Song W J, Baba Y, Otsuka T, Murakami F
Department of Electronic Engineering, Graduate School of Engineering, Osaka University, Suita 565-0871, Japan.
J Neurophysiol. 2000 Nov;84(5):2630-7. doi: 10.1152/jn.2000.84.5.2630.
The subthalamic nucleus (STN) plays a key role in motor control. Although previous studies have suggested that Ca(2+) conductances may be involved in regulating the activity of STN neurons, Ca(2+) channels in this region have not yet been characterized. We have therefore investigated the subtypes and functional characteristics of Ca(2+) conductances in STN neurons, in both acutely isolated and slice preparations. Acutely isolated STN cells were identified by retrograde filling with the fluorescent dye, Fluoro-Gold. In acutely isolated STN neurons, Cd(2+)-sensitive, depolarization-activated Ba(2+) currents were observed in all cells studied. The current-voltage relationship and current kinetics were characteristic of high-voltage-activated Ca(2+) channels. The steady-state voltage-dependent activation curves and inactivation curves could both be fitted with a single Boltzmann function. Currents evoked with a prolonged pulse, however, inactivated with multiple time constants, suggesting either the presence of more than one Ca(2+) channel subtype or multiple inactivation processes with a single channel type in STN neurons. Experiments using organic Ca(2+) channel blockers revealed that on average, 21% of the current was nifedipine sensitive, 52% was sensitive to omega-conotoxin GVIA, 16% was blocked by a high concentration of omega-agatoxin IVA (200 nM), and the remainder of the current (9%) was resistant to the co-application of all blockers. These currents had similar voltage dependencies, but the nifedipine-sensitive current and the resistant current activated at slightly lower voltages. omega-Agatoxin IVA at 20 nM was ineffective in blocking the current. Together, the above results suggest that acutely isolated STN neurons have all subtypes of high-voltage-activated Ca(2+) channels except for P-type, but have no low-voltage-activated channels. Although acutely isolated neurons provide a good preparation for whole cell voltage-clamp study, dendritic processes are lost during dissociation. To gain information on Ca(2+) channels in dendrites, we thus studied Ca(2+) channels of STN neurons in a slice preparation, focusing on low-voltage-activated channels. In current-clamp recordings, a slow spike was always observed following termination of an injected hyperpolarizing current. The slow spike occurred at resting membrane potentials and was sensitive to micromolar concentrations of Ni(2+), suggesting that it is a low-threshold Ca(2+) spike. Together, our results suggest that STN neurons express low-voltage-activated Ca(2+) channels and several high-voltage-activated subtypes. Our results also suggest the possibility that the low-voltage-activated channels have a preferential distribution to the dendritic processes.
丘脑底核（STN）在运动控制中起关键作用。尽管先前的研究表明Ca(2+)电导可能参与调节STN神经元的活动，但该区域的Ca(2+)通道尚未得到表征。因此，我们研究了急性分离和脑片制备中STN神经元Ca(2+)电导的亚型和功能特性。通过用荧光染料Fluoro-Gold逆行填充来鉴定急性分离的STN细胞。在所有研究的急性分离的STN神经元中，均观察到对Cd(2+)敏感、去极化激活的Ba(2+)电流。电流-电压关系和电流动力学具有高电压激活Ca(2+)通道的特征。稳态电压依赖性激活曲线和失活曲线均可用单个玻尔兹曼函数拟合。然而，用延长脉冲诱发的电流以多个时间常数失活，这表明STN神经元中要么存在不止一种Ca(2+)通道亚型，要么在单一通道类型中存在多个失活过程。使用有机Ca(2+)通道阻滞剂的实验表明，平均而言，21%的电流对硝苯地平敏感，52%对ω-芋螺毒素GVIA敏感，16%被高浓度的ω-阿加毒素IVA（200 nM）阻断，其余电流（9%）对所有阻滞剂的联合应用具有抗性。这些电流具有相似的电压依赖性，但硝苯地平敏感电流和抗性电流在略低的电压下激活。20 nM的ω-阿加毒素IVA对阻断电流无效。综上所述，上述结果表明，急性分离的STN神经元除了P型外具有所有高电压激活Ca(2+)通道亚型，但没有低电压激活通道。尽管急性分离的神经元为全细胞电压钳研究提供了良好的标本，但在解离过程中树突过程会丢失。为了获取有关树突中Ca(2+)通道的信息，我们因此在脑片制备中研究了STN神经元的Ca(2+)通道，重点关注低电压激活通道。在电流钳记录中，在注入的超极化电流终止后总是观察到一个慢波峰。慢波峰出现在静息膜电位，并且对微摩尔浓度的Ni(2+)敏感，表明它是一个低阈值Ca(2+)波峰。综上所述，我们的结果表明STN神经元表达低电压激活Ca(2+)通道和几种高电压激活亚型。我们的结果还表明低电压激活通道可能优先分布于树突过程。
