ELECTROPHYSIOLOGICAL ANALYSIS OF THE SYNAPTIC ACTIVITY OF NONSPIKING INTERNEURONS IN BEHAVING ANIMALS USING WHOLE CELL RECORDING TECHNIQUES

为了巩固将全细胞记录技术应用于克氏原虾中枢神经系统非尖峰中间神经元执行行为行为的实验背景，我重点研究了1)可应用微电极和贴片电极的非麻醉全动物制备的新进展，2)使用单电极电压钳(SEVC)技术对非尖峰中间神经元膜电流的电生理分析，以及3)在解剖显微镜下将贴片电极应用于原位的中枢神经元。使用新开发的跑步机系统，非尖峰中间神经元的突触活动可以通过动物行走在细胞内的玻璃微电极来记录。SEVC实验表明，非棘突中间神经元的树突膜具有三种类型的去极化依赖的钾电导。在解剖显微镜下，在腹神经节机械性脱脂暴露的中枢神经元胞体上应用贴片电极可记录到单个钾通道的自发活动。在无脊椎动物中，到目前为止，使用原位全细胞记录技术进行的研究还很少。这项研究的结果表明，在进行行为行为的动物中，将用于全细胞记录的贴片电极应用于原位的中枢神经元，以便从电生理学的角度分析与特定行为相关的突触电流，在技术上是可行的。

In order to consolidate the experimental background for applying the whole-cell recording technique to nonspiking interneurons in the central nervous system of crayfish, Procambarus clarkii Girard, performing behavioral acts, I have focused this study on 1) new development of an unanesthetized whole animal preparation to which micro- and patch electrodes could be applicable, 2) electrophysiological analyses of membrane current of nonspiking interneurons using the single electrode voltage clamp (SEVC) technique, and 3) application of patch electrodes to central neurons in situ under a dissecting microscope. Using the newly developed treadmill system, the synaptic activity of nonspiking interneurons could be recorded intracellular by a glass microelectrode from an animal walking on it. The SEVC experiments have revealed that the dendritic membrane of nonspiking interneurons possesses three types of depolarization-dependent potassium conductances. Under a dissecting microscope, spontaneous activities of single potassium channels could be recorded by a patch electrode applied to the cell bodies of central neurons in situ exposed by mechanical desheathing of the abdominal ganglion. In invertebrate animals, only a few studies have been carried out so far using the in situ whole-cell recording technique. The results of this study indicate that it is technically possible to apply patch electrodes for whole cell recording to the central neurons in situ in the animal performing behavioral acts so that the synaptic currents associated with specific behavior can be analyzed electrophysiologically.