Effects of adenosine and ATP on the membrane potential and synaptic transmission in neurons of the rat locus coeruleus

Effects of adenosine (Ado) and adenosine 5'-triphosphate (ATP) on the membrane potential and synaptic transmission in neurons of the rat locus coeruleus (LC) were examined, in vitro. Ado (30-300 microM) produced a hyperpolarizing response and inhibited spontaneous firing activity in neurons of the rat LC. Ado decreased input resistance of LC neurons. The Ado-induced hyperpolarization reversed polarity near the equilibrium potential of K+ (EK). Ado (100-300 microM) depressed both excitatory postsynaptic potential (EPSP) and inhibitory postsynaptic potential (IPSP). Ado (300 microM) did not alter the hyperpolarization induced by norepinephrine (30 microM). N6-Cyclopentyladenosine (CPA, 100 microM), an A1 receptor agonist, also produced a hyperpolarizing response and depressed both the EPSP and IPSP. Another A1 receptor agonist, adenosine amine congener (ADAC, 30 microM) also produced a hyperpolarizing response and consistently depressed the EPSP and IPSP. Application of ATP (100 microM) to LC neurons caused a depolarizing response associated with an increase in the firing rate of spontaneous action potential in LC neurons. The ATP-induced depolarization was accompanied by an increased input resistance and reversed polarity at--91 mV. ATP (100 microM) consistently depressed the IPSP, while it did not change the amplitude of the EPSP in a majority of neurons. alpha, beta-Methylene ATP (alpha, beta-meATP, 30 micro/M), a P2 receptor agonist, mimicked these effects of ATP. Adenosine 5'-(beta, gamma-imido) triphosphate (AMP-PNP, 100 microM), a non-metabolizable analogue of ATP, produced a depolarizing response in LC neurons, but it produced no obvious depression of the EPSP and IPSP. These results suggest that Ado and ATP cause inhibitory and excitatory modulation, respectively, of neuronal activity and synaptic transmission in the rat LC.     

Effects of milnacipran on the inhibitory postsynaptic potential in neurons of the rat locus coeruleus

Effects of milnacipran (MIL), a serotonin and noradrenaline reuptake inhibitor (SNRI), on synaptic transmission were examined in the rat locus coeruleus (LC). Bath-application of MIL produced a hyperpolarization associated with a decrease in input resistance of LC neurons. The MIL-induced hyperpolarization reversed polarity near the equilibrium potential of K+. The MIL-induced hyperpolarization was blocked by yohimbine (1 microM). Clonidine, but not serotonin (5-hydroxytryptamine; 5-HT), produced a hyperpolarizing potential in LC neurons. The MIL-induced hyperpolarization reversed polarity at -114 +/- 3 mV (n=4). MIL (0.1-10 microM) depressed the amplitude of the excitatory postsynaptic potential (EPSP), while it enhanced the amplitude and duration of the inhibitory postsynaptic potential (IPSP). These results suggest that MIL hyperpolarizes LC neurons and enhances the IPSP by increasing endogenous noradrenaline (NA) concentration at synapses in LC neurons.     

Effects of methylphenidate on the inhibitory postsynaptic potential in rat locus coeruleus neurons

Effects of methylphenidate (MPH), an agent used clinically for the treatment of children presenting the attention-deficit/hyperactivity disorder (AD/HD), on synaptic transmission in the rat locus coeruleus (LC) were examined by intracellular recording methods. Bath-application of MPH (30 nM-3 microM) increased the amplitude of the inhibitory postsynaptic potential (IPSP), while it did not change the amplitude of the excitatory postsynaptic potential (EPSP). MPH increased the time-to-peak and the half-decay time of the IPSP in LC neurons. MPH increased the amplitude of spontaneous IPSP: individual spontaneous IPSPs merged one into the other so as to produce regular, long-lasting waves of hyperpolarization. Clonidine (10 nM), a selective agonist for alpha 2-adrenoceptors, depressed the IPSP without affecting the EPSP in LC neurons. The results suggest that MPH enhances inhibitory synaptic transmission in the rat LC by depressing the norepinephrine (NE) re-uptake system.     

氯化钐对大鼠离体颈上神经节烟碱传递的影响

目的：研究氯化钐（Samarium Chloride,SmCl3)对大鼠离体颈上神经节烟碱传递过程的影响。方法：细胞内生物电记录技术。结果：SmCl3在10^-4--10^-1mmol/L的浓度时,可逆地抑制快兴奋性突触后电位（f-EPSP）,但对乙酰胆碱（ACh)和氨甲酰胆碱（Carb)膜除极化反应无显著的影响。对细胞膜电位、膜电阻亦无明显影响。SmCl3(10^-1mmol/L)能拮抗高钙（10mmol/L),对f-EPSP的易化4作用（P＜0．01）。结论：SmCl3对烟碱的抑制作用是通过突触前机制,可能与抑制钙的内流有关。     

Ionic basis of noradrenaline depolarization and hyperpolarization in toad dorsal root ganglia neurones

Summary The membrane conductance and reversal potential were estimated for neurones in toad dorsal root ganglia with intracellular recording technique during depolarization or hyperpolarization induced by noradrenaline (NA). The effects of blocking agents for potassium or calcium ion current on NA-induced membrane potential responses were observed as well. The NA depolarization was accompanied by a 32.6% decrease of membrane conductance. In a few neurones, the membrane conductance showed an initial increase and a following decrease. The NA hyperpolarization was associated with an increase of membrane conductance by 16.2%. The mean reversal potential of NA depolarization was -88.5 ± 0.9mV. The NA hyperpolarization responses were nullified at -89 to -92 mV of membrane potential. TEA superfusion enhanced NA depolarization amplitude by 73.7 ± 11.9% and depressed NA hyperpolarization amplitude by 40.5%. CsCl (intracelluar injection) increased phenylephrine depolarization by 34.5 %. MnCl₂ superfusion decreased the amplitudes of NA depolarization by 50.5 ± 9.9%, and of NA hyperpolarizalion by 89.5±4.9% respectively. The results suggest that depolarization or hyperpolarization induced by NA might be mediated by the alteration of K or Ca channels.     

人工体神经-内脏神经反射弧的神经电生理研究

目的从电生理学角度研究躯体运动神经能否再生替代内脏运动神经及再生神经的电生理性质,探讨新反射弧支配膀胱的可能机制。方法显微吻合大鼠左侧L4L6前根建立控制排尿的人工体神经-内脏神经反射弧,电刺激吻合口近端,记录盆神经节前、节后纤维的诱发电位和膀胱压力变化,使用特异性神经节受体阻断剂六烃季铵观察节后纤维诱发电位和膀胱压力的变化,刺激同侧坐骨神经观察的膀胱压力变化;以正常鼠为对照组。结果1．刺激吻合口近端,吻合口远端盆神经、盆神经节后神经纤维均可记录到诱发电位,并可引起膀胱收缩;2．刺激吻合神经同侧坐骨神经可记录到膀胱压的升高;3．六烃季胺可阻止盆神经节的讯号传导;4．人工反射弧再生的传出神经较对照具有更快的传导速度。结论躯体运动神经与内脏运动神经吻合后,体神经运动神经轴突可以再生替代内脏运动神经,并通过盆神经节支配膀胱逼尿肌。     

Effect of acetylcholine on membrane potential in toad dorsal root ganglion neurons and its underlying ionic basis

Intracellular recordings were made to investigate the responses of membrane potential to acetylcholine (ACh) on neurons in isolated toad dorsal root ganglion (DRG). In the 73 neurons examined, 67 were of type A, and the remaining 6 of type C cell. The resting membrane potential of these two types of cells was −67.5±1.3 mV (× ±SE). During the application of ACh (4 × 10^-4–6 × 10^-4 mol/L), the changes in membrane potential were as follows: 1) hyperpolarization, with amplitude of 9.1±3.0 mV (X ± SE; n = 23); 2) depolarization, with amplitude of 12.9 ±2.2 mV (X ±SE; n = 20); 3) biphasic response, i.e., hyperpolarization with amplitude of 8.0±2.4 mV (X±SE) followed by depolarization with amplitude of 10.9±2.1 mV (X±SE) (n=24); no effect (n=6). The hyperpolarization induced by ACh was blocked by superfusion with atropine (1.3 × 10^-5 mol/L; n = 23), while ACh depolarization was blocked by the mixture of d-tubocurarine (1.4 × 10^-5 mol/L) and hexamethonium (1.4 ×10^-5 mol/L) (n = 18). When ACh caused hyperpolarization, the membrane conductance wascin reased by 13.8% and the reversal potential was about -96 mV (n=3). TEA (20 mmol/L) superfusion enhanced ACh depolarization amplitude by 48.2 ±3.2 % (× ± SE;n = 6), and depressed ACh hyperpolarization amplitude by 79.4 ±4.3 % (× ± SE; n= 8).MnCl₂ (4 mmol/l) superfusion decreased the amplitudes of ACh depolarization and hyperpolarization by 54.2 ±7.2 % (X ±SE; n= 5) and by 69.2±6.4 % (X±SE; n = 14) respectively. These results suggest that the depolarization and hyperpolarization induced by ACh are mediated by nicotinic and muscarinic receptors on the soma of toad DRG neurons separately, and it seems that ACh hyperpolarization involves activation of calcium-activated potassium conductance.     

胆碱酯酶抑制剂对胆碱能突触传递的差异性作用

应用细胞内记录技术，观察了毒扁豆碱（Ｐｈｙｓ）和石杉碱甲（Ｈｕｐ－Ａ）对离休蟾蜍椎旁神经节胆碱能传递的影响。发现Ｐｈｙｓ或Ｈｕｐ－Ａ１μｍｏｌ／Ｌ升高节前神经刺激诱发的顺行动作电位（ＡＰ）发放率。Ｐｈｙｓ５０～１００μｍｏｌ／Ｌ同时抑制顺行ＡＰ及外源性ＡＣＨ电位（ｎ＝６），但Ｈｕｐ－Ａ　５０～３００μｍｏｌ／Ｌ不仅更加强烈、快速、持久地增大突触性及ＡＣｈ电位（ｎ＝２１），并在６２％的细胞使单次节前刺激诱发长时程去极化反应，结果表明，不同胆碱酯酶抑制剂对胆碱能传递有不同的药理和毒理学作用。     

Adenosine inhibits a GABAB receptor-mediated hyperpolarizing postsynaptic potential in neurons of rat septal nuclei

Intracellular recordings were made from neurons of rat dorsolateral septal nucleus (DLSN), in vitro. Adenosine and 2-chloroadenosine (1-500 microM) hyperpolarized DLSN neurons and blocked the excitatory postsynaptic potential (EPSP) and the late hyperpolarizing potential (LHP) in the presence of bicuculline. Adenosine did not depress the glutamate-induced potential. Bath-application of adenosine depressed the baclofen-induced potential in 60% of the neurons. Adenosine also inhibited the LHP in the remaining 40% of neurons, while it did not depress the baclofen-induced potential in these neurons. These results indicate that adenosine inhibits the EPSP pre-synaptically whereas it inhibits the LHP both pre- and postsynaptically in rat septal nuclei.     

大鼠脊髓前角与盆神经节之间植块联合培养的实验研究

目的:联合培养大鼠脊髓前角与盆神经节( major pelvic ganglia,MPG )植块并初步探讨两者相互作用的可能性?方法:建立大鼠脊髓前角与盆神经节之间植块的联合培养体系,观察神经元的生长变化?结果:神经组织在体外联合培养时均生长良好,脊髓前角植块生长出的运动神经元(spinal motor neurons,SMNs)发出突起与盆神经节的神经元形成明显形态上的接触?结论:联合培养的不同系统的神经元能彼此形成明显接触,初步提示运动神经元与盆神经节神经元之间可能形成功能性的相互作用?     

阿扑吗啡对离体蟾蜍交感神经节细胞的作用

应用离体蟾蜍椎旁神经节（ＰＶＧ）细胞内记录技术，初步观察了阿扑吗啡（ＡＰＯ）对交感神经节细胞的作用。ＡＰＯ１，３和１０ｍｍｏｌ／Ｌ灌流３０ｓ分别使６４％（ｎ＝２５），８３％（ｎ＝２４）和１００％（ｎ＝１１）的ＰＶＧ细胞产生浓度依赖性的缓慢去极化反应，在部分细胞可伴有输入膜电阻减少和去极化幅度随膜超极化而增大，但在全部受试细胞均未记录到确切的超极化反应。ＡＰＯ０．１～３ｍｍｏｌ／Ｌ灌流１５ｍｉｎ不仅使全部受试细胞产生去极化反应（ｎ＝２０），还使电刺激交感节前纤维诱发的顺行动作电位起始段上升速率减慢，超极化后电位增大、去极化后电位减小，并在０．３ｍｍｏｌ／Ｌ以上的浓度使顺行动作电位转为兴奋性突触后电位或完全取消突触反应，但对直接动作电位无明显影响。另外，ＡＰＯ还可逆地降低外源性Ａｃｈ电位的幅度（ｎ＝１０）。结果表明ＡＰＯ对离体蟾蜍交感神经节细胞有去极化作用并抑制其胆碱能突触传递。     

大鼠脊髓背角胶状质神经元的去极化反跳及调控机制

目的 研究大鼠脊髓背角胶状质(SG)神经元的去极化反跳及调控机制,以期对去极化反跳相关疾病的临床治疗提供参考.方法 选取3～5周龄SD大鼠,制作离体脊髓纵切片,应用全细胞膜片钳技术记录SG神经元的电生理学特点及接受超极化刺激后的反应,并观察超极化激活环核苷酸门控阳离子(HCN)通道阻断剂和T型钙(Cav3)通道阻断剂对去极化反跳的作用.结果 共记录了63个SG神经元的电活动,其中23个无去极化反跳,19个为去极化反跳无放电,21个为去极化反跳伴放电.无去极化反跳组SG神经元的动作电位阈值(-28.7±1.6 mY)明显高于去极化反跳伴放电组(-36.0±2.0 mV)(P＜0.05).HCN通道阻断剂氯化铯和ZD7288可显著延长去极化反跳伴放电的潜伏期,分别从45.9±11.6 ms增加到121.6±51.3ms(P＜0.05)和从36.2±10.3 ms增加到73.6±13.6ms(P＜0.05);ZD7288也能显著延长去极化反跳不伴放电的潜伏期,从71.9±35.1 ms增加到267.0±68.8 ms (P＜0.05),而T型钙通道阻断剂氯化镍和米贝地尔可显著降低去极化反跳伴放电的振幅,分别从19.9±46.3 mV降到9.5±4.5 mV(P＜0.05)和从26.1±9.4 mV降到15.5±5.0mV(P＜0.05),米贝地尔同样能显著降低去极化反跳不伴放电的振幅,从14.3±3.0 mV降低至7.9±2.0 mY(P＜0.05).结论 近2/3的SG神经元有去极化反跳,其潜伏期和振幅分别受HCN通道和T型钙通道调控.     

Additive potentiation by co application of propofol and sevoflurane on GABA induced chloride current in cultured neurons from rat dorsal root ganglia

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脑溢安对血红蛋白损伤神经元神经生长因子和白细胞介素1β表达的影响

目的：观察脑溢安对体外培养神经元血红蛋白损伤后的保护作用及神经生长因子与白细胞介素-1表达的影响。方法：以50umol/L^-1血红蛋白造成培养海马神经元损伤,运用活细胞计数、Northern杂交、酶联免疫检测神经元存活数量、神经生长因子与白细胞介素-1mRNA及其蛋白表达水平。结果：体外培养大鼠海马神经元在含2%牛血清的培养液中可存活5个月以上。加入50umol.L^-1血红蛋白培养24h神经元死亡率为38.5%。同时加入脑溢安药液能显著降低神经元死亡率。血红蛋白可引起培养神经元白细胞介素-1mRNA与蛋白表达水平显著升高及神经生长因子的短暂升高,脑溢安能降低白细胞介素-1表达,维持神经生长因子持续表达。结论：脑溢安对血红蛋白损伤神经元具有保护作用,其机理与调节神经元白细胞素-1表达,维持神经生长因子持续表达。结论：脑溢安对血红蛋白损伤神经元具有保护作用。其机理与调节神经元白细胞素-1、神经生长因子表达有关。     

脚趾截除后大鼠前扣带回锥体神经元突触传递持续性增强

截肢患者中50%-80%经历过幻肢疼痛.前扣带回(anterior cingulate cortex,ACC)是参与疼痛情绪反应形成的一个重要部位.为揭示前扣带回在幻肢疼痛中的作用,我们采用成年大鼠右后中趾截除慢性疼痛模型,麻醉固定后在体纪录大脑前扣带回锥体神经元之间突触传递特性的变化.所记录神经元经形态学确认为前扣带回锥体神经元.结果显示,在右后中趾截除后3-7 d,大鼠前扣带回锥体神经元兴奋性突触后电位持续性增强,同时,反映突触前机制参与突触可塑性的配对脉冲易化(paired-pulse facilitation,PPF)值增大.结果 表明,大鼠在体脚趾截除引起前扣带回锥体神经元兴奋性突触后电位持续性增强,这个增强的突触传递可能参与了幻肢疼痛的形成过程.     

Morphologic Relationship between the Pontine Micturition Center and the Sympathetic Center in the Spinal Cord of the Rat

To study whether the sympathetic nerves coordinate with the parasympathetic nerves during micturition in the rat. We used antegrade neural tracing with biotinylated dextran amine (BDA) injected into the pontine micturition center (PMC) to label the terminals in the L6-S1 cord，Preganglionic parasympathetic neurons (PPNs) in the L6-S1 segment were labelled by retrograde transport of Fluorogold (FG) from the major pelvic ganglion (MPG). We detected retrograde neurons in L6-S1 using retrograde transport of horseradish peroxidase (HRP) from the intermediolateral cell column (IML) of the L1-L2 segment where sympathetic preganglionic neurons (SPNs) are located, lmmunohistochemical methods showed that PPNs were identified to he choline acetyltransferase-immunoreactive (ChAT-IR). HRP-labelled neurons were not ChAT-IR and located dorsal to PPNs. BDA labelled terminals were located mainly in the bilateral IML of L6-S1, some of which had synaptic contact with the HRP-labelled neurons. In addition, there were some wheat germ agglutinin-horseradish peroxidase (WGA-HRP) labelled terminals in the ipsilateral IML of the L1-L2 segment after WGA-HRP was microinjected into SPN. We conclude that PMC may control the preganglionic neurons of sympathetic nerves through the interneurons located dorsal to PPNs.     

蛙皮素对离体交感神经节细胞的易化作用

目的:探讨蛙皮素(BOM)对离体交感神经节细胞的易化作用.方法:选用豚鼠离体交感神经节(主要是肠系膜下神经节,IMG),应用细胞内生物电记录技术观察压力注射和灌流BOM对神经节细胞膜电位的影响.结果:BOM使绝大部分IMG细胞膜电位缓慢除极,该除极反应与剂量相关,可为特异性受体阻断剂Tyr4,[D-phe12]BOM所阻断.结论:可能有BOM相应受体存在于豚鼠交感神经节细胞膜;BOM可使除极电位幅度增大并转化为动作电位(AP),并使电刺激突触前神经所致的快兴奋性突触后电位(f-EPSP)转化为AP,提示BOM对神经元兴奋性起易化作用.     

慢性粒细胞白血病骨髓巨噬细胞表达与细胞因子的相关性

目的 探讨慢性粒细胞白血病(CML)患者骨髓中巨噬细胞及白细胞介素(IL)-1β、IL-2、IL-4、IL-10、γ干扰素(INF-γ)的表达及相关性以明确其临床意义.方法 选择CML慢性期30例(CML-CP组)、加速期21例(CML-AP组)、急变期20例(CML-BP组)、经治疗后完全缓解的患者44例、未缓解7例以及30例缺铁性贫血患者(对照组),采用免疫组化染色法检测上述各组患者骨髓组织中CD68、CD163的表达变化;应用流式细胞仪分析相应患者骨髓IL-1β、IL-2、IL-4、IL-10、INF-γ表达;比较骨髓巨噬细胞在CML患者不同时期骨髓组织CD68、CD163的表达差异与IL-1β、IL-2、IL-4、IL-10、INF-γ表达变化的关系.结果 CML各组患者骨髓组织中CD68、CD163均有不同程度的表达,且随着病情的进展表达逐渐增高,即对照组＜CML-CP组＜CML-AP组＜CML-BP组,组间比较差异有统计学意义(P＜0.05);同时,CD163与CD68阳性细胞数密切相关(P＜0.05).与对照组比较,CML-CP组骨髓IL-2、INF-γ明显降低(P＜0.05),IL-1β、IL-4、IL-10表达明显升高(P＜0.05);随着病情进展,IL-2、INF-γ表达逐渐降低,即对照组＞CML-CP组＞CML-AP组＞CML-BP组,其组间差异有统计学意义(P＜0.05),且与CD163表达呈负相关;IL-1β、IL-4、IL-10表达逐渐升高,对照组＜CML-CP组＜CML-AP组＜CML-BP组,组间差异有统计学意义(P＜0.05),并与CD163表达呈正相关.患者经治疗完全缓解后,CD68及CD163仍高于对照组(P＜0.05);IL-2、INF-γ有所回升,但仍低于对照组(P＜0.05),IL-1β、IL-4、IL-10有所降低,同样也高于对照组(P＜0.05).未缓解组所测指标与CML-BP组相比,差异无统计学意义(P＞0.05).结论 巨噬细胞在CML患者骨髓中的异常浸润并逐渐从M1型向M2型巨噬细胞激化及IL-1β、IL-2、IL-4、IL-10、INF-γ的异常表达,改变了正常骨髓微环境,有利于白血病细胞的生存和增殖分化,可能与CML的发生、发展相关.