Louiset E, Mei Y A, Valentijn J A, Vaudry H, Cazin L
European Institute for Peptide Research, Laboratory of Molecular Endocrinology, CNRS URA 650, UA INSERM, University of Rouen, Mont-Saint-Aignan, France.
J Neuroendocrinol. 1994 Feb;6(1):39-46. doi: 10.1111/j.1365-2826.1994.tb00553.x.
The molecular mechanisms regulating GABAA receptor activity in cultured frog melanotrophs were studied using the patch-clamp technique. In the whole-cell configuration, application of GABA evoked a dose-related increase of inward chloride currents. The ED50 value, estimated from the sigmoidal dose-response curve was 2 x 10(-6) M and the Hill coefficient was 1.55. The amplitude of the GABA-induced current decayed with time. Kinetics analysis of the desensitization revealed that the time-course of the current decrement was fitted by one exponential. Graded doses of GABA or association of GABA with the benzodiazepine receptor agonist flunitrazepam accelerated the desensitization process. In contrast, the time-course of the current did not significantly vary at different holding potentials. In the outside-out configuration, GABA was found to activate channels which displayed three unitary conductance levels (8, 15 and 30 pS). The channel openings of the more frequent conductance level (30 pS) exhibited short and long lasting open states (1.2 and 28.3 ms at -60 mV). Altogether these data reveal that frog melanotrophs possess a single population of GABAA receptors which interconvert into a higher affinity state in the presence of benzodiazepine receptor agonists. Two GABA molecules must bind to the receptor to trigger long lasting channel openings. In addition, the activity of the GABAA receptor appears to be independent of the accumulation of intracellular chloride ions.
利用膜片钳技术研究了培养的青蛙黑素细胞中调节γ-氨基丁酸A（GABAA）受体活性的分子机制。在全细胞模式下，应用γ-氨基丁酸（GABA）可引起内向氯离子电流呈剂量相关增加。根据S形剂量反应曲线估计的半数有效剂量（ED50）值为2×10⁻⁶ M，希尔系数为1.55。GABA诱导的电流幅度随时间衰减。脱敏动力学分析表明，电流衰减的时间进程符合单指数函数。不同剂量的GABA或GABA与苯二氮䓬受体激动剂氟硝西泮联合使用可加速脱敏过程。相反，在不同的钳制电位下，电流的时间进程没有显著变化。在外侧向外模式下，发现GABA可激活显示三种单位电导水平（8、15和30 pS）的通道。更频繁出现的电导水平（30 pS）的通道开放呈现短和长两种持续时间的开放状态（在-60 mV时分别为1.2和28.3 ms）。这些数据表明，青蛙黑素细胞具有单一群体的GABAA受体，在苯二氮䓬受体激动剂存在时可转变为更高亲和力状态。两个GABA分子必须与受体结合才能触发长时间的通道开放。此外，GABAA受体的活性似乎与细胞内氯离子的积累无关。
