Levels of vasoactive intestinal peptide, cholecystokinin and calcitonin gene-related peptide in plasma and jejunum of rats following traumatic brain injury and underlying significance in gastrointestinal dysfunction

AIM: To study the alterations of brain-gut peptides following traumatic brain injury (TBI) and to explore the underlying significance of these peptides in the complicated gastrointestinal dysfunction. METHODS: Rat models of focal traumatic brain injury were established by impact insult method, and divided into 6 groups (6 rats each group) including control group with sham operation and TBI groups at postinjury 3, 12, 24, 72 h, and d 7. Blood and proximal jejunum samples were taken at time point of each group and gross observations of gastrointestinal pathology were recorded simultaneously. The levels of vasoactive intestinal peptide (VIP) in plasma, calcitonin gene-related peptide (CGRP) and cholecystokinin (CCK) in both plasma and jejunum were measured by enzyme immunoassay (EIA). Radioimmunoassay (RIA) was used to determine the levels of VIP in jejunum. RESULTS: Gastric distension, delayed gastric emptying and intestinal dilatation with a large amount of yellowish effusion and thin edematous wall were found in TBI rats through 12 h and 72 h, which peaked at postinjury 72 h. As compared with that of control group (247.8+/-29.5 ng/L), plasma VIP levels were significantly decreased at postinjury 3, 12 and 24 h (106.7+/-34.1 ng/L, 148.7+/-22.8 ng/L, 132.8+/-21.6 ng/L, respectively), but significantly increased at 72 h (405.0+/-29.8 ng/L) and markedly declined on d 7 (130.7+/-19.3 ng/L). However, Plasma levels CCK and CGRP were significantly increased through 3 h and 7 d following TBI (126-691% increases), with the peak at 72 h. Compared with control (VIP, 13.6+/-1.4 ng /g; CGRP, 70.6+/-17.7 ng/g); VIP and CGRP levels in jejunum were significantly increased at 3 h after TBI (VIP, 35.4+/-5.0 ng/g; CGRP, 103.8+/-22.1 ng/g), and declined gradually at 12 h and 24 h (VIP, 16.5+/-1.8 ng/g, 5.5+/-1.4 ng/g; CGRP, 34.9+/-9.7 ng/g, 18.5+/-7.7 ng/g), but were significantly increased again at 72 h (VIP, 48.7+/-9.5 ng/g; CGRP, 142.1+/-24.3 ng/g), then declined in various degrees on d 7 (VIP, 3.8+/-1.1 ng/g; CGRP, 102.5+/-18.1 ng/g). The CCK levels in jejunum were found to change in a similar trend as that in plasma with the concentrations of CCK significantly increased following TBI (99-517% increases) and peaked at 72 h. CONCLUSION: Traumatic brain injury can lead to significant changes of brain-gut peptides in both plasma and small intestine, which may be involved in the pathogenesis of complicated gastrointestinal dysfunction.     

Enteric nerves in diabetic rats: increase in vasoactive intestinal polypeptide but not substance P

The distribution of vasoactive intestinal polypeptide (VIP) and substance P-like immunoreactivities was studied by immunohistochemistry in the myenteric plexus and circular muscle layer of the ileum and proximal colon of rats 8 wk after induction of diabetes with streptozotocin. A consistent increase was observed in fluorescence intensity of VIP-like immunoreactivity in the nerve fibers, and intensely stained cell bodies were significantly more frequent in the myenteric plexus of the ileum (p less than 0.001) from diabetic animals. Some varicosities of VIP-like immunoreactive fibers in the myenteric plexus appeared to be enlarged. Vasoactive intestinal polypeptide-like immunoreactivity was increased and VIP-like immunoreactive nerves appeared thicker in the circular muscle layer of both diabetic ileum and proximal colon. The VIP levels were measured biochemically in tissue consisting of the smooth muscle layers and myenteric plexus. A significant increase in the VIP content per centimeter of intestine was found in both the ileum (p less than and proximal colon (p less than 0.01) from diabetic rats. In contrast, no apparent change in substance P innervation was observed immunohistochemically in the myenteric plexus and circular muscle layer of either diabetic ileum or proximal colon when compared with controls. The results are discussed in relation to the symptoms of autonomic neuropathy of the gut in diabetes.     

Effects of tactile and electrical stimuli upon release of vasoactive intestinal polypeptide in the mammalian penis

Plasma levels of vasoactive intestinal polypeptide (VIP) in the corpora cavernosa penis and dorsal penile veins greatly exceeded those measured in the limb or caudal veins during anaesthesia in various mammals (Bennett's wallaby, Barbary sheep, cheetah, puma, sooty mangabey, pigtail macaque and chimpanzee). Tactile stimulation of the penis immediately before or during collection of blood samples resulted in an increase. In the wallaby, VIP levels (mean +/- S.E.M.) in blood samples collected from the flaccid penis in the absence of tactile stimulation were very low (0.6 +/- 0.5 pmol/l). A 36-fold increase in VIP occurred after manual extension of the flaccid penis (24.8 +/- 3.2 pmol/l) or during manually stimulated erections (25.1 +/- 1.7 pmol/l). Electrical stimulation of erection produced no significant increase in VIP levels (2.3 +/- 0.9 pmol/l) unless accompanied by tactile stimulation (17.5 +/- 1.4 pmol/l). These studies provide the first demonstration that sensory feedback from the penis plays an important role in regulating vasoactive intestinal polypeptidergic activity. Since VIP is a potent vasodilator its release due to tactile stimuli during copulation may play a role in the maintenance of penile erection.     

Mechanisms of vasoactive intestinal peptide release in short-term culture of vasoactive intestinal peptide-producing tumor

Vasoactive intestinal peptide-producing tumor tissue fragments obtained at surgery were maintained in short-term culture. Functional cellular integrity of vasoactive intestinal peptide-producing tumor tissue was reflected by progressive protein synthesis and the ability of tumor tissue to release vasoactive intestinal peptide when stimulated by the intracellular second messengers cyclic adenosine monophosphate and calcium. Studies with verapamil and ethyleneglycol-bis (beta-aminoethylether)-N,N'-tetraacetic acid suggest that cyclic nucleotide- and ionophore A23187-mediated vasoactive intestinal peptide release are dependent, at least in part, upon the availability and transmembrane transport of extracellular calcium.     

Neuropeptide control of rat gastric mucosal blood flow. Increase by calcitonin gene-related peptide and vasoactive intestinal polypeptide, but not substance P and neurokinin A.

Submucosal blood vessels of the mammalian stomach are densely innervated by neurons containing calcitonin gene-related peptide (CGRP), substance P, neurokinin A, and vasoactive intestinal polypeptide (VIP). Because all these peptides are vasodilators in certain vascular beds, we tested the hypothesis that rat alpha-CGRP, rat VIP, substance P, and neurokinin A are candidate mediators of noncholinergic vasodilator neurons in the gastric mucosa and submucosa. The experiments were performed on urethane-anesthetized Sprague-Dawley rats. Gastric mucosal blood flow (GMBF) was measured by the hydrogen gas clearance technique, and the peptides were infused close arterially to the stomach via a catheter inserted retrogradely in the splenic artery. Basal GMBF was in the range of 35-50 ml/min/100 g. Infusion of rat alpha-CGRP (15 and 75 pmol/min) significantly increased GMBF in a dose-dependent manner, whereas mean arterial blood pressure was significantly lowered only by the higher dose of CGRP. Substance P (125 and 625 pmol/min) and neurokinin A (50 and 250 pmol/min) failed to alter GMBF, although the higher dose of each peptide led to a significant decrease in mean arterial blood pressure. Infusion of rat VIP (25 pmol/min) failed to affect GMBF and mean arterial blood pressure, whereas a fivefold higher dose of VIP (125 pmol/min) led to a significant rise of GMBF and to significant hypotension. These findings indicate that substance P and neurokinin A are unlikely to be of physiological significance for the regulation of GMBF. CGRP and VIP, however, can be considered as candidate mediators of submucosal nerve endings involved in the neural control of GMBF.(ABSTRACT TRUNCATED AT 250 WORDS)     

CGRP在大鼠胃痛觉过敏形成机制中的作用

目的:探索降钙素基因相关肽(CGRP)相关的干预措施对胃痛觉过敏的影响,了解CGRP在胃痛觉过敏形成过程中发挥的作用．方法:成年SD古大鼠,均植入胃内气囊．观察伤害性扩张或CGRP iv对大鼠疼痛阈值的影响:观察由上述措施诱发内脏过敏的大鼠在给予CGRP受体特异性拮抗剂hCGRP8-37后疼痛阈值的变化:观察不同剂量CGRP和hCGRP8-37对疼痛阈值的影响．结果:CGRP iv后胃疼痛阈值为11．7±2．6 mmHg,对照组疼痛阈值为19．2±2．0 mmHg,生理盐水对照组则为18．3±2．5 mmHg,实验组与其他两组比较尸均<0．05．CGRP使大鼠的疼痛阈值降低．hCGRP8-37能逆转伤害性扩张和CGRP引起的内脏敏感性增高,该作用呈剂量依赖性(r=0．821,P<0．01)．结论:胃扩张刺激能引起胃敏感性增高,在此过程中CGRP具有重要的作用．     

Cardiovascular effects of central calcitonin gene-related peptide in conscious rats

Calcitonin gene-related peptide (CGRP) is a potent peripheral vasodilator. In the present study, the cardiovascular effects of centrally administered CGRP were examined in conscious, normotensive rats. The rats were chronically instrumented with miniaturized pulsed Doppler flow probes to allow measurement of regional blood flow in the conscious animals. Injection of increasing doses of CGRP (0.3 to 3.0 micrograms/kg) in the lateral cerebroventricles transiently increased arterial pressure (maximal change = 13 +/- 3 mm Hg) and markedly increased heart rate (maximal increase = 88 +/- 10 b/min). The heart rate response was sustained over a period of 20 to 30 minutes. Central CGRP decreased hindquarter vascular resistance but had no effect on renal or mesenteric vascular resistances. In contrast, intravenous injections of CGRP reduced arterial pressure and renal, mesenteric, and hindquarter vascular resistances. The tachycardia response to central CGRP was attenuated by pretreatment with propranolol or hexamethonium, indicating that the heart rate response was mediated, in part, through increases in cardiac sympathetic tone. These data indicate that central CGRP may alter cardiovascular function through alterations in sympathetic outflow.     

Effect of passive immunization with antisera to vasoactive intestinal polypeptide and peptide histidine isoleucine amide on 5-hydroxy-L-tryptophan-induced prolactin release in rats

The present study was aimed to clarify, by use of the passive immunization method, the involvement of endogenous vasoactive intestinal polypeptide (VIP) and peptide histidine isoleucine amide (PHI)-like peptides in the stimulation of PRL-like immunoreactive material release induced by 5-hydroxy-L-tryptophan (5HTP), a serotonin precursor. We used conscious, freely moving male rats of the Wistar strain (BW, 250-300 g) chronically cannulated with atrial catheters. Anti-VIP serum (AVS) and anti-PHI serum (APS), each generated in rabbits against synthetic porcine VIP and natural porcine PHI, respectively, were highly potent [maximum binding capacity (Bmax): AVS, 55.5 nmol/ml; APS, 5.53 nmol/ml] and specific. Bolus injection of 5HTP (10 mg/kg BW) through the catheter caused a significant increase in plasma PRL (nanograms per ml) in rats pretreated with normal rabbit serum (NRS) [14.3 +/- (SE) 3.8----56.3 +/- 11.2], with AVS (12.3 +/- 3.5----48.5 +/- 6.2), with APS (10.5 +/- 3.9----43.5 +/- 8.8), and with AVS plus APS (9.0 +/- 1.4----28.5 +/- 2.7). The basal PRL concentrations did not differ significantly among these groups, whereas the PRL responses to 5HTP were significantly blunted in AVS plus APS-pretreated rats (P less than 0.05 vs. NRS). To eliminate the modification by dopaminergic control of 5HTP-induced PRL release, the next experiment was performed in rats repeatedly injected with sulpiride, a dopamine receptor antagonist (5 mg/kg BW), every 30 min. The first injection of sulpiride caused a prompt and marked increase in plasma PRL, followed by decreasing but still high levels of plasma PRL upon the subsequent injections of sulpiride every 30 min. The cumulative release area of PRL after pretreatment with AVS plus APS or APS alone was significantly lower than that after NRS (P less than 0.05). The same dose of 5HTP resulted in a significant further increase in plasma PRL exceeding the levels elevated by sulpiride injections in NRS-treated rats. Prior simultaneous administration of AVS and APS resulted in a complete suppression of 5HTP-induced PRL release, whereas pretreatment with either AVS or APS showed only a minimal effect. These results suggest that endogenous VIP and PHI-like peptides are PRL-releasing factors, involved at least in the mechanism of 5HTP-induced PRL release, in which the dopaminergic control may be also involved.     

Effects of vasoactive intestinal peptide and pancreatic polypeptide in rabbit intestine.

The effects of porcine vasoactive intestinal peptide (VIP) and bovine pancreatic polypeptide (PP) on jejunal, ileal, and colonic fluid transport were studied in the rabbit. VIP produced secretion in the small intestine (jejunum greater than ileum) but did not affect absorption in the colon. PP had no secretory effects in jejunum, ileum, or colon. The small intestinal secretion induced by VIP was not associated with raised cAMP concentrations in the mucosa; this suggests that the secretory effects of VIP in vivo are mediated by a mechanism other than stimulation of adenylate cyclase.     

Regional differences of adenylate cyclase stimulation by calcitonin and calcitonin gene-related peptide in the human kidney

Calcitonin (CT) gene-related peptide (CGRP)-like immunoreactivity was detected in both the cortex and medullo-papillary portion of human kidneys. The two forms of human CGRP as well as rat CGRP were capable of stimulating renal cortical adenylate cyclase activity in a concentration-related manner, with a half-maximally effective concentration (EC50) similar to that of human CT and approximately 100-1000 times higher than that of salmon CT. However, in the medullo-papillary portion, in which both salmon CT and human CT were inactive, the two forms of human and rat CGRP increased adenylate cyclase activity by 100%, with EC50 values ranging from 36 nmol/L to 1 mumol/L. In cortical membrane preparations the effect of CGRP was additive to that of salmon CT. We concluded that regional differences exist in the effect of CT and CGRP in human renal tissue and that in the medullo-papillary portion and possibly in the cortex, CGRP stimulates adenylate cyclase activity through a CT-independent mechanism.     

Effects of in vivo stimulation on molecular forms of circulatory calcitonin and calcitonin gene-related peptide in man

Multiple molecular weight forms of the potent vasodilator peptide calcitonin gene-related peptide (CGRP) are present in the circulation in man. The monomeric form of CGRP(1-37) and CT(1-32) account for only a fraction of the total immunoreactive CGRP and calcitonin (CT) in the circulation. In vivo release of various molecular weight forms of CGRP and CT, including the monomeric form, into the circulation was demonstrated by gel-permeation chromatography, following stimulation of the gastrointestinal tract by a meal, and C-cells of the thyroid gland with pentagastrin. Following these stimulations an increase of the levels of C-terminal fragment of CGRP was also observed indicating a rapid metabolic breakdown of CGRP in vivo. In addition to a number of endogenous peptides and other substances, acute release of monomeric CGRP into the circulation may also contribute to the vasodilation observed following ingestion of food and administration of pentagastrin in man.     

Plasma levels of vasoactive intestinal polypeptide and oxytocin in response to suckling, electrical stimulation of the mammary nerve and oxytocin infusion in rats

The aim of the present study was to measure plasma levels of vasoactive intestinal polypeptide (VIP) and oxytocin following suckling, electrical stimulation of the mammary nerve and oxytocin infusion in lactating rats. Trunk blood was collected by decapitation after 5 and 20 min of suckling in conscious lactating rats. Repeated blood samples were drawn from the carotid artery in anesthetized rats, in connection with suckling, oxytocin infusion (0.22 nmol/l/kg/h) and electrical stimulation of the mammary nerve (5 V, 5 Hz, 2 ms). VIP and oxytocin levels were measured by radioimmunoassay. In conscious rats, VIP levels rose significantly from 18 +/- 5 to 102 +/- 30 pM after 5 min of suckling and to 123 +/- 25 pM after 20 min of suckling when milk ejection occurred. Oxytocin levels rose significantly from 90 +/- 24 to 269 +/- 45 pM during milk ejection. Suckling, oxytocin infusion and mammary nerve stimulation in anesthetized rats raised VIP levels significantly from 13 +/- 2, 18 +/- 5 and 10 +/- 2 to 43 +/- 8, 45 +/- 16 and 53 +/- 22 pM, respectively, whereas oxytocin levels rose from 111 +/- 34 to 294 +/- 66 pM after 20 min of suckling and to a peak value of 500 +/- 70 pM after oxytocin infusion. This study shows that VIP is elevated in plasma in lactating rats when the pups are suckling. The results showing that VIP levels rise following mammary nerve stimulation and oxytocin infusions indicate that both neurogenic and hormonal mechanisms can contribute to the regulation of VIP levels in plasma.     

The effect of calcitonin gene-related peptide on healing of intestinal anastomosis in rats with experimental obstructive jaundice

Purpose

Intestinal anastomotic healing is a complex procedure in which several mediators and cytokines play roles. Calcitonin gene-related peptide is an important neuropeptide in inflammation. In this study we aimed to investigate the effect of calcitonin gene-related peptide on healing of intestinal anastomosis in rats with obstructive jaundice.

Materials and methods

Obstructive jaundice was induced in rats by the ligation and division of the common bile duct. Four days after the operation, intestinal anastomosis was performed, and either calcitonin gene-related peptide or 0.9% NaCl was administered intraperitoneally to these jaundiced rats and controls. The concentrations of serum tumor necrosis factor-α (TNF-α) and triglyceride levels of all rats were measured, and healing of the anastomosis was evaluated by measuring the bursting pressure and hydroxyproline content on the 7th postoperative day.

Results

Calcitonin gene-related peptide was found to have positive effects on healing of the anastomosis by inhibiting the effects of TNF-α and increasing the bursting pressure and hydroxyproline content of the anastomosis.

Conclusion

Calcitonin gene-related peptide increases anastomotic wound healing in experimental anastomosis in the presence of obstructive jaundice in rats.     

Administration of antisera to vasoactive intestinal polypeptide and peptide histidine isoleucine attenuates ether-induced prolactin secretion in rats

The effect of immunoneutralization of endogenous vasoactive intestinal polypeptide (VIP) and peptide histidine isoleucine (PHI)-like peptides by administration of specific and potent antisera to the respective peptides on ether stress-induced prolactin (PRL) release was examined in male rats bearing an indwelling atrial catheter. Forty-five minutes after an injection of 1 ml of either normal rabbit serum (NRS), anti-VIP serum (AVS), anti-PHI serum (APS) or AVS plus APS, the rat was placed for 1 min in a beaker containing an ether-impregnated cotton ball. Ether exposure caused a prompt and significant increase in plasma PRL in all of the animal groups tested. Pretreatment with either antisera did not affect basal plasma PRL levels, whereas plasma PRL rises after ether exposure were significantly lower in rats pretreated with AVS, APS or AVS plus APS than those with NRS. These results suggest that hypothalamic VIP and PHI-like peptides may be involved, at least in part, in the mechanism by which ether stress stimulates PRL secretion in rats.     

Concentration of substance P, neurokinin A, calcitonin gene-related peptide, neuropeptide Y and vasoactive intestinal polypeptide in synovial fluid from knee joints in patients suffering from rheumatoid arthritis

We have studied the presence of five neuropeptides in knee joint synovial fluid from either patients suffering from rheumatoid arthritis and pain (n = 18) or being subjected to arthroscopy due to meniscal/cruciate ligament injuries (n = 13). Radioimmunoassay technique was used for peptide analysis using antisera SP2 against substance P (SP), K12 against neurokinin A (NKA), CGRPR8 against calcitonin gene-related peptide (CGRP), NPY1 against neuropeptide Y (NPY) and VIP2 against vasoactive intestinal polypeptide (VIP). No SP could be detected, and lower levels of NKA was found in arthritic joints vs controls. CGRP and NPY was found in higher concentrations in arthritic patients vs controls. VIP was found sporadically in both arthritis and control patients. Our data show some quantitative differences between patients suffering rheumatoid arthritis and pain, and patients with non-inflamed joints without pain; indicating an involvement of peptidergic fibers in arthritis in humans.     

Inhibitory action of islet amyloid polypeptide and calcitonin gene-related peptide on release of insulin from the isolated perfused rat pancreas

Islet (or insulinoma) amyloid polypeptide (IAPP) is a 37-residue peptide recently purified from amyloid deposits in the pancreas of patients with type 2 diabetes and from amyloid deposits of a human insulinoma. IAPP immunoreactivity has been identified in islet B cells of diabetic and nondiabetic humans. IAPP is structurally similar to calcitonin gene-related peptide (CGRP). The purpose of this study was to examine the effects of IAPP and CGRP on glucose- and carbachol-stimulated release of insulin and pancreatic polypeptide (PP) from the isolated perfused rat pancreas. IAPP and CGRP, at 10(-7) M, failed to inhibit glucose-stimulated (16.7 mM) release of insulin. At the same concentration, however, IAPP significantly (p less than 0.05) inhibited carbachol-stimulated (10(-7) M) release of insulin by 30%, and CGRP significantly inhibited carbachol-stimulated release of insulin by 33% when compared with the control group. IAPP also significantly decreased carbachol-stimulated release release of PP. IAPP and CGRP, at 10(-8) M, did not inhibit carbachol-stimulated release of insulin and PP. These results suggest that IAPP and CGRP may have roles in the regulation of secretion of insulin. IAPP may inhibit secretion of insulin, at least in part, by blocking cholinergic mechanisms.     

Effects of the three neuropeptides, vasoactive intestinal peptide, substance P, and calcitonin gene related peptide on basal and stimulated calcitonin release in the rat

Calcitonin is secreted from the thyroidal C-cells. Except that calcitonin secretion is stimulated by calcium, little is known of its regulation. Vasoactive intestinal peptide (VIP), substance P, and calcitonin gene related peptide (CGRP) have recently been detected within intrathyroidal neurons, and CGRP also within the C-cells, and may therefore affect calcitonin secretion. In this study, we investigated whether VIP, substance P or CGRP could influence calcitonin secretion in the rat. Each of these peptides was administered as a single injection (1.5 nmol/animal) or as a 30-min infusion (1.5 nmol/animal per 30 min) during which calcium chloride, 456 mumol/animal, was injected iv. We found that the peptides had no effect on basal calcitonin secretion, but that VIP potentiated the calcium-induced calcitonin release. Thus, the peak plasma calcitonin level following calcium chloride injection was doubled by the infusion of VIP (P less than 0.001). In contrast, neither substance P nor CGRP significantly influenced the calcium-induced calcitonin release. We conclude that VIP, a neuropeptide within intrathyroidal nerves, has the capacity to augment calcium-induced calcitonin secretion in the rat and we therefore suggest that VIP is a regulator of calcitonin secretion.     

Serotonin,calcitonin and calcitonin gene-related peptide in acute pancreatitis

Objective: The aim of this study was to investigate plasma levels of serotonin, calcitonin and calcitonin gene-related peptide (CGRP) in the course of acute pancreatitis (AP) taking organ failure, etiology and severity into consideration.

Material and methods: Sixty consecutive patients with alcohol- or gallstone-induced AP were included over a 15-month period. Patients were treated according to a standardized algorithm and monitored for organ specific morbidity and mortality. Organ functions and blood samples were assessed on days 0, 1, 2 and 14 after hospital admission. Twenty healthy volunteers, matched for age and gender, comprised the reference group.

Results: Lower levels of serotonin were observed in patients at admission compared to healthy volunteers (p?=?.021). Serotonin levels increased from day 2 to 14 (p?<?.001), but with no relation to severity, etiology or organ failure. No difference in calcitonin levels was found in patients at admission compared to healthy volunteers. However, calcitonin levels decreased over time (p?<?.001) and higher levels were found in patients with respiratory failure (p?=?.039). No difference was observed in relation to severity or etiology. CGRP levels in patients at admission did not differ from healthy volunteers, nor did CGRP change over time or show any relationship to severity, etiology or organ failure.

Conclusion: Our data suggest serotonin and calcitonin levels to be associated to time-course of AP, and calcitonin levels to organ dysfunction. We hypothesize that serotonin plays a pathogenic role in the compromised pancreatic microcirculation, and calcitonin a role as a biomarker of severity in AP.