King T S, Potter D, Kang I S, Norris C, Chen E, Schenken R S, Javors M A
Department of Cellular and Structural Biology, The University of Texas Health Science Center, San Antonio, TX 78284-7762, USA.
Brain Res. 1999 Apr 3;824(1):56-62. doi: 10.1016/s0006-8993(99)01163-4.
Immortalized GT1-7 neurons were used to characterize the effect of muscimol, a GABAA receptor agonist, to enhance pulsatile gonadotropin-releasing hormone (GnRH) release. GT1-7 neurons were grown on Cytodex-3 beads and placed in special superfusion microchambers. The cells were superfused at a rate of 6.2 ml x h-1 with Media 199 (pH 7.35) using a commercially available perfusion system. After a pre-muscimol period of 120 min, the cells were exposed for 5 min to 0.35, 1, 5 or 10 microM muscimol or 5 microM muscimol+20 microM of the GABAA receptor antagonist, bicuculline. Following removal of the muscimol (and bicuculline, in the case of the latter experiment), the superfusion was continued for another 115 min. Sample fractions were collected at 5 min intervals throughout the perfusion. Basal GnRH release from the GT1-7 neurons was pulsatile with an average interpulse interval of 45.4+/-0.5 min and an average pulse amplitude of 191.5+/-22.6 pg x min x ml-1. Our results also demonstrated that the GABAA receptor agonist, muscimol, enhances pulsatile GnRH release from GT1-7 neurons in culture. The response to muscimol was saturable and concentration-dependent with an EC50 of 0.47 microM. The effects of 5 microM muscimol to increase GnRH pulsatility were blocked by co-exposure to the GABAA receptor antagonist, bicuculline. The average GnRH interpulse intervals were 41.7+/-1.8 min, 32.5+/-2.9 min, 30.6+/-0.7 min and 25.5+/-0.4 min in the period following exposure to 0.35, 1, 5 and 10 microM of muscimol, respectively (post-muscimol period). GnRH pulse amplitude (mean-area for each pulse) was increased during exposure to muscimol but not during the pre- or post-muscimol periods. The GABAA receptor antagonist, bicuculline, itself had no effect on pulsatile GnRH release. These results are consistent with previously published reports suggesting that activation of the GABAA receptor stimulates hypothalamic GnRH release in embryonic and neonatal animals.
永生化的GT1-7神经元被用于研究γ-氨基丁酸A型（GABAA）受体激动剂蝇蕈醇增强促性腺激素释放激素（GnRH）脉冲式释放的作用。GT1-7神经元生长在Cytodex-3微珠上，并置于特殊的灌流微室中。使用市售的灌流系统，以6.2 ml×h-1的速率用199培养基（pH 7.35）对细胞进行灌流。在给予蝇蕈醇前120分钟后，将细胞暴露于0.35、1、5或10 μM的蝇蕈醇或5 μM蝇蕈醇 + 20 μM的GABAA受体拮抗剂荷包牡丹碱中5分钟。去除蝇蕈醇（在后一实验中还包括荷包牡丹碱）后，继续灌流115分钟。在整个灌流过程中，每隔5分钟收集一次样品馏分。GT1-7神经元基础GnRH释放呈脉冲式，平均脉冲间期为45.4±0.5分钟，平均脉冲幅度为191.5±22.6 pg×min×ml-1。我们的结果还表明，GABAA受体激动剂蝇蕈醇可增强培养的GT1-7神经元的GnRH脉冲式释放。对蝇蕈醇的反应具有饱和性且呈浓度依赖性，半数有效浓度（EC50）为0.47 μM。同时暴露于GABAA受体拮抗剂荷包牡丹碱可阻断5 μM蝇蕈醇增加GnRH脉冲频率的作用。暴露于0.35、1、5和10 μM蝇蕈醇后的时期（蝇蕈醇作用后时期），GnRH平均脉冲间期分别为41.7±1.8分钟、32.5±2.9分钟、30.6±0.7分钟和25.5±0.4分钟。GnRH脉冲幅度（每个脉冲的平均面积）在暴露于蝇蕈醇期间增加，但在蝇蕈醇作用前或作用后时期未增加。GABAA受体拮抗剂荷包牡丹碱本身对GnRH脉冲式释放无影响。这些结果与先前发表的报告一致，表明GABAA受体的激活可刺激胚胎和新生动物下丘脑GnRH的释放。
