在不均匀牵张大鼠膈肌标本上新霉素对接头作用的进一步研究

在不均匀牵张的大鼠离体膈肌标本上,同时记录同一终板的小终板电位(MEPP),终板电位(EPP)和微电泳外源性乙酰胆碱(ACh)诱发的乙酰胆碱电位(AChP)。以AChP幅度为突触后膜兴奋性大小指标,对MEPP和EPP进行校正;以量子含量M和二项统计参数即刻可释放量子贮存n及平均量子释放机率P为突触前递质释放指标,定量观察了新霉素(neomycin)对突触前和突触后的作用。结果表明新霉素(30μmol/L,100μmol/L)作用20min,对AChP,MEPP无影响,M_l,M_x,n和P明显下降,串EPPs(50Hz)出现脱落现象,表明此浓度下的新霉素对接头的作用主要是抑制突触前诱发的递质释放,对自发释放的量子大小并无影响。其机理可能是新霉素拮抗Ca~(2+)内流,减小n和P,从而降低诱发的递质释放。     

Fura—2荧光测定眼镜蛇心脏毒引起细胞胞浆Ca~(2+)浓度的变化

用Fura—2荧光技术直接测定眼镜蛇心脏毒(CTX)对大鼠泪腺细胞胞浆Ca~(2+)浓度的影响。在无Ca~(2+)的介液下,CTX可使胞浆Ca~(2+)浓度升高;随之加入1mmol/L CaCl_2可使胞浆Ca~(2+)浓度进一步升高。表明CTX能引起胞内Ca~(2+)释放和胞外Ca~(2+)内流。8mmol/L CaCl_2能阻断CTX升高胞浆Ca~(2+)浓度的作用。     

T淋巴细胞膜上离子通道与免疫性疾病的关系

T淋巴细胞上存在多种类型的离子通道,主要有电压门控性K~+通道(KV1.3),Ca~(2+)激活K~+通道(Kca3.1)、Ca~(2+)释放激活的Ca~(2+)+通道(CRAC)、瞬时受体电位通道(TRPM7)及细胞容积调节的Cl~-通道(Clswell)5种类型。这些离子通道共同参与T淋巴细胞的激活、分化等免疫调节过程,参与T淋巴细胞凋亡及体积调节等过程,与很多自身免疫性疾病息息相关。就T淋巴细胞上离子通道的分类、离子通道与淋巴细胞功能的关系、离子通道与免疫性疾病及在临床上的应用等方面进行综述。     

蛇床子素对家兔主动脉条的钙拮抗作用

蛇床子素30μmol·L~(-1)及100μmol·L~(-1)使NE、CaCl_2和高K~+除极化所致的家兔主动脉条收缩量—效曲线右移,最大反应降低.表明蛇床子素有松弛血管平滑肌作用,并与Ca~(2+)呈非竞争性拮抗作用.和阻滞α受体或激动β受体无关;选择性作用于电位依赖性Ca~(2+)通道,抑制细胞外Ca~(2+)内流;在100μmol·L~(-1)时,明显减弱NE诱导的依赖细胞内Ca~(2+)收缩。证明蛇床子素松弛血管平滑肌作用可能与其Ca~(2+)拮抗作用有关。     

不同条件下吡喹酮对日本血吸虫雄虫摄入钙的影响

吡喹酮仅在较短时间内增加体外培养的日本血吸虫♂虫对~(45)Ca~(2+)的摄入量。在含30 mMMg~(2+)的HBS中,吡喹酮对♂虫摄入~(45)Ca~(2+)的量无明显影响。在4℃的HBS中,吡喹酮可使♂虫对~(45)Ca~(2+)的摄入量至少在1小时内持续增加。此外,在无Ca~(2+)的HBS中,血吸虫♂虫已摄入的~(45)Ca~(2+)的释放量较在HBS中的显著为多。     

在不均匀牵拉的离体大鼠膈肌标本上梭曼的突触前和突触后作用

在离体大鼠膈肌上制备了一种不均匀牵拉肌肉以制动,便于用微电极记录终板电位(EPP)的标本(INSMP),避免了常规制动方法带来的制动药物、台氏液Ca~(2+),Mg~(2+)浓度改变或钳压肌肉导致膜电位(MP)下降的种种干扰。INSMP的MP和小终板电位(MEPP)正常,EPP幅度高达30mV以上,是研究药物对接头作用的较好标本。在INSMP上,梭曼(5.5μM)使MEPP频率加快,串刺激(50Hz)诱发的平均串EPPs幅度及其平均ACh 量子含量减少80％和77％。小剂量三碘季铵酚和六烃季铵可部分对抗梭曼的作用。梭曼引起强直收缩抑制主要原因为终板区蓄积的ACh作用于突触前N受体、负反馈地抑制ACh 量子释放;次要原因为作用突触后N受体、使其对ACh敏感性降低。     

人精子膜的离子通道和男性抗生育中草药的避孕机制(英文)

利用离子通道-脂双层重组技术,即将分离的人精子通道蛋白重组于人工脂双层膜,利用膜片箝技术记录、分析通道性质的方法,已鉴定了人精子膜数种通透Ca~(2+)、K~+、Na~+和Cl~-通道的生物物理和药理学性质,提供了成熟精子离子通道的基本电生理资料。利用小鼠生精细胞和膜片箝技术分析棉酚及由避孕中药雷公藤分离的多种单体对Ca~(2+)通道的作用,发现它们均在抑制精子顶体反应的浓度水平上抑制T型Ca~(2+)通道,表明对Ca~(2+)通道的抑制与它们的避孕效应密切相关;观察药物对小鼠生精细胞Ca~(2+)通道的作用,是筛选抗生育化合物特别是局部急用避孕药的一个有效方法。     

1-烯丙基氯-3对神经纤维轴浆运输作用的机理研究

本实验结果是神经纤维MT、NF变性和数量减少,线粒体肿胀并呈变性空泡;轴浆内并为轴浆运输成分的ChE活性降低或消失;轴浆Ca~(2+)增加,Mg~(2+)无改变;cAMP增加,cGMP及游离CaM降低;神经微丝肌球蛋白ATPase活性降低,Ca~(2+)-ATPase活性无改变。表明1-烯丙基氯-3促进或加强神经膜Ca~(2+)通道磷酸化作用,或CGMP减少负反馈PIP_2信息系统增强,或内质网及线粒体扩张     

尼莫地平对离体豚鼠左心房的抑制作用及其机理探讨

尼莫地平(50μmol/L)明显减低豚鼠左心室收缩幅度,其强度与戊脉安(25μmol/L)相当;明显抑制正阶梯现象,但不象戊脉安那样使其翻转成负阶梯现象;明显抑制静息后增强效应;逆转或预防哇巴因诱发的心律失常。以上结果提示尼莫地平既能减少胞外Ca~(2+)通过Ca~(2+)通道内流,又能减少胞内贮Ca~(2+)释放。     

Ca~(2+)激动剂的药理研究近况

<正> Ca~(2+)及调节Ca~(2+)的功能性蛋白——Ca~(2+)通道的药理学研究正如60年代对环核苷酸的研究一样,已进入了一个非常活跃的时期。许多临床疾病如高血压、心律失常、心绞痛、动脉硬化以及糖尿病、胃十二指肠溃疡等均与Ca~2的活动有关.应用Ca~(2+)通道拮抗剂作为研究工具并用于临床治疗已成为生物医学多个领域的重要内容。近几年来有关的研究有两个比较显著的特点:首先是应用二氢吡啶类     

Cadmium: effects on transmitter release at the frog neuromuscular junction

Cd2+ competitively antagonizes Ca2+ in the evoked release of acetylcholine from nerve terminals in the frog sciatic-sartorius neuromuscular junction. The dissociation constant between Cd2+ and its receptor sites was calculated to be 1.7 microM. In agreement with previous work we found that brief exposures to 100-500 microM Cd2+ had little or no effect on spontaneous transmitter release. However, in contrast to other reports, prolonged exposures to 100-100 microM Cd2+ or tetanic nerve stimulation during brief exposures to high concentrations of Cd2+ produced substantial increases in spontaneous transmitter release. We conclude that Cd2+, when present at low concentrations, is a specific Ca2+ channel blocker. At higher concentrations, Cd2+ is either relatively impermeable to the presynaptic nerve membrane or, if it does enter the nerve terminal, it is less effective than such metals as Pb2+, La3+ or Hg2+ in increasing the rate of spontaneous transmitter release.     

Interactions of lead and cadmium on acetylcholine release at the frog neuromuscular junction

The interactive effects of Cd2+ and Pb2+ on evoked and spontaneous transmitter release were studied in the sciatic nerve-sartorius muscle preparation of the frog (Rana pipiens). Either Pb2+ or Cd2+ competitively inhibited the actions of Ca2+ in bringing about evoked release, as measured by the endplate potential (EPP) amplitude. Combinations of Pb2+ and Cd2+ were additive in their effects on the EPP. The rate of spontaneous transmitter release was measured as the miniature endplate potential (MEPP) frequency. In contrast to their effects on the EPP, exposure of preparations to combinations of Pb2+ and Cd2+ actually increased the MEPP frequency less than exposures of the preparations to Pb2+ alone. The degree of reduction in MEPP frequency produced by Cd2+ depended upon the relative ratio of Pb2+ and Cd2+ ions. These results suggest that Pb2+ and Cd2+ ions competed for a common presynaptic receptor site during evoked release. They also suggest that Pb2+ ions may enter the nerve terminal, possibly through the Ca2+ channel, and that this entry is inhibited by Cd2+.     

Endothelin receptor blockade attenuates air embolization-induced pulmonary hypertension in sheep

CuCl2, ZnCl2 and NiCl2, but not CdCl2 or CoCl2, induced transmitter release from superfused rat hippocampal and striatal synaptosomes preloaded with, respectively, [3H]noradrenaline and [3H]dopamine. Cu2+ was the most potent and effective, acting in a concentration- (0.1-300 microM) and time-dependent (peak effect occurring at 2-3 min) manner. The amount of Cu2+-induced release over a 5 min period is similar to that induced by depolarization with high KCl or the K+ channel blocker 4-aminopyridine. However, the time course of the Cu2+-induced release is slower and the effect of Cu2+ is not reversed by washout. Cu2+-induced catecholamine release requires extracellular calcium (Ca2+) and is inhibited by the Ca2+ channel blocker Cd2+, and in the case of noradrenaline, by the voltage-gated Na+ channel blocker tetrodotoxin. The ability of Cu2+ to induce massive Ca2+-dependent transmitter release from brain catecholaminergic nerve terminals may contribute to the neuropathological processes associated with Cu2+ toxicity in Wilson's disease.     

Inhibition of evoked acetylcholine release: two different mechanisms in the Torpedo electric organ

The action of agents inhibiting evoked transmitter release was investigated by analysing the Ca2+ secretion relationship, electrophysiologically and biochemically, and by measuring the stimulation-induced 45Ca accumulation in the tissue. Transmitter release saturated at external Ca2+ concentrations higher than 4 mM, the releasing mechanism probably being the limiting step. Antagonists of Ca2+ entry (Mg2+, Cd2+, diltiazem) decreased the sensitivity of acetylcholine release to Ca2+, acting as competitive inhibitors, and reduced the stimulation-induced 45Ca accumulation. Quinacrine produced similar effects, indicating that it interacts primarily with Ca2+ entry. In contrast, oxotremorine and adenosine depressed transmitter release proportionally at all Ca2+ concentrations, acting as non-competitive inhibitors, and did not modify the stimulation-induced 45Ca accumulation. Their inhibitory effects were additive and reflected a decrease in the quantal content of the responses evoked. It is concluded that both drugs inhibit cholinergic transmission in the electric organ without altering Ca2+ entry into the nerve endings.     

Ryanodine facilitates calcium-dependent release of transmitter at mouse neuromuscular junctions.

1. Quantal release of transmitter was measured intracellularly at mouse neuromuscular junctions in the presence and absence of ryanodine (Rnd). 2. Rnd at concentrations up to 1 microM did not significantly alter the frequency of miniature endplate potentials (m.e.p.ps) in the presence or absence of Ca2+, suggesting that Rnd is unlikely to alter the internal concentration of Ca2+ ([Ca2+]i) at rest. 3. In a high-K+ (10 mM) bathing solution, Rnd further potentiated the facilitatory effect of Ca2+ on the frequency (F, s-1) of m.e.p.ps. Rnd shifted the relationship between 1n(F) and 1n[Ca2+]o to lower concentrations. 4. In a high-Mg2+ bathing solution, Rnd did not affect the frequency of m.e.p.ps at any value of [Ca2+]o. However, Rnd slightly but significantly increased the quantal content (m) of e.p.ps. It shifted the relationship between 1n(m) and 1n[Ca2+]o to lower concentrations. These results suggest that Rnd potentiates the quantal release of transmitter after depolarization of the membrane or nerve impulse, in keeping with the cooperativity of Ca2+ at the active site. 5. A series of two closely spaced nerve impulses produced a facilitation of transmitter release, as judged by the quantal content (m2) of the second response in relation to that of the first one (m1), m2/m1. Rnd did not change the ratio m2/m1. Thus Rnd is unlikely to affect the rapid phase of the sequestration of Ca2+ inside the nerve terminal. 6. High levels of K+ (5 mM) and caffeine (2 mM) potentiated both modes of transmitter release, in a manner dependent on [Ca2+]o. Caffeine did not potentiate facilitation of transmitter release.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of modulating Ca2+ entry and activating prejunctional receptors on facilitation of excitatory junction potentials in the guinea-pig vas deferens in vitro

This study investigated the effects of changing the extracellular Ca2+ concentration on facilitation of excitatory junction potential (EJP) amplitude during trains of 20 stimuli at 1 Hz at sympathetic neuroeffector junctions in the guinea-pig vas deferens. These effects were compared with those of Ca2+ channel blockers and agents which act at prejunctional receptors to increase or decrease neurotransmitter release. In these experiments, alpha-adrenoceptor-mediated autoinhibition of neurotransmitter release was blocked by the alpha-adrenoceptor antagonist, phentolamine (1 microM). Varying the extracellular Ca2+ concentration (0.75-6 mM) changed the amplitude of EJPs without affecting the maximum level of facilitation during the trains of stimuli. Reductions in Ca2+ concentration (from 2 mM) were associated with a slowing in the rate of development of facilitation. The Ca2+ channel antagonists, Cd2+ (2 microM and 5 microM) and omega-conotoxin GVIA (10 nM), and agents which act at prejunctional receptors to reduce neurotransmitter release, adenosine (100 microM and 1,000 microM) and prostaglandin E2 (PGE2; 0.1 nM and 1 nM), produced similar effects to those of lowering the extracellular Ca2+ concentration. Raising the extracellular Ca2+ concentration (from 2 mM) increased the rate of development of facilitation. Angiotensin II (AII; 0.5 microM) produced similar effects to raising extracellular Ca2+. However, isoprenaline (1 microM), while increasing EJP amplitude, reduced the maximum level of facilitation and was without effect on the rate of development of facilitation. In the guinea-pig vas deferens EJPs are produced by neurally released ATP. Thus, the findings support the idea that adenosine, PGE2 and AII change ATP release by modifying Ca2+ entry into the nerve terminal. However, the effects of isoprenaline may not solely be accounted for by modifications in Ca2+ entry.     

Stannous chloride-induced increase in calcium entry into motor nerve terminals of the frog

The effects of SnCl2 on potentials recorded extracellularly from motor nerve terminals of the bullfrog were studied to elucidate the mechanism of the SnCl2-induced facilitation of evoked transmitter release. Under conditions in which the muscle preparations were pretreated with d-tubocurarine and tetraethylammonium in a K+-free medium, SnCl2 (50 microM) augmented the prolonged positive deflection ascribed to the inward Ca2+ current, an effect which was reduced by addition of Cd2+. The results suggest that SnCl2 could increase Ca2+ entry into the nerve terminals.     

Antagonism of calcium currents and neurotransmitter release by barium ions at frog motor nerve endings

1. The effects of Ba(2+) (0.1 - 2 mM) on the component of the perineural voltage change associated with nerve terminal calcium currents (prejunctional Ca(2+) currents) were compared with the effects of this ion to antagonize calcium-dependent acetylcholine (ACh) release. These experiments were made on isolated neuromuscular junctions of the frog. 2. In the presence of sufficient concentrations of K(+) channel blockers to eliminate measurable prejunctional K(+) currents, low concentrations of Ba(2+) selectively antagonized prejunctional Ca(2+) currents in normal Ca(2+) solutions. Higher concentrations of Ba(2+) also substantially reduced the Na(+) component of the perineural waveform. 3. Ba(2+) inhibited the prolonged prejunctional Ca(2+) currents that developed in the presence of higher concentrations of K(+) channel blockers. 4. Simultaneous measurements of the prejunctional Ca(2+) currents and the electrophysiological correlates of ACh release (i.e. end-plate potentials, EPPs) were made under conditions of modest K(+) channel blockade. Under these conditions, Ba(2+) generally produced simultaneous decreases in both Ca(2+) currents and EPP amplitudes. In some instances, a prolongation of prejunctional Ca(2+) currents and a transient increase in EPP amplitudes preceded the decreases in both electrophysiological events. 5. These results suggest that Ba(2+) ions can antagonize the entry of calcium into motor nerve endings and this effect is likely to be responsible for the inhibitory effects of Ba(2+) on evoked ACh release.     

Effects of charybdotoxin, a blocker of Ca2+-activated K+ channels, on motor nerve terminals.

1. The contribution of Ca2+-activated K+ currents (IK,Ca) to the control of electrical excitability of motor nerve terminals and the control of acetylcholine release was assessed by studying the effects of the specific K(Ca) channel blocking toxins charybdotoxin and apamin. Electrical activity of the terminal regions of motor nerves was assessed by extracellular recording from an electrode placed in the perineural sheaths of nerves in the mouse triangularis sterni and frog cutaneous pectoris preparations. Acetylcholine release was monitored by intracellular recording of endplate potentials (e.p.ps). 2. Charybdotoxin (20-300 nM), but not apamin (10 nM-2.5 microM), selectively reduced the amplitude of an IK,Ca unmasked by prior blockade of the delayed rectifier K+ current with 3,4-diaminopyridine (3,4-DAP). 3. In the combined presence of 3,4-DAP and charybdotoxin, large Ca2+-dependent plateau responses developed, but only moderate and transient increases in acetylcholine release occurred. 4. In the absence of 3,4-DAP, charybdotoxin did not alter the electrical activity of, or the transmitter release from motor nerve terminals. 5. A possible role of the charybdotoxin-sensitive IK,Ca in the control of transmitter release is discussed.     

Effects of inorganic mercury on [3H]dopamine release and calcium homeostasis in rat striatal synaptosomes

Inorganic mercury (Hg2+) in vitro increases spontaneous transmitter release from nerve terminals. The mechanisms of action are not well understood but may involve alterations in intraterminal Ca2+ dynamics. In this study we describe actions of Hg2+ in vitro on isolated mammalian CNS striatal nerve terminals (synaptosomes). Cobalt (2 mM) completely blocked the effect of 2 microM Hg2+ on spontaneous [3H]dopamine release. Cadmium (100 microM) was equipotent to Co2+ in blocking depolarization-dependent [3H]dopamine release, but did not alter the 2 microM Hg2(+)-induced spontaneous [3H]dopamine release. Depolarization-dependent [3H]dopamine release was not altered by 5 microM Hg2+. It appears that the site of action of Hg2+ on spontaneous [3H]dopamine release is not the Ca2+ channel. The effects of Hg2+ on intraterminal ionized Ca2+ [( Ca2+]i) were evaluated using the Ca2(+)-specific fluorescent probe, fura-2. Hg2+ (1-8 microM) had no effect on [Ca2+]i in 1.2 mM Ca2(+)-containing buffers. In nominal Ca2+ media, 4 and 8 microM Hg2+ significantly decreased [Ca2+]i. Following exposure to 4 and 8 microM Hg2+ the quenching of extrasynaptosomal fura-2 by Mn2+ was increased, suggesting that Hg2+ facilitated the leakage of fura-2. This apparent leakage was probably due to a nonspecific increase in membrane permeability since 2 microM Hg2+ produced a Co2(+)-insensitive increase in [3H]deoxyglucose phosphate efflux. Hg2+ did not increase the leakage of either lactate dehydrogenase or soluble protein from synaptosomes. Hg2+ produced a concentration-dependent (1-8 microM) increase in 45Ca2+ efflux from superfused synaptosomes which was insensitive to blockade either by 2 mM Co2+ or by 100 microM Cd2+. These data suggest that the transmitter releasing action of Hg2+ involves interactions with sites that also interact with Co2+ but not with Cd2+. Furthermore, Hg2+ may have direct transmitter releasing actions (i.e., Ca2(+)-mimetic properties), as well as nonspecific actions on plasma membrane permeability which may not necessarily be linked to [3H]dopamine release.