Effects of general anesthetics on calcium ion concentration in presynaptic nerve terminals attached to dissociated mammalian neurons

众所周知，全麻药在较低浓度下影响神经传递，而不是损害传导。越来越多的证据表明，除5-羟色胺外，全身麻醉药可增强GABA或甘氨酸的抑制性传递，抑制乙酰胆碱和谷氨酸的兴奋性传递。已经用分离神经元的突触后受体进行了复杂的传递实验。然而，几乎没有证据表明麻醉剂对突触前过程的影响，而突触前过程是传递的重要组成部分。在本研究中，我们测量了机械分离的大鼠海马神经元突触前神经末梢的钙离子浓度。Wistar大鼠在戊巴比妥麻醉下断头，取脑切片。将火抛光微玻璃吸管固定在海马区，以2赫兹频率振动3min。用突触前…获得分离的神经元附加了更多IC端子(Bouton)。FM1-43鉴定胞浆内钙离子浓度，FM-3AM激光共聚焦显微镜下测定胞内钙离子浓度。1)高浓度的细胞外钾使神经元胞体和胞外胞体内[Ca~(2+)&Gt；]_i升高，无钙溶液中这种升高被取消，说明去极化诱导的[Ca~(^&lt；2+&Gt；]_i-升高依赖于钙离子的进入。2)谷氨酸(1-300微米)使[Ca^&lt；2+&Gt；]_i升高。[_i]_i在神经元胞体和胞核均呈浓度依赖性。3)CNQX(30微米)显著抑制谷氨酸诱导的[Ca^&lt；2+&gt；]_i升高。4)L类钙通道阻滞剂(尼莫地平、FS2、钙胞苷)抑制谷氨酸诱导的[Ca^&lt；2+&gt；]_i升高。5)上述结果提示谷氨酸诱导的[Ca^&lt；2+&gt；]i升高。6)戊巴比妥抑制谷氨酸引起的[Ca~(2+)]_i升高，对胞体和胞体[Ca~(2+)]_i的升高有较大影响。这一结果提示，戊巴比妥通过抑制突触前神经末梢的钙通道来调节神经传递。较少

It is known that general anesthetics affect neurotransmission at lower concentrations than they impair conduction. Accumulating evidence shows that general anesthetics enhance inhibitory transmission carried by GABA or glycine, and that they depress excitatory transmission by Ach or glutamate, with an exception of that by 5HT. Sophisticated experiments on transmission have been performed with post synaptic receptors of dissociated neurons. There is little evidence, however, about the anesthetics effects on the presynaptic processes, which compose an important part of transmission. In the present study, we measure the calcium ion concentration in the presynaptic nerve terminal, attached to mechanically dissociated neurons of rat hippocampus.Wistar rats were decapitated under pentobarbital anesthesia and the brain was removed and sliced. Fire-polished micro glass pipette was attached on the hippocampal region and vibrated at 2Hz for 3min. Dissociated neurons were obtained, with presynapt … More ic terminal (bouton) attached. The boutons were identified with FM1-43 and the intracellular calcium ion concentration [Ca^<2+>]_i was measured with fm-3AM under confocal laser microscope1) High concentration of extracellular K increased the [Ca^<2+>]_i both in neuronal soma and in the bouton This increase was abolished in Ca-free solution, indicating that depolarization induced [Ca^<2+>]_i-increase depends on Ca entry.2) Glutamate (1-300 micro M) increased the [Ca^<2+>]_i both in neuronal soma and in the bouton in a concentration dependent manner. The increases did not occur in the absence of Ca^<2+>.3) CNQX (30 micro M) markedly suppressed the glutamate-induced increases in [Ca^<2+>]_i..4) L-type calcium channel blockers (nimodipine, FS2, calcicludine) inhibited the glutamate-induced increases in [Ca^<2+>]_i. concentration dependently.5) These findings suggest that the glutamate-induced increases in [Ca^<2+>]_i. may be attributed, in part, to opening of L-type calcium channels, which is produced by activation of AMPA/kainate type glutamate receptor.6) Pentobarbital depressed the glutamate-induced increases in [Ca^<2+>]_i. both in soma and in boutons, with greater effects on boutons. This result suggests that pentobarbital modulates neurotransmission via inhibition of calcium channel at the presynaptic nerve terminal. Less