responses.2 When the ambient temperature was reduced, the adrenoceptors mediating cyclic AMP production changed very little; they were blocked as effectively as at the higher temperature by propranolol and were not blocked by phenoxybenzamine. However, the adrenoceptors mediating the inotropic response were markedly changed by the decrease in temperature; phenoxybenzamine now inhibited this response and the inhibitory activity of propranolol was reduced about tenfold.3 These results indicate that the adrenoceptors that mediate cardiac inotropic responses at physiological temperatures are distinct from those that mediate the production of cyclic AMP, and that the activation of adenylate cyclase and the accumulation of cyclic AMP are probably not intermediate steps in cardiac inotropic responses to catecholamines.     

垂体腺苷酸环化酶激活肽对大鼠缺血性脑损伤的保护作用

脑缺血是严重危害人类健康的常见病，对其的预防和治疗一直是人们研究的重点课题。近年来，垂体腺苷酸环化酶激活肽（ＰＡＣＡＰ）在脑损伤中的作用日益受到人们的重视。ＰＡＣＡＰ由３８个氨基酸残基组成，是血管活性肠肽家族的新成员。体外研究结果表明，ＰＡＣＡＰ能保...     

Inhibition of bronchoconstriction by pituitary adenylate cyclase activating polypeptide (PACAP 1-27) in guinea-pigs in vivo.

1. We studied the inhibitory effect of pituitary adenylate cyclase activating polypeptide (PACAP 1-27) on the increase in total pulmonary resistance (RL) caused either by allergen or histamine in anaesthetized, ventilated guinea-pigs. 2. PACAP 1-27 given via i.v. infusion (0.045-4.5 nmol kg-1 min-1) dose-dependently reduced the increase in RL caused by inhaled ovalbumin and histamine. At the highest dose, PACAP 1-27 prevented the increase in RL caused by ovalbumin and histamine completely. Infusion of PACAP 1-27 and the beta 2-adrenoceptor agonist, salbutamol (0.045-4.5 nmol kg-1 min-1) inhibited the increase in RL similarly, but salbutamol increased the heart rate more than PACAP 1-27. 3. PACAP 1-27 and salbutamol given via inhaled aerosol (0.1 mM, 20 breaths) significantly reduced the increase in RL caused by histamine infused i.v., whereas aerosolised sterile saline did not. Both PACAP 1-27 and salbutamol caused bronchodilator effects within 1 min of drug inhalation and these effects remained throughout the 20 min of study. 4. Because PACAP 1-27 produced significant bronchodilatation and rapid onset of sustained action in vivo and without pronounced cardiovascular side effects, we conclude that this peptide may have therapeutic potential as a bronchodilator.     

Pituitary adenylate cyclase activating polypeptide is a potent vasodilator in humans.

The vasodilator effect of the novel peptide pituitary adenylate cyclase activating polypeptide (PACAP) was investigated in humans. Forearm blood flow was measured in six healthy men by venous occlusion plethysmography. Infusion of PACAP into the brachial artery at 0.01, 0.1, 1, 3, and 10 pmol/min produced a dose-related increase in forearm blood flow in the cannulated arm from 2.8 +/- 0.6 to 8.6 +/- 2.4 ml/100 ml/min at the highest dose (mean +/- SEM, p less than 0.05). In a subsequent experiment, where the highest dose of PACAP was repeated after a 36 min interval, there was no tachyphylaxis of the forearm blood flow response, with the forearm blood flow increasing by 129 +/- 9% during the first infusion and 128 +/- 31% during the second infusion (N.S.). In further experiments, microvascular blood flow was measured by a laser-Doppler flow probe to compare the effects of intradermally injected PACAP, vasoactive intestinal polypeptide (VIP), and calcitonin gene-related peptide (CGRP). When injected into the skin of normal volunteers at 10(-12) to 10(-11) mol/site, each peptide caused a rapid flare lasting 2-3 min, which became erythematous after 5 min. At 10(-12) mol/site, intradermally injected PACAP and VIP caused a maximum increase in skin blood flow at 15 min of 379 +/- 96 and 307 +/- 121% (% increase above basal +/- SEM), respectively, and these responses were not significantly affected by oral aspirin (600 mg) taken 1.5 h beforehand. The vasodilation induced by PACAP at 10(-12) mol/site lasted approximately 6 h, whereas the effect of the same dose of CGRP and VIP lasted less than 2 h. These data suggest that PACAP is a potent and long-lasting vasodilator in humans.     

Differential regulation by Ca(2+) of calmodulin- and PKC-dependent contractile pathways in cat lower oesophageal sphincter

1. In the present investigation we examined the regulation of calmodulin (CaM)- and protein kinase C (PKC)-dependent pathways by cytosolic Ca(2+) in the contraction of cat lower oesophageal sphincter (LES). 2. Force developed in response to increasing doses of acetylcholine (ACh) was directly related to the increase of the [Ca(2+)](i) measured by fura-2. Thapsigargin, which depletes Ca(2+) stores, reduced the contraction and the [Ca(2+)](i). In addition, contraction in response to maximal ACh was reduced by the CaM inhibitor CGS9343B but not by the PKC inhibitor chelerythrine. The contraction in response to submaximal ACh was reduced by chelerythrine but not by CGS9343B. 3. In permeabilized cells, the contraction in response to low Ca(2+) (0.54 microm) was also reduced by CGS9343B. 4. The response to high Ca(2+) (1.0 microm) was reduced by CGS9343B. ACh also inhibited PKC activation induced by diacylglycerol, which activation is inhibited by the N-myristoylated peptide inhibitor derived from pseudosubstrate sequences of PKCalphabetagamma (myr-PKC-alphabetagamma), but not of myr-PKC-alpha. 5. These data are consistent with the view that activated CaM-dependent pathways inhibit PKC-dependent pathways, this switch mechanism might be regulated by Ca(2+) in the LES.     

Differential effects of baclofen on lower oesophageal sphincter pressure and proximal gastric motility in humans

BACKGROUND: The gamma-aminobutyric acid receptor type B agonist, baclofen, inhibits transient lower oesophageal sphincter relaxations by influencing a vagal pathway. Although post-prandial proximal gastric function, which is vagally mediated, is important in the occurrence of transient lower oesophageal sphincter relaxations, the effects of baclofen on post-prandial proximal gastric motility in humans remains undetermined. AIM: To determine the effects of baclofen on post-prandial lower oesophageal sphincter function and proximal gastric motility in healthy subjects. METHODS: In 11 healthy volunteers, a barostat bag and an oesophageal manometric catheter with a sleeve were simultaneously positioned; 40 mg of oral baclofen or placebo was then given in a randomized, double-blind manner. Subsequently, the intragastric bag volume, oesophageal and lower oesophageal sphincter pressure and oesophageal pH were recorded during the 90 min before and 120 min after a meal. RESULTS: During the post-prandial period, unlike the fasting period, baclofen decreased the rate of transient lower oesophageal sphincter relaxations and increased the basal lower oesophageal sphincter pressure compared with placebo. However, the meal-induced decrease in the tone and phasic contractility of the fundus was not affected by baclofen. CONCLUSION: The gamma-aminobutyric acid receptor type B agonist, baclofen, has a potent effect on post-prandial lower oesophageal sphincter motility without altering post-prandial proximal gastric motility, suggesting differential effects of baclofen on different signals of gastrointestinal vagal afferents.     

Effects of pituitary adenylate cyclase activating polypeptide27 on cyclic AMP efflux and atrial dynamics in perfused beating atria

Although pituitary adenylate cyclase activating polypeptide (PACAP) has been shown to increase cardiac force of contraction and to change the heart rate, the effect of PACAP on cyclic (c) AMP production in the atrium still has to be defined. In the present experiments, a simple protocol was developed for the evaluation of cAMP production in real-time base in the perfused beating left atria. The PACAP27-induced cAMP efflux in the atrial perfusate reflected changes in the production of cAMP in the atrial tissue. cAMP efflux was measured as an indicator of cAMP production in beating perfused rabbit atria. PACAP27 increased cAMP production in a dose- and time-dependent manner with a minor effect on atrial dynamics. These results suggest that PACAP27 has other roles besides control of force of contraction through cAMP production in the atrium.     

Mechanical and electrophysiological effects of endothelin-1 on guinea-pig isolated lower oesophageal sphincter circular smooth muscle.

1. The effects of endothelin-1 (ET-1) on guinea-pig lower oesophageal sphincter (LOS) circular smooth muscle were investigated by using intracellular microelectrodes and isometric tension recording techniques. 2. ET-1 produced biphasic mechanical responses; an initial transient relaxation followed by a sustained contraction. The initial relaxation was not inhibited by either tetrodotoxin (TTX, 1 microM) or L-N(G)-nitroarginine (L-NOARG, 100 microM). The sustained contraction was greatly attenuated by nifedipine (1 microM). 3. ET-1 (1 - 30 nM) induced a concentration-dependent hyperpolarisation that was unaffected by TTX or L-NOARG. The ET(A) receptor antagonist, BQ123 (0.3 microM) abolished the ET-1-induced hyperpolarisation, whereas the ET(B) receptor antagonist, BQ788 (0.3 microM) had no detectable effect. Sarafotoxin S6c (10 nM) did not change the membrane potential. 4. The ET-1-induced hyperpolarisation was abolished by apamin (0.1 microM). Interestingly, apamin abolished the ET-1-induced transient relaxation but potentiated the sustained contraction. 5. In Ca(2+)-free Krebs solution, the ET-1-induced hyperpolarisation was greatly attenuated and returned to the control value when the tissue was reperfused with Krebs solution containing Ca(2+). The ET-1-induced hyperpolarisation was insensitive to nifedipine but was attenuated by SK&F 96365 (1 - [beta-[3-(4 - methoxy - phenyl)propoxy] - 4 - methoxyphenethyl] - 1H-imidazole hydrochloride, 50 microM), an inhibitor of receptor-mediated Ca(2+) entry. The residual component of the ET-1-induced hyperpolarisation was sensitive to thapsigargin (1 microM). 6. These results demonstrate that, in guinea-pig LOS circular smooth muscle, ET-1 hyperpolarizes the membrane by activating apamin-sensitive K(+) channels, mainly as a result of receptor-mediated Ca(2+) entry and partly by Ca(2+) release from intracellular stores. The hyperpolarisation triggers the initial transient relaxation, which acts to oppose the sustained contraction.     

Localization and activity of haem oxygenase and functional effects of carbon monoxide in the feline lower oesophageal sphincter.

1. In the feline lower oesophageal sphincter (LOS), the distribution of the carbon monoxide (CO) producing enzymes haem oxygenase (HO)-1 and -2 was studied by immunohistochemistry and confocal microscopy, the HO activity was measured and the possible role for CO as a mediator of relaxation was investigated. 2. HO-2 immunoreactivity was abundant in nerve cell bodies of the submucosal and myenteric plexus. Approximately 50% of the HO-2-containing myenteric cell bodies were also nitric oxide synthase- and vasoactive intestinal peptide (VIP)-immunoreactive. In addition, HO-2 immunoreactivity was seen in nerve fibres, in non-neuronal cells dispersed in the smooth muscle and in arterial endothelium. HO-1 immunoreactivity was confined to non-neuronal cells in the smooth muscle, similar to those positive for HO-2. 3. Activity of HO, measured as CO production, was observed in LOS homogenates at a rate of 1.00 +/- 0.05 nmol mg-1 protein h-1. This production was inhibited by the HO inhibitor, zinc protoporphyrin-IX (ZnPP). 4. In isolated circular smooth muscle strips of LOS, developing spontaneous tone, exogenously administered CO evoked a concentration-dependent relaxation reaching a maximum of 93 +/- 3%. This relaxation was accompanied by an increase in cyclic GMP, but not cyclic AMP levels. The relaxant response was attenuated by methylene blue, but unaffected by tetrodotoxin. Repeated exposure to CO resulted in a progressive reduction of the relaxant response. 5. ZnPP caused a rightward-shift of the concentration-response curves for the relaxant responses to VIP, peptide histidine isoleucine, and pituitary adenylate cyclase activating peptide 27. 6. ZnPP and tin protoporphyrin-IX (another inhibitor of HO) did not affect nonadrenergic, noncholinergic relaxations induced by electrical field stimulation. Nor did ZnPP affect relaxations induced by 3-morpholino-sydnonimine or forskolin. 7. The present findings, showing localization of HO immunoreactivity to both neuronal and nonneuronal cells of the feline LOS, ability of LOS to produce CO and a relaxant effect of CO in circular LOS muscle, suggest a role for CO as a peripheral messenger.     

小脑颗粒细胞的高电压激活钙电流及多巴胺D4和垂体腺苷酸活化酶激活多肽

Cerebellar granule cells were a good mold for electrophysiologic studies at the single neuron level. Two distinct types of high-voltage-activated Ca2+ channels were present in cerebellar granule cells. These calcium channels change their expression, gating, and pharmacological properties during development, suggesting that calcium channel must be related to the processes of granule cell maturation and excitability. Dopamine inhibited L-type calcium current by activating D4 receptor, and this effect might involve another signaling system with the exception of cAMP system. The functional D4 receptor discovered in cerebellum not only gave a possibility to find other antipsychotics, but also supported the existence of a dopaminergic system in the granule cell involving the D4 receptor. Pituitary adenylate cyclase activating polypeptide (PACAP) could increase intracellular Ca2+ content by activation of Ca2+ channel and mobilization of intracellular Ca2+ stores. The effects were also cAMP-independent. Activating Ca2+ currents might be an important and necessary role of PACAP as a neurotropic factor involved in the control of multiplication, differentiation, and migration of granule cells.     

Effect of propranolol on the lower oesophageal sphincter in man

Summary

The effect of an intravenous dose (2?mg) of propranolol on the lower oesophageal sphincter was studied in 10 human volunteers. A hydraulic-capillary infusion manometric technique was used to measure the lower oesophageal sphincter pressure. A significant increase in sphincter pressure was recorded, together with increased amplitude and duration of oesophageal peristaltic activity. This study confirms the presence of specific beta-receptors in the lower oesophageal sphincter.