Dendritic outgrowth of myenteric plexus neurons in primary culture.

Myenteric plexus neurons derived from neonatal guinea pigs, when exposed to serum, demonstrated a characteristic pattern of growth, including a proliferating outgrowth zone of glial cells, peripheral extension of dendritic processes, and progressive dendritic growth. Serum effects upon dendritic growth, measured morphometrically, was strongly dose- and temporally dependent. Dendritic density was increased 10-fold (120 hr) by the addition of 6% serum, while mean dendritic length was increased 3-fold. Development of cholinergic function was reflected by release of [3H]ACh in response to cholecystokinin octapeptide, vasoactive intestinal peptide, substance P, and calcitonin gene-related peptide (10(-10) and 10(-8) M).     

Inhibition by beta-CCM of cholecystokinin-induced release of acetylcholine from the longitudinal muscle-myenteric plexus preparation in the guinea-pig ileum

The antagonism between cholecystokinin (CCK) and methyl beta-carboline-3-carboxylate (beta-CCM) in the nervous system was studied by measuring the release of acetylcholine (ACh) from the longitudinal muscle-myenteric plexus preparation of guinea-pig. The ACh release was assessed by measuring [3H] output from the preparation preincubated with [3H] choline. Thirty mM of KCl caused a pronounced release of [3H] ACh from the preparation. Sulfated cholecystokinin octapeptide (CCK8) also increased the release of [3H] ACh in a dose-dependent manner at concentrations ranging from 10(-10)M to 10(-8)M. CCK8 at a concentration of 10(-8)M released [3H] ACh by about 300% of the 30 mM KCl-induced [3H] ACh release. In the presence of beta-CCM (10(-8)M to 10(-4)M), the release of [3H] ACh by KCl was not affected, but that by CCK8 was significantly inhibited depending on the concentrations of beta-CCM. These results show that the action of CCK8 to stimulate neurons in the myenteric plexus can be selectively antagonized by beta-CCM.     

Calcitonin gene-related peptide inhibits gallbladder contractility

Calcitonin gene-related peptide (CGRP), a 37-amino acid peptide, is widely distributed in the gastrointestinal tract where it is colocalized with substance P. The effect of CGRP on gallbladder motility is unknown. The objective of this study was to examine the effect of CGRP on cholecystokinin-octapeptide (CCK-8) and substance P-stimulated gallbladder contraction in vivo and in vitro. In in vivo studies intragallbladder pressure was measured in response to bolus administration of CCK-8 (10(-15) to 10(-9) mol/kg) or substance P (10(-12) to 10(-7) mol/kg), either alone or with a continuous infusion of CGRP (10(-9) mol/kg/hr), in anesthetized guinea pigs. In in vitro studies the contractile force of guinea pig gallbladder muscle strips was examined in response to CCK-8 (10(-12) to 10(-7) mol/L) and substance P (10(-9) to 10(-6) mol/L), alone or with CGRP (10(-10) to 10(-6) mol/L). CGRP (10(-9) mol/kg/hr) inhibited in vivo gallbladder contraction that was stimulated by CCK-8, but not by substance P. CGRP alone produced a significant (p less than 0.05) dose-related decrease in the resting tension of gallbladder strips in vitro. CGRP (10(-6) mol/L) inhibited gallbladder muscle tension in vitro, stimulated by both CCK-8 and substance P. These studies show that CGRP can affect gallbladder motor activity by decreasing smooth muscle tone and that CGRP can antagonize the action of CCK and substance P. CGRP may be involved in the physiologic control of gallbladder emptying and refilling.     

Release and effects of calcitonin gene-related peptide in myocardial ischaemia

This thesis is based on the following papers, which will be referred to by their Roman numerals. I. Franco-Cereceda A, Källner G, Lundberg JM: Capsazepine-sensitive release of calcitonin gene-related peptide from C-fibre afferents in the guinea-pig heart by low pH and lactic acid. Eur J Pharmacol 1993;238:311-316. II. Franco-Cereceda A, Källner G, Lundberg JM: Cyclo-oxygenase products released by low pH have capsaicin-like actions on sensory nerves in the isolated guinea pig heart. Cardiovasc Res 1994;28:365-369. III. Källner G, Franco-Cereceda A: Attenuation of low pH-, but not capsaicin- or PGI-evoked CGRP-release by endothelium removal using saponin. Acta Physiol Scand 1995;155:251-256. IV. Källner G, Franco-Cereceda A: Ion channels involved in the release of calcitonin generelated peptide by low pH, prostacyclin and capsaicin in the isolated guinea-pig heart. Eur J Pharmacol 1998;in press. V. Källner G, Gonon A, Franco-Cereceda A: Calcitonin gene-related peptide in myocardial ischaemia and reperfusion in the pig. Cardiovasc Res 1998;38:493-499. VI. Källner G, Franco-Cereceda A: Aggravation of myocardial infarction in the porcine heart by capsaicin-induced depletion of calcitonin gene-related peptide (CGRP). J Cardiovasc Pharmacol , accepted for publication. VII. Källner G, Öwall A, Franco-Cereceda A: Myocardial outflow of calcitonin gene-related peptide in relation to metabolic stress during coronary artery bypass grafting without cardiopulmonary bypass. Submitted for publication.     

Inhibition of parietal cell H+ secretion by transforming growth factor alpha: a possible autocrine regulatory mechanism

Transforming growth factor-alpha (TGF alpha) and TGF alpha/epidermal growth factor receptor messenger ribonucleic acid have recently been demonstrated in isolated parietal cells. The aim of this study was to investigate the effects of TGF alpha on basal and stimulated secretion in vitro with the isolated rabbit parietal cell model. Acid secretion was assessed indirectly with cell uptake of carbon 14-labeled aminopyrine [( 14C]AP). TGF alpha (10(-11) to 10(-7) mol/L) had no effect on unstimulated [14C]AP uptake. TGF alpha dose dependently inhibited histamine (10(-5) to 10(-6) mol/L)-stimulated but not forskolin (10(-5) to 10(-7) mol/L)-stimulated [14C]AP uptake. This effect on histamine-stimulated activation was reversed by pertussis toxin (200 ng/ml) before incubation. TGF alpha had no effect on carbachol (10(-5) to 10(-6) mol/L)-stimulated [14C]AP uptake. Specific HCO3-buffer studies demonstrated that these observations were independent of extracellular buffer and possible TGF alpha effects on intracellular pH. Our data indicate that TGF alpha inhibits acid secretion by specifically uncoupling histamine/cyclic adenosine monophosphate transduction at the guanosine triphosphate-binding protein. TGF alpha, unlike epidermal growth factor, has no effect on carbachol stimulation, which suggests a qualitative difference between the biologic actions of TGF alpha and epidermal growth factor. Possible autocrine-paracrine modulation of histamine stimulation by TGF alpha invokes a novel regulatory mechanism of parietal cell secretion.     

Natriuretic peptide responsive, cyclic guanosine monophosphate producing structures in the guinea pig bladder

PURPOSE: We examined the localization of natriuretic peptide responsive, cyclic guanosine monophosphate producing cells in the guinea pig bladder. MATERIALS AND METHODS: The bladder was removed from male guinea pigs sacrificed by cervical dislocation. The lateral wall of the bladder was cut into strips 2 mm thick. The tissue pieces were incubated in the presence of human atrial natriuretic peptide, rat brain natriuretic peptide and C-type natriuretic peptide or the nitric oxide donor DEANO (diethylamine NONOate or 1,1-diethyl-2-hydroxy-2-nitrosohydrazine) (Sigma). Cyclic guanosine monophosphate immunoreactivity was localized using an antibody against formaldehyde fixed cyclic guanosine monophosphate. RESULTS: Atrial natriuretic peptide and brain natriuretic peptide stimulated cyclic guanosine monophosphate synthesis in suburothelial interstitial cells, whereas C-type natriuretic peptide was not effective. In contrast, DEANO stimulated cyclic guanosine monophosphate synthesis in urothelial umbrella cells, suburothelial interstitial cells, muscle interstitial cells and neurons. The effect of atrial natriuretic peptide and brain natriuretic peptide was not inhibited by ODQ (1H-[1, 2, 4]oxadiazolo[4-3a]quinoxalin-1-one), an inhibitor of nitric oxide responsive soluble guanylyl cyclase. CONCLUSIONS: To our knowledge our findings show for the first time a localized effect of atrial natriuretic peptide and brain natriuretic peptide to the suburothelial cells of the guinea pig bladder. These cells express the soluble guanylyl cyclase and particulate guanylyl cyclase-A isoforms. The specific physiological role of these cells is not known but it was suggested that they may be involved in the generation or modulation of sensation. The results imply a role for natriuretic peptide-cyclic guanosine monophosphate signaling in the processing of sensory information in the bladder.     

Involvement of TRPV1-containing peripheral sensory efferents in hemodynamic responses in a rat hemorrhagic shock model

BackgroundMechanisms underlying hemodynamic disturbance in hemorrhagic shock are not completely understood. Transient receptor potential vanilloid 1–expressing afferents are involved in hemorrhagic shock pathology, and transient receptor potential vanilloid 1 antagonist, capsazepine, acts on the central nervous system to improve mortality in a rat hemorrhagic shock model. In contrast, transient receptor potential vanilloid 1-positive efferents promote vasoactive reactions through the release of neuropeptides, including calcitonin gene-related peptides. This study aimed to investigate whether transient receptor potential vanilloid 1-positive peripheral sensory efferents are involved in hemodynamic responses after hemorrhagic shock.MethodsMale rats underwent hemorrhagic shock (mean arterial pressure 30 mm Hg for 90 min, followed by resuscitation for 30 min) and received capsazepine (5 μM/kg) 30 min after shock induction. A separate cohort of rats subjected to hemorrhagic shock received hCGRP_8-37 (300 μg/kg), a calcitonin gene-related peptide receptor antagonist, at 30, 60, or 90 minutes after shock induction. The 24-hour survival rate, mean arterial pressure, heart rate, arterial blood gas, and plasma calcitonin gene-related peptide levels were measured. Tissue blood flow and oxygenation both in the mesentery and skeletal muscle were also assessed.ResultsCapsazepine treatment prevented the hemorrhagic shock-induced increase in plasma calcitonin gene-related peptide levels, and hCGRP_8-37 treatment improved the 24-h survival rates after hemorrhagic shock at a time-dependent manner. The hCGRP_8-37- or capsazepine-treated rats exhibited tissue oxygenation and metabolic conditions comparable to those in control rats at the end of the experiment.ConclusionTransient receptor potential vanilloid 1 plays a crucial role in hemodynamic responses to hemorrhagic shock, partly via calcitonin gene-related peptide release, involved in its peripheral sensory-efferent functions. The hCGRP_8-37 appears to improve peripheral circulatory failure, which may be useful as adjunct treatment after hemorrhagic shock.     

Effect of vasoactive intestinal peptide on the motility of guinea-pig colon in vitro

The effects of vasoactive intestinal peptide (VIP) on longitudinal and circular muscle strips of guinea-pig proximal and distal colons, and on propulsive activity of guinea-pig distal colon were investigated in vitro. VIP (10(-9)-10(-6) M) produced relaxations of longitudinal and circular muscle strips in proximal colon and of circular muscle strip in distal colon, but produced a contraction of longitudinal muscle strip in distal colon. VIP-induced responses of the muscle strips were not influenced by indomethacin (10(-6) M). Tetrodotoxin (10(-6) M) and atropine (10(-6) M) converted VIP-induced contraction into relaxation in longitudinal muscle strip of distal colon, although these nerve blockers did not influence VIP-induced relaxations of longitudinal and circular muscle strips in proximal colon and of circular muscle strip in distal colon. VIP (10(-6) M) inhibited spontaneous and carbachol (10(-8) M)-stimulated propulsive activities of the isolated segment in distal colon. These results suggest that VIP may directly relax colonic smooth muscle cells and may indirectly contract longitudinal muscle strip of distal colon, mainly via stimulation of cholinergic neurones in the myenteric plexus of the muscle strip. It is also suggested that VIP-induced watery diarrhea in WDHA syndrome may not due to a direct stimulation of colonic motility.     

NK2 tachykinin receptors mediate contraction of the pig intravesical ureter: tachykinin-induced enhancement of non-adrenergic non-cholinergic excitatory neurotransmission

The current study was designed to characterize the functionally active tachykinin receptors involved in tachykinin-elicited contractions in the pig intravesical ureter, and to investigate the possible modulation exerted by the natural tachykinins substance P (SP) and neurokinin A (NKA) on the non-adrenergic non-cholinergic (NANC) excitatory ureteral neurotransmission. In pig intravesical ureteral strips pretreated with phosphoramidon (10(-5) mol/L) to block the endopeptidase activities, isometric force recordings showed that SP, NKA, and the NK2 receptor selective agonist [beta-Ala(8)]-NKA (4-10), all three induced contractions, with the following potency order: NKA > [beta-Ala(8) ]-NKA (4-10) > SP. [Sar(9), Met(O(2))(11)]-SP and senktide, selective agonists of the NK1 and NK3 receptors, respectively, failed to modify the ureteral tone. Urothelium removal and incubation with tetrodotoxin (10(-6) mol/L), phentolamine (10(-7) mol/L), propranolol (3 x 10(-6) mol/L), atropine (10(-7) mol/L) and indomethacin (3 x 10(-6) mol/L), did not alter the contraction induced by a submaximal (10(-7) mol/L) dose of [beta-Ala(8)]-NKA (4-10). MEN 10,376 (10(-8)-10(-7) mol/L), a NK2 receptor antagonist, reduced the contraction to 3 x 10(-8) mol/L NKA. GR 82334 (10(-6) -10(-5) mol/L) and SR 142801 (10(-8)-10(-7) mol/L), selective antagonists of the NK1 and NK3 receptors, respectively, did not modify that contraction. In pig intravesical ureteral strips in NANC conditions, SP and NKA induced a potentiation of the contractions to electrical field stimulation (EFS) and to exogenous ATP. The results suggest that the tachykinins evoke a direct contraction of pig intravesical ureteral strips through NK2 receptors located in the smooth muscle. SP and NKA exert an enhancement of the NANC excitatory neurotransmission of the pig intravesical ureter.     

Mechanism mediating nitric oxide-induced inhibition in human jejunal longitudinal smooth muscle

BACKGROUND: Enteric neurotransmission is a complex process involving multiple neurotransmitters, including nitric oxide (NO). Our aim was to evaluate the role and mechanism(s) of action of NO in normal human jejunal longitudinal smooth muscle. METHODS: Transmural strips of normal human jejunum obtained from subjects undergoing gastric bypass were studied in organ chambers. Effects of exogenous NO (7 x 10(-6) mol/L to 7 x 10(-5) mol/L) and electrical field stimulation (nonspecific release of endogenous neurotransmitters) on spontaneous contractile activity and on precontracted muscle strips (substance P, 10(-5) mol/L) were evaluated in the presence and absence of the competitive NO synthase inhibitor N(G)-amino-L-arginine (L-NNA, 10(-3) mol/L) and the specific soluble guanylyl cyclase inhibitor 1H-[1,2,4]-oxadiazaolo-[4,3-a]-quinoxalin-1-one (ODQ, 10(-5) mol/L and 10(-4) mol/L). RESULTS: Exogenous NO dose-dependently inhibited spontaneous contractility and relaxed precontracted smooth muscle strips. The effects of NO were markedly attenuated or completely inhibited in the presence of ODQ. Electric field stimulation under nonadrenergic, noncholinergic conditions also inhibited spontaneous contractility and relaxed precontracted smooth muscle strips; both of these effects were attenuated, but not completely inhibited, in the presence of both ODQ and L-NNA. CONCLUSIONS: NO is an endogenous inhibitory neurotransmitter in human jejunal longitudinal smooth muscle, acting at least in part via a mechanism mediated by guanylyl cyclase. Other (non-nitrergic) nonadrenergic, noncholinergic inhibitory neurotransmitters are likely active in this portion of the human gut.     

Hypergastrinemia after vagotomy is not associated with decreased gastric somatostatin

Although hypergastrinemia occurs after vagotomy, the mechanisms responsible are not understood. Somatostatin (SRIF) is a peptide that inhibits gastrin release, is present within the gastric fundus and antrum, and is under vagal control. In this study we have investigated the hypothesis that hypergastrinemia is associated with a decrease in gastric SRIF. We examined tissue levels of SRIF in gastric mucosa and muscle wall in rabbits undergoing vagotomy and pyloroplasty compared with sham-operated controls. We also compared the release of SRIF from gastric glands in response to vasoactive intestinal peptide and calcitonin gene-related peptide. Vagotomy resulted in an increase in gastrin compared with controls in both antrum (1062 +/- 176 pmol/gm vs 484 +/- 48 pmol/gm) and plasma (236 +/- 72 pmol/L vs 29 +/- 4 pmol/L; p less than 0.05). This was accompanied by an increase in the number of gastrin cells (25 +/- 4 vs 9 +/- 3; p less than 0.05). No significant differences were observed in gastric SRIF levels in either the fundus or antrum (p greater than 0.5). In addition, there were no differences in the release of SRIF from gastric glands of vagotomized animals compared with controls in response to vasoactive intestinal peptide and calcitonin gene-related peptide (p greater than 0.5). These data suggest that the elevations in plasma and antral gastrin levels after vagotomy are not a result of alterations in gastric SRIF.     

A role for calcitonin gene-related peptide in protection against gastric ulceration.

OBJECTIVE: The goal of this investigation was to determine the role of calcitonin gene-related peptide (CGRP) in gastric mucosal resistance to ulceration. SUMMARY BACKGROUND DATA: CGRP is a 37-amino acid peptide found in the peripheral ends of afferent gastric neurons. CGRP is known to inhibit acid secretion, stimulate mucosal blood flow, and stimulate release of somatostatin. METHODS: The release of CGRP in response to intragastric and intra-arterial administration of capsaicin in the isolated, vascularly perfused rat stomach was measured by radioimmunoassay. The molecular forms of CGRP released were analyzed by gel filtration chromatography. The effect of intravenous CGRP or intragastric capsaicin on gastric ulceration induced by 100 mmol/L HCl and indomethacin was studied in intact and endogenous CGRP-depleted rats. RESULTS: Intra-arterial capsaicin (concentration range, 10(-7) to 10(-5) mol/L) stimulated a prompt and sustained release of immunoreactive CGRP, of which 84% coeluted with rat 1-37 CGRP I by gel filtration. Intragastric capsaicin (range, 10(-5) to 10(-4) mol/L) failed to release CGRP into the vascular perfusate. In intact rats, intragastric capsaicin (10(-6) mol/L) or intravenous CGRP I (10 micrograms/kg/hr) reduced the number and area of mucosal lesions caused by HCl and indomethacin compared with the findings in control rats. Rats depleted of endogenous CGRP were more susceptible to gastric ulceration than were normal rats. Intragastric capsaicin failed to protect the mucosa of CGRP-depleted rats, whereas exogenous intravenous CGRP was effective. CONCLUSIONS: These data support the hypothesis that CGRP released from gastric enteric neurons mediates gastric mucosal resistance to ulceration by noxious agents.     

Drug Insight: biological effects of botulinum toxin A in the lower urinary tract

Botulinum toxins can effectively and selectively disrupt and modulate neurotransmission in striated muscle. Recently, urologists have become interested in the use of these toxins in patients with detrusor overactivity and other urological disorders. In both striated and smooth muscle, botulinum toxin A (BTX-A) is internalized by presynaptic neurons after binding to an extracellular receptor (ganglioside and presumably synaptic vesicle protein 2C). In the neuronal cytosol, BTX-A disrupts fusion of the acetylcholine-containing vesicle with the neuronal wall by cleaving the SNAP-25 protein in the synaptic fusion complex. The net effect is selective paralysis of the low-grade contractions of the unstable detrusor, while still allowing high-grade contraction that initiates micturition. Additionally, BTX-A seems to have effects on afferent nerve activity by modulating the release of ATP in the urothelium, blocking the release of substance P, calcitonin gene-related peptide and glutamate from afferent nerves, and reducing levels of nerve growth factor. These effects on sensory feedback loops might not only help to explain the mechanism of BTX-A in relieving symptoms of overactive bladder, but also suggest a potential role for BTX-A in the relief of hyperalgesia associated with lower urinary tract disorders.     

Effect of calcitonin gene-related peptide on anastomotic healing in the presence of endotoxin

BACKGROUND: Intestinal anastomotic healing is a complex procedure in which several mediators, cytokines and other substances play roles, as well as calcitonin gene-related peptide (CGRP). CGRP is capable of stimulating DNA synthesis and cell proliferation in endothelial cells by increasing vasodilatation and inflammatory response and promoting epithelial, vascular and mesothelial cell proliferation. This study was undertaken to investigate whether CGRP has a beneficial effect on intestinal anastomotic healing, even in septic conditions. METHODS: Four groups of 10 rats were administered normal saline (0.5 mL), lipopolysaccharide (LPS) (0.5 mg/kg), CGRP (0.5 mL 6.5 x 10(-10) mol/L) and LPS + CGRP (0.5 mg/kg + 0.5 mL 6.5 x 10(-10) mol/L) via intraperitoneal route, respectively, 24 h prior to operation and postoperatively. All rats underwent ileo-ileal end-to-end anastomosis. Anastomotic bursting pressure and tissue hydroxyproline levels were measured on postoperative day 7. RESULTS: Calcitonin gene-related peptide was found to have positive effects on both parameters of healing. The LPS-injected group showed intestinal anastomotic healing disorder suggesting impaired collagen production, which showed improvement after CGRP administration. CONCLUSIONS: Calcitonin gene-related peptide increases anastomotic wound healing in experimental intestinal anastomosis in the presence of endotoxin.     

Basal and acetylcholine-stimulated nitric oxide formation mediates relaxation of rabbit cavernous smooth muscle

Externally applied acetylcholine (ACh) in human corpus cavernosum has been shown to cause endothelium-dependent smooth muscle relaxation. Changes in isometric tension in rabbit cavernous smooth muscle strips mounted in organ bath chambers were monitored in the presence of blocking agents. Nitric oxide (NO) is known as an endothelium-derived relaxation factor (EDRF). Addition of specific inhibitors of nitric oxide synthesis, such as L-n-monomethyl arginine (L-NMMA) at 5 x 10(-4) mol/l.. or L-n-nitro arginine (L-NOARG) at 2 x 10(-4) mol/l. to strips precontracted with phenylephrine (PE) at 3.16 x 10(-6) mol/l. led to significant increases in tension. In the presence of L-NMMA or L-NOARG, relaxing effects of ACh at 10(-8)-3.16 x 10(-5) mol/l. mediated by muscarinic receptors were almost completely abolished. These data indicate that rabbit cavernous smooth muscle is under the control of basal NO release. They constitute strong evidence that cholinergically induced endothelial formation of NO plays a crucial role in relaxing cavernous smooth muscle.     

17β-雌二醇对豚鼠胆囊平滑肌收缩的影响

目的 观察17p-雌二醇对豚鼠胆囊平滑肌的基因组和非基因组效应,初步探究其机制。方法 构建去卵巢豚鼠动物模型,皮下注射17β-雌二醇,检测各组豚鼠血清雌二醇和八肽胆囊收缩素(CCK-8)含量;采用张力换能器观察17p-雌二醇对胆囊离体肌条收缩的影响。观察17β-雌二醇对豚鼠胆囊平滑肌肌条的急性效应及可能机制。结果 与假手术组相比,去卵巢组豚鼠血清雌二醇和CCK-8含量降低(P＜0.05),离体胆囊肌条对CCK-8以及乙酰胆碱的敏感性升高(P＜0.05)。随着皮下注射17β-雌二醇时间（1、3和7 d）的延长,二者含量升高(P＜0.05),离体胆囊肌条对CCK-8和乙酰胆碱的敏感性下降(P＜0.05)。10-9～10-7 mol/L的17β-雌二醇对正常豚鼠离体胆囊肌条收缩无明显影响(P＞0.05),10 6～10 5mol/L的17β-雌二醇可以抑制胆囊肌条收缩(P＜0.05),这种抑制效应可以被尼莫地平、阿托品、地伐西匹、ICI 182,780和Y-27632等阻断剂阻断。结论 17β-雌二醇与胆囊动力相关,可以通过基因组和非基因组机制影响胆囊平滑肌的收缩。     

Regulation of rat mesangial cell migration by platelet-derived growth factor, angiotensin II, and adrenomedullin

This study sought to determine whether platelet-derived growth factor (PDGF) and angiotensin II (AngII) stimulate migration of cultured rat glomerular mesangial cells. After finding that this was so, the effects of adrenomedullin (ADM) and cAMP-elevating agents on basal and stimulated mesangial cell migration were examined. Two isoforms of PDGF, AB and BB, stimulated migration in a concentration-dependent manner between 1 and 50 ng/ml, while the AA isoform lacked significant effect. AngII modestly but significantly stimulated migration in a concentration-dependent manner between 10(-7) and 10(-6) mol/L. Rat ADM significantly inhibited the PDGF BB- and AngII-stimulated migration in a concentration-dependent manner between 10(-8) and 10(-7) mol/L. Inhibition by rat ADM was accompanied by an increase in cellular cAMP. cAMP agonists or inducers such as 8-bromo cAMP, forskolin, and prostaglandin I2 also significantly reduced the stimulated migration. H 89, a protein kinase A (PKA) inhibitor, attenuated the inhibitory effect of ADM, and a calcitonin gene-related peptide (CGRP) receptor antagonist, human CGRP (8-37), abolished the inhibitory effects of rat ADM. These results suggest that PDGF AB and BB as well as AngII stimulate rat mesangial cell migration and that ADM can inhibit PDGF BB- and AngII-stimulated migration, at least in part through cAMP-dependent mechanisms likely to involve specific ADM receptors with which CGRP interacts. The adenylate cyclase/cAMP/PKA system may be involved in the migration-inhibitory effect of ADM in these cells.     

人体桡动脉与大隐静脉内皮细胞一氧化氮与超极化因子的差异及其意义

目的 通过直接检测人体大隐静脉与桡动脉内皮细胞释放的一氧化氮及平滑肌细胞膜电位信号,对比两者一氧化氮和内皮超极化因子(EDHF)作用的差异.方法 分别将长度为5 mm的桡动脉(n=8)、大隐静脉(n=23),以及压力扩张后的大隐静脉(PV,n=9,将肝素生理盐水注入大隐静脉管腔,使压力维持在100～ 600 mmHg,1 mmHg =0.133 kPa)血管条放置于37℃血管灌流室中,依次加入乙酰胆碱( 10-8～ 10-5 mmol/L)和缓激肽(10-10～l0-7mmol/L),以激发内皮细胞释放一氧化氮与EDHF.应用一氧化氮测定仪与膜电位记录仪,直接检测桥血管释放的一氧化氮与平滑肌细胞膜电位信号,研究其一氧化氮释放动力学与EDHF介导的超极化反应.结果 桡动脉内皮细胞一氧化氮基础释放浓度为(11.9±1.8) nmol/L,高于大隐静脉的(9.9±2.8) nmol/L(q=3.422,P=0.041).当缓激肽10-7 mol/L时,一氧化氮的刺激性释放比大隐静脉低,为(25.8±3.6)nmoL/L比(43.7±8.2)nmol/L,P=0.006.PV一氧化氮的基础释放为(3.4±1.4)nmoL/L,低于桡动脉和大隐静脉(P均＜0.05);刺激性释放亦小于桡动脉和大隐静脉(P均＜0.05).乙酰胆碱10-5 mol/L的条件下,桡动脉EDHF调节的平滑肌细胞超极化幅值高于大隐静脉,为(-9.7±1.9)mV比(-4.5±1.1)mV(P =0.002).结论 桡动脉内皮细胞一氧化氮基础释放与EDRF介导的内皮细胞功能明显优于大隐静脉.压力扩张会损害大隐静脉的一氧化氮介导内皮细胞功能,导致大隐静脉桥远期通畅率低下.     

Effects of calcitonin, amylin, and calcitonin gene-related peptide on osteoclast development

Amylin and calcitonin gene-related peptide (CGRP) are homologous 37 amino acid peptides that are found in the circulation. Both peptides belong to the calcitonin family. Similar to calcitonin, amylin and CGRP inhibit osteoclast activity, although they are much less potent than calcitonin. Calcitonin is known to act on the latter stages of osteoclast development, inhibiting the fusion of committed preosteoclasts to form mature multinucleated cells; however, whether or not calcitonin acts earlier in the formation of the precursor osteoclasts is controversial. The question of osteoclast development has never been examined with respect to amylin and CGRP. These issues are addressed in the present study. We studied the effects of calcitonin (salmon and rat), amylin (human and rat), and CGRP (human and rat) in mouse bone marrow cultures stimulated to generate osteoclasts using 1alpha,25-dihydroxyvitamin D3. Calcitonin dose-dependently decreased the numbers of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells as well as TRAP-positive mono-/binucleated cells at concentrations >10(-13) mol/L. Amylin and CGRP showed similar effects at concentrations >10(-9) mol/L. In addition, calcitonin substantially reduced the ratio of TRAP-positive multinucleated to mono-binucleated cells, indicating an effect on fusion of osteoclast precursors. The present data establish that this family of peptides not only acts on mature osteoclasts but also inhibits their development in bone marrow cultures. This activity is shared by amylin and CGRP. The much greater potency of calcitonin than amylin and CGRP is consistent with the action of these peptides being mediated by calcitonin receptors.     

Chronic alcoholism associated with diabetes impairs erectile function in rats

OBJECTIVE

To investigate the effects of chronic ethanol consumption and diabetes on nitric oxide (NO)‐mediated relaxation of cavernosal smooth muscle (CSM).

MATERIAL AND METHODS

Male Wistar rats were divided into four groups: control, isocaloric, diabetic and ethanol‐diabetic. The CSMs were mounted in organ chambers for measurement of isometric tension. Contraction of the strips was induced by electrical field stimulation (EFS, 1–32 Hz) and phenylephrine. We also evaluated the effect of ethanol consumption on the relaxation induced by acetylcholine (ACh; 0.01–1000 µmol/L), sodium nitroprusside (SNP, 0.01–1000 µmol/L) or EFS (1–32 Hz) in strips pre‐contracted with phenylephrine (10 µmol/L). Immunoexpression of endothelial NO synthase (eNOS) and inducible NOS (iNOS) was also accessed.

RESULTS

The endothelium‐dependent relaxation induced by ACh was decreased in CSM from ethanol‐diabetic rats when compared with the controls, with a mean (sem ) of 21 (4) vs 37 (2)%. Similarly, the potency and maximal responses induced by SNP were reduced in the ethanol‐diabetic [3.97 (0.38) and 85 (1)%, respectively] and diabetic groups [3.78 (0.56) and 81 (2)%, respectively] when compared with the controls [5.3 (0.22) and 90 (3)%, respectively] and isocaloric [5.3 (0.19) and 92 (1)%, respectively] groups. Noradrenergic nerve‐mediated contractions of CSM in response to EFS were increased in rats from ethanol‐diabetic and diabetic groups when compared with the control and isocaloric groups. Conversely, there were no differences in EFS‐induced relaxation among the groups. The immunostaining assays showed overexpression of eNOS and iNOS in the CSM from diabetic and ethanol‐diabetic rats when compared with the control and isocaloric rats.

CONCLUSION

There was an impairment of relaxation of CSM from ethanol‐diabetic and diabetic rats that involved a decrease in the NO‐cyclic guanosine monophosphate signalling pathway by endothelium‐dependent mechanisms accompanied by a change in the CSM contractile sensitivity.