Zhan R Z, Fujiwara N, Tanaka E, Shimoji K
Department of Anesthesiology, Niigata University School of Medicine, Asahimachi-dori, Japan.
Brain Res. 1998 Jan 5;780(1):86-94. doi: 10.1016/s0006-8993(97)01149-9.
To elucidate the mechanism of pHi changes induced by membrane depolarization, the variations in pHi and [Ca2+]i induced by a number of depolarizing agents, including high K+, veratridine, N-methyl-D-aspartate (NMDA) and ouabain, were investigated in rat hippocampal slices by the fluorophotometrical technique using BCECF or fura-2. All of these depolarizing agents elicited a decrease in pHi and an elevation of intracellular calcium ([Ca2+]i) in the CA1 pyramidal cell layer. The increases in [Ca2+]i caused by the depolarizing agents almost completely disappeared in the absence of Ca2+ (0 mM Ca2+ with 1 mM EGTA). In Ca2+ free media, pHi acid shifts produced by high K+, veratridine or NMDA were attenuated by 10-25%, and those produced by ouabain decreased by 50%. Glucose-substitution with equimolar amounts of pyruvate suppressed by two-thirds the pHi acid shifts induced by both high K+ and NMDA. Furthermore, lactate contents were significantly increased in hippocampal slices by exposure to high K+, veratridine or NMDA but not by ouabain. These results suggest that the intracellular acidification produced by these depolarizing agents, with the exception of ouabain, is mainly due to lactate accumulation which may occur as a result of accelerated glycolysis mediated by increased Na+-K+ ATPase activity. A Ca2+-dependent process may also contribute to the intracellular acidification induced by membrane depolarization. Since an increase in H+ concentration can attenuate neuronal activity, glycolytic acid production induced by membrane depolarization may contribute to the mechanism that prevents excessive neuronal excitation.
为阐明膜去极化诱导细胞内pH值（pHi）变化的机制，我们采用荧光光度技术，利用BCECF或fura-2，研究了多种去极化剂（包括高钾、藜芦碱、N-甲基-D-天冬氨酸（NMDA）和哇巴因）在大鼠海马切片中诱导的pHi和细胞内钙浓度（[Ca2+]i）的变化。所有这些去极化剂均引起CA1锥体细胞层pHi降低和细胞内钙（[Ca2+]i）升高。在无钙（含1 mM乙二醇双乙醚二胺四乙酸（EGTA）的0 mM Ca2+）条件下，去极化剂引起的[Ca2+]i升高几乎完全消失。在无钙培养基中，高钾、藜芦碱或NMDA引起的pHi酸偏移减弱了10%-25%，而哇巴因引起的pHi酸偏移降低了50%。用等摩尔量的丙酮酸替代葡萄糖可使高钾和NMDA诱导的pHi酸偏移减少三分之二。此外，暴露于高钾、藜芦碱或NMDA可使海马切片中的乳酸含量显著增加，但哇巴因不会。这些结果表明，除哇巴因外，这些去极化剂引起的细胞内酸化主要是由于乳酸积累，这可能是由增加的钠钾ATP酶活性介导的糖酵解加速所致。钙依赖过程也可能导致膜去极化诱导的细胞内酸化。由于氢离子浓度升高可减弱神经元活动，膜去极化诱导的糖酵解产酸可能有助于防止神经元过度兴奋的机制。
