GABA-induced currents and their modulation by neurosteroids in rat GnRH neurons

分析大鼠促性腺激素释放激素(GnRH)神经元电压门控性离子通道电流和GABAA受体电流。在深度麻醉下，取成年GnRH-EGFP转基因大鼠的脑组织。解剖后，用酶法分散大脑，将神经元接种在盖片上，培养过夜。然后应用神经元进行穿孔膜片钳实验。GnRH神经元呈电压门控性钙电流，由L型、N-型、P/Q-型、R-型和T型组成。R型钙电流占总钙电流的30%。L型、N型、P/Q型和T型分别占25%、20%、12%和13%(Kato等人，2006年)。这种表达模式与新生GnRH神经元不同，但与青春期神经元的表达模式相似。因此，大鼠促性腺激素释放激素神经元中电压门控钙通道的表达模式是在青春期前后发育调节和完成的。为了研究这些钙通道的作用，我们研究了钙激活的…更多酸化的钾电流。大鼠促性腺激素释放激素神经元呈现两种钙激活钾电流。它们是钙激活和电压激活的钾电流(BK)和钙激活的慢后超极化电流。后者可被SK通道阻滞剂阿帕明特异性抑制。因此，这种缓慢的后超极化电流通过SK通道传输。此外，这种电流有助于强去极化引起的持续放电(Kato等人，2005年，日本神经科学会议；Kato等人，2006年)。最后，我们用钙显像法分析了GABA对大鼠GnRH神经元的作用。GABA作用于GABAA受体，使50%以上的成年大鼠GnRH神经元内钙离子浓度升高。这种GABA诱导的反应被细胞外钙的去除和氯离子转运体NKCC1的阻断所消除。因此，GABA通过作用GABAA受体使大鼠GnRH神经元去极化，从而激活电压门控钙通道(Tanaka等人。2006年，国际神经内分泌学大会；渡边等人，2006年正在筹备中)。较少

Voltage-gated ionic channel currents and GABAA receptor current in rat GnRH neurons were analyzed. Under deep anesthesia, brains were excised from adult GnRH-EGFP transgenic rats. After dissection, brains were enzymatically dispersed and the neurons were plated on coverslips and cultured overnight. Then the neurons were applied for perforated patch-clamp experiment. GnRH neurons, identified with EGFP-fluorescnce, exhibited the voltage-gated calcium currents, which were composed with L-, N-, P/Q-, R- and T-types. The R-type contributed 30% for total calcium current. L-, N-, P/Q- and T-type contributed 25, 20, 12 and 13%, respectively (Kato et al., 2006). This expression pattern differs from those of neonatal GnRH neurons but is similar to those from pubertal neurons. Therefore, expression pattern of voltage-gated calcium channels in rat GnRH neurons are developmentally regulated and completed around puberty. To study the roles of these calcium channels, we investigated the calcium-activ … More ated potassium currents. Rat GnRH neurons showed two types of calcium activated potassium currents. They were calcium- and voltage-activated potassium current (BK) and calcium-activated slow afterhyperpolarization current. The latter was specifically inhibited by SK channel blocker apamin. This slow afterhyperpolarization current is, therefore, carried through the SK channels. Furthermore, this current contributed for persistent firing evoked by strong depolarization (Kato et al., 2005, Japan Neuroscience Meeting ; Kato et al., 2006). Finally, we analyzed GABA action on rat GnRH neurons by means of calcium imaging method. GABA acted on the GABAA receptor and elevated intracellular calcium concentration in more than 50% of GnRH neurons isolated from adult rats. This GABA-induced response was abolished by the removal of extracellular calcium and by blocking the chloride transporter NKCC1. Therefore, GABA depolarizes rat GnRH neurons by acting GABAA receptor, thereby activating the voltage-gated calcium channels (Tanaka et al. 2006, International Congress of Neuroendocrinology; Watanabe et al., 2006 in preparation). Less