Function of voltage-dependent Ca^<2+> channels in central nervous system neurons.

本项目的目的是利用钙离子成像技术研究中枢神经系统神经元树突中功能电压门控钙离子通道的空间分布，以了解钙离子通道的作用。几种不同类型的电压门控钙离子已被证明分布在中枢神经系统神经元中，并根据它们的药理和动力学特性进行分类。这些通道包括T、N、L和P型钙通道。从分离和培养神经元的研究中发现，小脑浦肯野神经元富含P型钙通道，低阈值钙通道也分布在浦肯野神经元中。我们制作了大鼠的小脑切片，并用全细胞移液管对单个浦肯野神经元进行了荧光指示剂钙离子标记。普肯野神经元的树突在TTx，a钠通道b…的存在下通过胞体去极化来刺激细胞内Ca~(2+)水平的变化用冷却的CCD摄像机进行测量。加入P型通道阻断剂可消除钙依赖性动作峰和伴随的一过性钙升高，使树突内钙显著升高。钙显像学研究表明，这种钙升高是由于低阈值钙通道的激活，在电压门控性K通道被药物阻断的情况下，这种钙升高被低阈值钙通道阻滞剂Ni^；根据这些发现，我们认为低阈值钙通道和低阈值钾通道沿小脑浦肯野神经元的树突分布。一项使用隔室模型计算重建浦肯野神经元的模拟研究表明，这些低阈值的钙通道和K^通道负责依赖于钙的平台电位。还需要进一步的研究来了解树突中的这些低阈值离子通道如何为整合中枢神经系统神经元的突触输入贡献机制。较少

The objective of this project was to study spatial distribution of functional voltage-gated Ca^<2+> channels in the dendrites of central nervous system neurons in slice preparations using a Ca^<2+> imaging technique in order to understand the roles of Ca^<2+> channels. Several different types of voltage-gated Ca^<2+> has been shown to be distributed in central nervous system neurons and classified according to their pharmacological and kinetic properties. Those include T, N, L, and P-type of Ca^<2+> channels.It has been reported from the studies using isolated or cultured neurons that cerebellar Purkinje neurons are rich in P-type Ca^<2+> channels, and that low-threshold Ca^<2+> channels are also distributed in Purkinje neurons. We made cerebellar slices from rats and stained individual Purkinje neurons with fluorescent Ca^<2+> indicator dye using whole-cell pipetts. The dendrites of Purkinje neurons were stimulated by depolarizing the cell body in the presence of TTX, a Na^+ channel b … More locker, and the change in intracellular Ca^<2+> level was measured with a cooled CCD camera. Addition of a blocker for P-type channel abolished Ca^<2+> dependent action spikes and accompanying transient Ca^<2+> increase leaving a significant Ca^<2+> rise in the dendrites. Ca^<2+> imaging study showed that this Ca^<2+> rise was due to activation of low-threshold Ca^<2+> channels and that under a condition where voltage-gated K^+ channels were pharmacologically blocked, this Ca^<2+> rise was suppressed by Ni^<2+> , a blocker for low-threshold Ca^<2+> channels. Based on these findings we concluded that low-threshold Ca^<2+> channels and low-threshold K^+ channels are distributed along the dendrites of cerebellar Purkinje neurons. A simulation study to computationally reconstruct Purkinje neurons using a compartmental model suggested that those low-threshold Ca^<2+> and K^+ channels are responsible for the Ca^<2+>-dependent plateau potential.Further studies are needed to understand how these low-threshold ion channels in the dendrites contribute the mechanisms for integrating synaptic inputs in central nervous system neurons. Less