Analysis of induction mechanisms of intrinsic optical signals by simultaneous measurement of extracellular K~ concentration and space volume.

众所周知，神经元兴奋引起中枢神经系统的细胞肿胀（或细胞外空间体积减少）。人们认为，细胞膨胀是光散射变化的基础，称为内在光信号（lOSs），从未染色的切片中记录下来。其他人和我们已经报道了速尿敏感的阳离子-氯共转运体在诱导lOSs中起重要作用。共转运体有Na-K-2C1和K-Cl两种类型，后者有4种异构体（KCC1 ~ 4）。我们建立了一种新的实时光学成像系统，利用冷却CCD相机来测量损耗。我们记录了外部K~浓度变化引起的损失。我们得到了以下结果：K^+浓度的微小变化（<2mM）可引起损失，响应的幅度取决于K^+浓度变化的量。降低K^+浓度引起相反的反应。此外，TTX.2对K^+诱导的lOSs没有抑制作用。在抑制共转运体的条件下（应用速尿或在低CL溶液中），K^+诱导的损失被抑制。低Na^+溶液对K^+诱导的损耗影响不大。对内整流K^+通道阻滞剂对K^+诱导的损失抑制作用不大。这些结果表明，在脊髓背角中，K-Cl共转运蛋白主要参与了lOSs的诱导，而在神经元中特异性表达的KCC2可能是这种反应的候选物质。

It is known that neuronal excitation causes cell swelling (or a decrease in extracellular space volume)in the central nervous system.It is thought that cell swelling underlies changes in light scattering, called inirinsic optical signals(lOSs), recorded from slices not stained with dyes.Others and we have reported that furosemide-sensitive cation-chloride cotransporters play an important role in the induction of lOSs.There are two types of cotransporters, Na-K-2C1 and K-Cl cotransporters, and the latter has 4 isoforms (KCC1〜4).We have set up a new reaJ-time optical imaging system using a cooled CCD camera to measure lOSs.We have recorded lOSs induced by changes in externai K~ concentration. We have obtained the following results.1.lOSs could be induced by small (<2mM) changes in K^+ concentration, and the amplitude of the responses was dependent on the amount of K^+ concentration changes.Lowering of K^+ concentration caused reversed responses.In addition, K^+-induced lOSs were not inhibited by addition of TTX.2.K^+-induced lOSs were suppressed under the conditions (application of furosemide or in low CL solution) to inhibit the cotransporters.3.K^+-induced lOSs were not much affected in low Na^+ solution.4.K^+-induced lOSs were little inhibited by blockers of the inwardly-rectifying K^+ channels.These results indicate that in the dorsal horn of the spinal cord the K-Cl cotransporters are mainly involved in the induction of lOSs and that KCC2, which expresses specifically in neurons, may be the likely candidate for the responses.