O2 deprivation inhibits Ca2+-activated K+ channels cytosolic factors in mice neocortical neurons
O2 deprivation inhibits Ca2+-activated K+ channels cytosolic factors in mice neocortical neurons
复制标题
DOI：
10.1172/jci7291
复制
发表时间：
1999-09-01
影响因子：
15.9
通讯作者：
Haddad, GG
中科院分区：
文献类型：
--
作者：
Liu, HJ;Moczydlowski, E;Haddad, GG
O-2 剥夺会诱导哺乳动物中枢神经元膜去极化。这种缺氧引起的去极化可能部分是由 K+ 通道的抑制介导的。因此，我们使用膜片钳技术进行了实验，并从小鼠新皮质中分离了神经元。在细胞贴壁和内向外结构中均观察到三种类型的 K+ 通道，但只有其中一种对 O-2 缺乏敏感。这种 O-2 敏感 K+ 通道被鉴定为大电导 Ca2+ 激活 K+ 通道 (BKCa)，因为它在对称 K+ (140 mM) 条件下表现出 210 pS 的大电导、激活的强烈电压依赖性以及对 Ca2+ 的显着敏感性。低 O-2 介质（PO2 = 10-20 mmHg）以电压依赖性方式显着抑制细胞附着斑块中 BKCa 通道开放概率，但在由内而外的斑块中则不然，表明 O-2 剥夺对小鼠新皮质神经元 BKCa 通道的影响是通过细胞质依赖性过程介导的。在 Mg-ATP 存在的情况下，降低细胞内 pH (pH(i)) 或在胞质中添加 cAMP 依赖性蛋白激酶 A 的催化亚基，可通过降低 BKCa 通道对 Ca2+ 的敏感性来导致 BKCa 通道活性降低。相反，还原剂谷胱甘肽和 DTT 增加了单个 BKCa 通道开放概率，而不影响单一电导。我们认为，在新皮质神经元中，(a) BKCa 通过胞质因子和胞质依赖性过程受到 O-2 剥夺的调节，(b) 缺氧期间通道活性的减少可能是由于胞质变化（例如 pH(i) 和磷酸化）导致 Ca2+ 敏感性降低。由于 BKCa 通道电导率大且在新皮质中普遍存在，因此可将其视为急性缺氧或缺血情况下药物干预的目标。
O-2 deprivation induces membrane depolarization in mammalian central neurons. It is possible that this anoxia-induced depolarization is partly mediated by an inhibition of K+ channels. We therefore performed experiments using patch-clamp techniques and dissociated neurons from mice neocortex. Three types of K+ channels were observed in both cell-attached and inside-out configurations, but only one of them was sensitive to lack of O-2. This O-2-sensitive K+ channel was identified as a large-conductance Ca2+-activated K+ channel (BKCa) as it exhibited a large conductance of 210 pS under symmetrical K+ (140 mM) conditions, a strong voltage-dependence of activation, and a marked sensitivity to Ca2+. A low-O-2 medium (PO2 = 10-20 mmHg) markedly inhibited this BKCa channel open probability in a voltage-dependent manner in cell-attached patches, but not in inside-out patches, indicating that the effect of O-2 deprivation on BKCa channels of mice neocortical neurons was mediated via cytosol-dependent processes. Lowering intracellular pH (pH(i)) or cytosolic addition of the catalytic subunit of a cAMP-dependent protein kinase A in the presence of Mg-ATP, caused a decrease in BKCa, channel activity by reducing the sensitivity of this channel to Ca2+. In contrast, the reducing agents glutathione and DTT increased single BKCa channel open probability without affecting unitary conductance. We suggest that in neocortical neurons, (a) BKCa is modulated by O-2 deprivation via cytosolic factors and cytosol-dependent processes, and (b) the reduction in channel activity during hypoxia is likely due to reduced Ca2+ sensitivity resulting from cytosolic alternations such as in pH(i) and phosphorylation. Because of their large conductance and prevalence in the neocortex, BKCa channels may be considered as a target for pharmacological intervention in conditions of acute anoxia or ischemia.