Wu Yuanming, Wang Wengang, Richerson George B
Department of Neurology, LCI-712B, Yale University School of Medicine, 15 York St., PO 208018, New Haven, CT 06520-8018, USA.
J Neurophysiol. 2006 Nov;96(5):2425-36. doi: 10.1152/jn.00545.2006. Epub 2006 Jul 26.
Tonic inhibition is widely believed to be caused solely by "spillover" of GABA that escapes the synaptic cleft and activates extrasynaptic GABA(A) receptors. However, an exclusively vesicular source is not consistent with the observation that tonic inhibition can still occur after blocking vesicular release. Here, we made patch-clamp recordings from neurons in rat hippocampal cultures and measured the tonic current that was blocked by bicuculline or gabazine. During perforated patch recordings, the tonic GABA current was decreased by the GAT1 antagonist SKF-89976a. Zero calcium solution did not change the amount of tonic current, despite a large reduction in vesicular GABA release. Perturbations that would be expected to alter the transmembrane sodium gradient influenced the tonic current. For example, in zero calcium Ringer, TTX (which can decrease cytosolic [Na(+)]) reduced tonic current, whereas veratridine (which can increase cytosolic [Na(+)]) increased tonic current. Likewise, removal of extracellular sodium led to a large increase in tonic current. The increases in tonic current induced by veratridine and sodium removal were completely blocked by SKF89976a. When these experiments were repeated in hippocampal slices, similar results were obtained except that a GAT1- and GAT3-independent nonvesicular source(s) of GABA was found to contribute to the tonic current. We conclude that multiple sources can contribute to ambient GABA, including spillover and GAT1 reversal. The source of GABA release may be conceptually less important in determining the amount of tonic inhibition than the factors that control the equilibrium of GABA transporters.
人们普遍认为，强直性抑制完全是由逸出突触间隙并激活突触外GABA(A)受体的GABA“溢出”引起的。然而，仅囊泡来源与以下观察结果不一致，即阻断囊泡释放后仍可发生强直性抑制。在这里，我们对大鼠海马培养物中的神经元进行了膜片钳记录，并测量了被荷包牡丹碱或加巴喷丁阻断的强直电流。在穿孔膜片记录期间，GAT1拮抗剂SKF-89976a可降低强直GABA电流。零钙溶液并没有改变强直电流的大小，尽管囊泡GABA释放大幅减少。预期会改变跨膜钠梯度的扰动会影响强直电流。例如，在零钙林格氏液中，TTX（可降低胞质[Na⁺]）可降低强直电流，而藜芦碱（可增加胞质[Na⁺]）可增加强直电流。同样，去除细胞外钠会导致强直电流大幅增加。藜芦碱和钠去除诱导的强直电流增加被SKF89976a完全阻断。当在海马切片中重复这些实验时，得到了类似的结果，只是发现一种不依赖GAT1和GAT3的非囊泡GABA来源对强直电流有贡献。我们得出结论，多种来源可导致周围环境中的GABA增加，包括溢出和GAT1逆转。在确定强直性抑制的量方面，GABA释放的来源在概念上可能不如控制GABA转运体平衡的因素重要。
