铃蟾肽介导的豚鼠肠系膜下神经节非胆碱能迟慢兴奋性突触后电位

应用细胞内记录技术,对铃蟾肽(bombesin,BOM)在豚鼠离体肠系膜下神经节(inferior mesenteric ganglion,IMG)非胆碱能兴奋性突触传递中的作用进行了研究。重复电刺激突触前结肠神经,有74．3％(52／70)IMG细胞可诱发迟慢兴奋性突触后电位(ls-EPSP)。在可引出ls-EPSP的细胞中,22％(4／18)细胞同时对BOM和SP敏感。用BOM持续灌流IMG,可明显抑制对BOM敏感细胞的ls-EPSP,对BOM不敏感细胞的ls-EPSP则无影响,且BOM受体与SP受体间无交叉脱敏。BOM受体阻断剂tyr^4[D-phe^12]bombesin能明显可逆性地抑制BOM敏感细胞的ls-EPSP和去极化,但对BOM不敏感细胞则无影响。研究结果提示,BOM可能是介导豚鼠IMG细胞ls-EPSP的一种递质。     

蛙皮素对豚鼠肠系膜下神经节细胞的生物电影响

应用离休细胞内记录技术,观察了蛙皮素（BOM）对豚鼠离体肠系膜下神经节（IMG）细胞膜电位和膜电阻的影响,结果表明,181个IMG细胞在压力注射BOM（10^-5mol/L,1-15pulse,3-15ms)时呈现缓慢去极化（84．0％）,先超极化后去极化（8．3％）,和无明显反应（7．7％）,在10个细胞上灌流BOM（10^-7-10^-6mol/L,60s),90％的细胞亦缓慢去极化,该去极化反应受低钙／高镁溶液的影响,但不为胆碱和肾上腺素受体阻断剂所阻断;膜电阻表现为减小（60．0％）,不变（35．0％）和增大（5．0％）,说明BOM可能存在于豚鼠IMG细胞上且发挥易化作用。     

豚鼠肠系膜下神经节细胞电活动与结肠运动的关系

在豚鼠肠系膜下神经节(IMG)及其支配的结肠段联合标本上,对IMG细胞内电位与肠段纵肌或环肌舒缩活动进行了同步记录。实验结果表明:(1)肠段预置张力为零时,约50％IMG细胞有自发的快兴奋性突触后电位(EPSP)活动,切断结肠神经或以筒箭毒(50μmol/L)灌流IMG后消失;(2)筒箭毒或低钙高镁溶液阻断神经节传递时,环肌节律性收缩幅度增大,节律变慢,但对纵肌节律性收缩无明显影响,(3)串刺激节前神经,在IMG细胞引起一串快EPSP或动作电位并常跟随迟慢的EPSP,同时,纵肌在0.1-0.2s潜伏期后出现迅速的、时程基本与动作电位串一致的舒张波,后者在筒箭毒灌流IMG后消失,而环肌运动可见舒张、舒张波延长或收缩波增大。结果提示:IMG不仅中继经典的胆碱能传出功能,还参与以胆碱能传递为中介的肠-肠反射,该反射活动的传出效应主要在于抑制环肌收缩。     

5—羟色胺介导的豚鼠肠系膜下神经节突触传递

本实验通过豚鼠离体肠系膜下神经节(IMG)的细胞内生物电记录方法观察到:(1)5-羟色胺(5-HT 1-100μmol/L)灌流可在部分 IMG 细胞引起与非胆碱能迟慢兴奋性突触后电位(Is-EPSP)相似的缓慢去极化;(2)持续灌流5-HT 可使对5-HT 敏感的 IMG 细胞的Is-EPSP 明显阻抑;(3)5-HT 去极化及5-HT 敏感细胞的 Is-EPSP 均可为5-HT 再摄取抑制剂氟苯氧丙胺(50μmol/L)所增大,而对5-HT 不敏感细胞的 Is-EPSP 则不受这种药物的影响,(4)5-HT 合成抑制剂对氯苯丙氨酸(PCPA)预处理可使 IMG 细胞的 Is-EPSP 的出现率和去极幅度均明显减低。上述结果表明:5-HT 可能参与介导豚鼠部分 IMG 细胞的Is-EPSP。     

河蟹眼柄神经内分泌细胞Glu受体研究

作者采用全细胞膜片钳技术测定了中华绒螯蟹 (Eriocheirsinensis)眼柄视神经节端髓X器官 (MTXO)神经内分泌细胞对 0 0 1— 10mmol/L谷氨基酸 (Glu)的反应 ,并结合药理学方法进行了Glu受体研究。结果表明 ,Glu激活A型和B型细胞离子型Cl-通道蛋白受体 ,诱导快速激活、快速失活的配体门控Cl-通道电流 (IGlu)。依据内外液的Cl-浓度比例引发去极化或超极化反应 ,继续施加Glu ,细胞快速出现脱敏反应 ;去除Glu后 ,细胞约需 2 0s恢复对Glu的敏感状态。IGlu幅值呈浓度依赖性 ,量 效关系曲线呈线形 ,激活阈值为 0 0 1mmol/L ,约 5mmol/L达到饱和。河蟹眼柄神经内分泌细胞IGlu明显受到Cl-通道阻断剂picrotoxin(0 5mmol/L)抑制 ;对离子型Glu受体激动剂Quisqualate、Kainate、NMDA、AMPA不敏感。Ibotenicacid(IA)可模拟Glu诱导快速激活、快速脱敏的Cl-电流 ,并与Glu产生交互脱敏作用。Glu和GABA对河蟹眼柄神经内分泌细胞无交叉脱敏和交叉激活作用 ,甘氨酸 (Gly)没有诱导细胞产生任何反应 ,提示中枢神经系统通过Glu和GABA两套系统实现对眼柄神经内分泌系统的精确调控。     

5—羟色胺抑制离体脊髓运动神经元的突触传递

应用新生大鼠脊髓切片运动神经元(MN)细胞内记录技术,发现5-羟色胺(5-HT)10～100μmol/L灌流可浓度依从地抑制背、腹根刺激在MN诱发的兴奋性突触后电位(EPSP)和抑制性突触后电位,但可增大外源性谷氨酸引起的MN去极化。5-HT对背根性EPSP的抑制无刺激频率依赖性,可为5-HT_(1A)受体激动剂8-OH-DPAT模拟,但不受士的宁、酮色林及MDL 72222的影响。结果表明5-HT可直接激活初级传入纤维末梢5-HT_(1A)受体而抑制MN的突触传递。     

GABA_A和GABA_B受体介导的蟾蜍背根神经节神经元胞体膜反应

实验在蟾蜍离体背根神经节(DRG)标本进行细胞内记录。浴槽滴加10~(-4)-10~(-3)mol/LGABA引起膜电位改变如下:(1)去极化(79/100);(2)双相反应;先为去极化,继后为超极化(10/100);(3)无反应(11/100)。以上去极化反应均可为荷包牡丹碱所阻断。GABA-去极化时膜电导增加,逆转电位值为-15——25mV。低Cl~-和高Cl~-任氏液分别使GABA-去极化反应增大和减小。10~(-4)mol/Lbaclofen不引起膜电位改变。在GABA-去极化期间,观察到大部分细胞的动作电位时程(ApD)缩短。ApD的此种变化可为baclofen所模拟,但不为荷包牡丹碱所阻断。结果提示:蟾蜍DRG神经元胞体膜有GABA_A和GABA_B受体共存,前者介导膜电位的改变,后者介导ApD的缩短。本文并联系到初级传入终末的突触前抑制的产生机制进行了讨论。     

低频刺激后海马CA1区场兴奋性突触后电位与群体锋电位的变化

在大鼠海马脑片上使用双电极在CA1区进行细胞外记录 ,观察低频刺激 (LFS)诱发同突触长时程抑制 (LTD)时场兴奋性突触后电位 (fEPSP)的斜率 (S EPSP)和群体锋电位 (PS)的幅值 (A PS)的变化。给予 90 0脉冲 1HzLFS后 ,S EPSP和A PS降低的幅度分别是 35 4± 5 3%和 6 8 0± 7 2 % ;而给予 4 5 0脉冲 1HzLFS后 ,S EPSP和A PS分别降低 14 3± 2 3%和 36 8± 6 7%。上述两组中A PS的变化率均显著大于S EPSP (P <0 0 1) ,而 90 0脉冲数组中两个指标的变化率均大于 4 5 0脉冲数组 (P <0 0 5 )。高Mg2 + (4mmol/L)使突触的传递活动减弱 ,但不影响LTD的诱发 ,在高Mg2 + 介质中 ,LFS引起的A PS变化率仍显著大于S EPSP (P <0 0 1)。结果表明 ,由LFS诱发同突触LTD的水平不仅与LFS的脉冲数有关 ,还与评价指标的选择有关     

家兔第四脑室注射P物质对肺动脉压和颈动脉压的影响

本工作将P物质(SP)注入麻醉家兔第四脑室,观察其对肺动脉压和颈动脉压的影响。结果观察到:(1)脑室注射SP后,肺动脉压升高或降低,颈动脉压上升,心率减慢。(2)切断肺动脉压下降组家兔的两侧颈部迷走神经,再ivt.SP,则引起肺动脉压的升高,降心率反应则明显减弱。(3)预先用酚妥拉明或α_1受体阻断剂哌唑嗪均可阻断SP引起的肺动脉和颈动脉升压反应。(4)α_2受体阻断剂育亨宾或纳洛酮均可增强这二个升压反应。(5)心得安对这二个升压反应无明显影响。(6)SP的心血管效应可被SP受体拮抗剂[D-pro~2.D-Trp~(7.9)]-SP阻断。 实验结果表明:脑中SP升高可通过SP受体引起肺动脉压和颈动脉压上升,心率减慢;在SP引起的加压反应的中枢环节中有肾上腺素能α_1受体活动参与;中枢肾上腺素能α_2受体系统和内啡肽系统对传递SP中枢加压作用的路径有抑制性的调制作用。看来,SP与儿茶酚胺及阿片样物质一起参与脑干对血压的调节。     

血管钠肽对离体人乳内动脉的舒张作用

为了研究血管钠肽(VNP)对人乳内动脉(human intramammary artery,HIMA)的舒张作用及其机制,采用离体血管灌流的方法,观察VNP对内皮完整和去内皮HIMA的舒张作用,以及HS—142—1、TEA、8—Br—cGMP和镁蓝(MB)对这一过程的影响。实验中观察到,VNP(0．0001—1μmol/L)可引起剂量依赖性的舒张效应,且无内皮依赖性;8—Br—cGMP(0．1—1000μmol/L)也可引起剂量依赖性的血管舒张效应。钠尿肽鸟苷酸环化酶(guanylate cyclase,GC)受体的特异性阻断剂HS—142—1(20μmol/L)使VNP舒张HIMA的作用几乎完全消失。MB是GC的抑制剂,10μmol/L的MB不但使VNP舒张HIMA的作用完全消失,而且可增强HIMA对去甲肾上腺素(NE)产生的收缩反应。钙激活钾通道(KCa)的阻断剂TEA(1mmol/L)可减弱(但是不完全阻断)VNP的舒血管作用。上述结果表明,VNP对HIMA具有不依赖内皮的舒张作用;此作用是通过作用于平滑肌细胞的钠尿肽GC受体,引起细胞内的cGMP水平升高实现的,并且与Kca有关。     

Slow excitatory transmission in rat spinal dorsal horn and the effects of capsaicin

High intensity repetitive stimulation of a dorsal root elicited slow depolarization in more than half of the dorsal horn neurons examined in the rat spinal cord slice preparation. There was a significantly smaller group of neurons showing slow hyperpolarization as well. Slow depolarization was not observed when synaptic activity was blocked by perfusing the slice with a TTX- or a low-Ca2+ high-Mg2+ solution. This result is consistent with a presynaptic origin of the slow response. Capsaicin treatment of neonatal rats significantly reduced the incidence of slow depolarization, suggesting that the slow depolarization was generated by small diameter afferent fibres, probably unmyelinated afferents. DR-evoked slow depolarization and SP-induced depolarization were similar in several important aspects: a) Both responses caused depolarization and increased the excitability of dorsal horn neurons; b) They were frequently associated with similar membrane conductance changes; c) The size of both responses varied in parallel when the membrane potential was shifted over a wide range; d) Both responses were markedly reduced or abolished by an analogue of SP having antagonist properties, and by polyclonal and monoclonal antibodies to SP; e) The depression of the DR-elicited slow depolarization during and after the SP-induced depolarization suggested that SP and the natural transmitter for the DR-elicited slow depolarization were bound to the same receptors. The results suggest that SP or, SP-like peptide, is an agonist that mimics in some aspects the action on the natural transmitter for the slow depolarizing potential.     

交感神经节细胞对P物质和5-羟色胺的反应

本工作旨在观察Ｐ物质（ＳＰ）受体与５－羟色胺（５－ＨＴ）受体是否分别还是同时存在于豚鼠腹腔神经节（ＣＧ）与肠系膜下神经节（ＩＭＧ）不同细胞,以及这两种递质之间是否存在相互作用。在１３３个ＣＧ细胞中,６６个（４９．６％）对ＳＰ及５－ＨＴ同时敏感,４０个（３０．１％）仅对其中一种递质敏感,此外２７个（２０．３％）对两都不敏感。     

Presynaptic nicotinic acetylcholine receptors in the myenteric plexus of guinea pig intestine

Presynaptic nicotinic acetylcholine receptors (nAChRs) were studied in myenteric plexus preparations from guinea pig ileum using intracellular electrophysiological methods. Microapplication of nicotine (1 mM) caused a biphasic depolarization in all AH neurons (n = 30) and in 36 of 49 S neurons. Cytisine (1 mM) caused fast depolarizations in S neurons and no response in AH neurons. Mecamylamine (10 microM) blocked all responses caused by nicotine and cytisine. TTX (0.3 microM) blocked slow excitatory synaptic potentials in S and AH neurons but had no effect on fast depolarizations caused by nicotine. Nicotine-induced slow depolarizations were reduced by TTX in two of twelve AH neurons (79% inhibition) and four of nine S neurons (90+/-12% inhibition). Slow nicotine-induced depolarizations in the remaining neurons were TTX resistant. TTX-resistant slow depolarizations were inhibited after neurokinin receptor 3 desensitization caused by senktide (0.1 microM); senktide desensitization inhibited the slow nicotine-induced depolarization by 81+/-5% and 63+/-15% in AH and S neurons, respectively. A low-calcium and high-magnesium solution blocked nicotine-induced slow depolarizations in AH neurons. In conclusion, presynaptic nAChRs mediate the release of substance P and/or neurokinin A to cause slow depolarizations of myenteric neurons.     

Desensitization of the inhibition of the M-current in sympathetic neurons: effects of ATP analogs, polyanions, and multiple agonist applications.

Desensitization occurs when the response to a neurotransmitter receptor agonist wanes in the continued presence of agonist. In amphibian sympathetic neurons, both muscarinic and peptidergic receptor agonists inhibit a K+ current, the M-current (IM), and this inhibition desensitizes. We have studied the desensitization to substance P (SP) by whole-cell recordings from dissociated sympathetic neurons from bullfrogs. When ATP in the recording pipette was replaced with AMP-PNP, SP still inhibited IM, but no desensitization was observed, indicating that ATP hydrolysis is required for desensitization. Desensitization inhibitors of beta-adrenergic receptors did not block desensitization to SP. When a low dose of muscarine sufficient to inhibit IM, but not to elicit desensitization, was applied simultaneously with a desensitizing dose of SP, IM remained depressed and did not desensitize. Thus, there may be separate systems controlling desensitization for different agonists, or the enzyme(s) involved is "compartmentalized."     

兔肠系膜下神经节细胞的两种非胆碱能性慢突触后电位

以常规细胞内记录技术对兔肠系膜下神经节细胞的跨膜电位进行了观察。对节前神经的短串脉冲刺激,可诱发出一串快兴奋性突触后电位(f-EPSP)或顺向动作电位;在此之后,大多数细胞还出现一个持续约2min 的缓慢去极化电位。该电位具有抗箭毒和阿托品性质,受低钙高镁溶液的可逆性阻抑,因而可称为非胆碱能性兴奋性突触后电位,或者也可归入迟慢兴奋性突触后电位(ls-EPSP)。多数细胞的 ls-EPSP 伴有膜电阻增大,电位的幅度随细胞静息电位的超极化而变小;提示在这些细胞上,钾电导的失活很可能参与了电位的发生。以P物质溶液灌流神经节未见该电位有显著改变。另外,在箭毒化加阿托品化的神经节中,还发现少数细胞对节前神经的串刺激发生一个持续约一分钟的超极化电位。它也具有抗胆碱能受体阻断剂的性质,受低钙高镁溶液可逆性阻抑,为此我们命之为“极慢抑制性突触后电位”(vs-IPSP),以区别于“慢抑制性突触后电位”(s-IPSP),后者是通常用以表示一种胆碱能性的慢电位。本文所述的这两种非胆碱能性的突触电位有关递质,尚待探索。     

P物质对大鼠分离的DRG细胞GABA激活电流的抑制作用

本文就用全细胞膜片箝技术,在新鲜分离的大鼠ＤＲＧ细胞上证明,在部分细胞Ｐ物质（１０＾－７－１０＾－５ｍｏｌ／Ｌ）可引起浓度依赖性的内向流（４／２６）;在多数细胞虽未检测到ＳＰ引起的膜电流,但却能对ＧＡＢＡＡ受体激活介导的膜内向流产生抑制效应（１８／２２）,并有加速去敏感的作用。本文就有关ＳＰ以ＧＡＢＡ激活电流抑制效应的可能意义进行了讨论。     

Substance P release evoked by capsaicin or potassium from rat cultured dorsal root ganglion neurons is conversely modulated with bradykinin

To clarify the molecular mechanism of substance P (SP) release from dorsal root ganglion (DRG) neurons, we investigated the involvement of several intracellular effectors in the regulation of SP release evoked by capsaicin, potassium or/and bradykinin. Bradykinin-evoked SP release from cultured adult rat DRG neurons was attenuated by either the mitogen-activated protein kinase kinase (MEK) inhibitor (U0126) or cycloheximide. As the long-term exposure of DRG neurons to bradykinin (3 h) resulted in extracellular signal-regulated kinase (ERK) phosphorylation at an early stage and thereafter induced cyclooxygenase-2 (COX-2) protein expression, which both contribute to the SP release triggered by bradykinin B2 receptor. The long-term exposure of DRG neurons to bradykinin enhanced the SP release by capsaicin, but attenuated that by potassium. Interestingly, the inositol 1,4,5-triphosphate (IP3)-induced calcium release blocker [2-aminoethyl diphenylborinate (2-APB)] not only inhibited the potassium-evoked SP release, but also completely abolished the enhancement of capsaicin-induced SP release by bradykinin from cultured DRG neurons. Together, these findings suggest that the molecular mechanisms of SP release by bradykinin involve the activation of MEK, and also require the de novo protein synthesis of COX-2 in DRG neurons. The IP3-dependent calcium release could be involved in the processes of the regulation by bradykinin of capsaicin-triggered SP release.     

Cardioprotection by ischemic postconditioning is lost in isolated perfused heart from diabetic rats: Involvement of transient receptor potential vanilloid 1, calcitonin gene-related peptide and substance P

We previously found that the expression of transient receptor potential vanilloid 1 (TRPV1) and contents of calcitonin gene-related peptide (CGRP) and substance P (SP), two main neuropeptides released from TRPV1, were decreased in diabetic hearts. This study aimed to test whether decreased TRPV1, CGRP and SP levels were responsible for the loss of cardioprotection by ischemic postconditioning (IPostC) in isolated perfused heart from streptozotocin-induced diabetic rats. IPostC effectively protected non-diabetic hearts against ischemia/reperfusion injury by improving cardiac function and lowering creatine kinase (CK) and cardiac troponin I (cTnI) release, which could be abolished by inhibiting TRPV1, CGRP receptor or SP receptor. However, IPostC had no effect on cardiac function and the release of CK and cTnI in diabetic hearts regardless of whether TRPV1, CGRP receptor or SP receptor were inhibited. CGRP or SP-induced postconditioning significantly prevented both non-diabetic and diabetic hearts from ischemia/reperfusion injury by improving cardiac function and lowering CK and cTnI release. Additionally, IPostC markedly increased CGRP and SP release in non-diabetic hearts, which could be reversed with TRPV1 inhibition, but not CGRP receptor or SP receptor inhibition. However, IPostC failed to affect CGRP and SP release in diabetic hearts in the presence or absence of TRPV1, CGRP receptor or SP receptor inhibition. These results indicate that the loss of cardioprotection by IPostC during diabetes is partly associated with a failure to increase CGRP and SP release, likely due to decreased TRPV1 expression and CGRP and SP contents in diabetic hearts.