Developmental Regulation of inhibitory neuronal circuits by neural activity

利用gramicidin穿孔贴片记录模式，研究了不同发育阶段LSO神经元细胞内Cl- i的调节机制。在出生后第二周，由于[Cl-] i减少，对甘氨酸的反应由去极化变为超极化。速尿同样改变了未成熟和成熟LSO神经元的[Cl-] i，表明速尿敏感的细胞内Cl-调节因子有重要作用；即K+- cl -共转运体（KCC）和Na+-K+- cl -共转运体（NKCC），在整个早期发育过程中。细胞外K+浓度升高和细胞内K+被Cs+取代导致[Cl-] i在出生后(P) 13-15时升高，但在P -2时没有升高，这表明神经元Cl-挤压的机制对速尿和K+梯度都很敏感，并且在未成熟的LSO神经元中发育不良。此外，细胞外Na+的去除在p -2时降低了[Cl-] i，表明未成熟LSO神经元中存在细胞外Na+依赖性和速尿敏感的Cl积累。这些数据清楚地表明，Cl-共转运体的发育变化改变了[Cl-] i，并负责LSO神经元从新生儿Cl-外排到成熟Cl-内流的转换。Cl-共转运体的这种成熟变化可能对甘氨酸能和gaba能突触传递具有重要的功能作用，并对LSO和听觉发育具有更广泛的影响。

The regulatory mechanisms of intracellular Cl-concentration ([Cl-] i ) were investigated in the lateral superior olive (LSO) neurons of various developmental stages by taking advantage of gramicidin perforated patch recording mode which enables neuronal [Cl-] i measurement. Responses to glycine changed from depolarization to hyperpolarization during the second week after birth, resulting from [Cl-] i decrease. Furosemide equally altered the [Cl-] i of both immature and mature LSO neurons, indicating substantial contributions of furosemide-sensitive intracellular Cl- regulators ; i.e. K+-Cl- cotransporter (KCC) and Na+-K+-Cl- cotransporter (NKCC), throughout this early development. Increase of extracellular K+ concentration and replacement of intracellular K+ with Cs+ resulted in [Cl-] i elevation at postnatal day (P) 13-15, but not at P0-2, indicating that the mechanism of neuronal Cl- extrusion is sensitive to both furosemide and K+-gradient, and poorly developed in immature LSO neurons. In addition, removal of extracellular Na+ decreased [Cl-] i at P0-2, suggesting the existence of extracellular Na+- dependent and furosemide-sensitive Cl-accumulation in immature LSO neurons. These data show clearly that developmental changes of Cl--cotransporters alter [Cl-] i and are responsible for the switch from the neonatal Cl- efflux to the mature Cl- influx in LSO neurons. Such maturational changes in Cl--cotransporters might have the important functional roles for glycinergic and GABAergic synaptic transmission and the broader implications for LSO and auditory development.