ANTIEPILEPTIC DRUG EFFECTS ON CA2+ AND GABA C1 CURRENT

这项研究的长期目标是更好地理解 抗癫痫药物治疗癫痫的作用机制研究进展 癫痫。拟议的研究将集中在以下领域 神经生理学、神经药理学和分子生物学。实验将会 旨在研究抗癫痫药物对电压的基本作用 依赖离子通道和传输门控离子通道。 电压依赖性钙通道将在小鼠和大鼠的原代培养中进行研究 和急性分离的中央和初级传入细胞培养 神经元。记录在这些组织中的钙电流成分包括 暂态低阈值电流(T)，大的暂态高阈值 电流(N)和缓慢失活的高阈值电流(L)。vbl.使用 全电池电压钳制技术，我们将确定所有的效果 临床使用的抗癫痫药物、活性代谢物和 戊四氮对T、N和L全细胞钙电流的影响。线性泄漏 并从随后记录的电流中减去电容电流 通过分析峰值电流、激活和失活动力学、电压 依赖性，以及从失活中恢复的时间过程。使用 细胞附着单通道记录技术，我们将确定 电压依赖性T、N和L钙通道的特性及 抗癫痫药物、活性代谢物和戊四唑的作用 在单通道电流上。记录的数据将被数字化并进行分析 通过计算机脱机。个别洋流将由 通道幅度、打开和关闭时间直方图以及 突发特征，包括频率、持续时间、分布、数量 以及开口和闭合的类型，以及冲击间隔。 非洲爪哇卵母细胞的递质门控氯离子通道将通过 注射后的细胞内记录技术及其表达 分离或克隆GABAA受体基因。单通道电流将是 在卵母细胞和快速稳定地转导cdna的哺乳动物中的研究 细胞系。重点将放在表达的特征上 受体对苯二氮卓类、巴比妥类和惊厥类药物的反应 并对其动力学性质进行了分析。

The long term objective of this research is a better understanding of the mechanisms of action of antiepileptic drugs used in the treatment of epilepsy. The proposed studies will concentrate within the fields of neurophysiology, neuropharmacology and molecular biology. Experiments will be designed to study the basic action of antiepileptic drugs on voltage dependent and transmitter gated ion channels. Voltage dependent calcium channels will be studied in mouse and rat primary and acutely dissociated cell cultures of central and primary afferent neurons. The calcium current components recorded in these tissues include a transient low threshold current (T), a large transient high threshold current (N), and a slowly inactivating high threshold current (L). Using the whole cell voltage clamp technique, we will determine the effect of all clinically used antiepileptic drugs, active metabolites, and pentylenetetrazol on T, N, and L whole cell calcium currents. Linear leak and capacitance currents will be subtracted from recorded currents followed by analysis of peak currents, activation and inactivation kinetics, voltage dependency, and the time course of recovery from inactivation. Using the cell attached single channel recording technique, we will determine the properties of voltage dependent T, N, and L calcium channels and the effects of antiepileptic drugs, active metabolites, and pentylenetetrazol on single channel currents. Recorded data will be digitized and analyzed off line by computer. Individual currents will be analyzed by the distribution of channel amplitudes, open and closed time histograms and burst characteristics including frequency, duration, distribution, number and types of openings and closings, and interburst interval. Transmitter gated chloride channels will be studied in Xenopus oocytes by intracellular recording techniques after injection and expression of either isolated or cloned GABA A receptor mRNA. Single channel currents will be studied in the oocytes and in acutely and stably cDNA transfected mammalian cell lines. Emphasis will be placed on the characterization of expressed receptors' responses to benzodiazepines, barbiturates, and convulsant drugs and on the analysis of kinetic properties.