MICROGLIA GENERATE EXTERNAL H+ & K+ ION GRADIENTS UTILIZING H & K ATPASE FAMILY

从新生大鼠脑中分离小胶质细胞并培养7 天，积极挤出质子，以换取钾离子， 一个梯度的过剩质子和赤字的钾离子在接近 靠近细胞膜的外表面。 这些 浓度梯度，用离子选择性 自参比电极，在10 mm距离内消散 并被高浓度的 细胞外钾离子或质子。 药理学和 酶化学研究表明，H ~+/K ~+-ATP酶是细胞的主要酶。 这些梯度的主要生成器。 Na+/K+-ATP酶和 内向整流钾通道对 这些梯度的产生。 5 mM细胞外钾 浓度，记录梯度为-9.43 +4.2 mM（n=48） 在离细胞膜10 mm的距离上消散。 而 转运蛋白活性可被奥美拉唑（10 mM）阻断， 特异性H+/K+-ATP酶阻断剂SCH 28080（1 mM）对 哇巴因和毒毛旋花子定。 胶质细胞转运蛋白K 离子的数量级（3.7 mM）高于 上皮细胞H+/K+-ATP酶。 这是首次报道的H+/K+ 小胶质细胞中的转运蛋白，Kd在 [K+]出来。 H ~+/K ~+-ATP酶对钾稳态的影响 在高细胞外钾和低pH下的小胶质细胞，如在 脑损伤的部位进行了讨论。

Microglia isolated from newborn rat brains and cultured for 7 days, actively extrude protons in exchange for potassium ions creating a gradient of excess protons and a deficit of potassium ions in close proximity to the outer surface of the cell membrane. These concentration gradients, measured with an ion selective self-referencing electrode, dissipate over a distance of 10mm away from the cell and are activated by high concentrations of extracellular potassium ions or protons. Pharmacological and immunohisto-chemical studies revealed that an H+/K+-ATPase is the major generator of these gradients. Neither the Na+/K+-ATPase nor the inward rectifier potassium channel make significant contributions to the generation of these gradients. At 5mM extracellular potassium concentration, a gradient of -9.43+ 4.2 mM (n=48) was recorded dissipating over a distance of 10mm from the cell membrane. While the transporter activity could be blocked by Omeprazole (10mM) and by the specific H+/K+-ATP ase blocker SCH28080 (1mM), it was insensitive to ouabain and strophantidine. The Kd of the glial transporter for K ions is an order of magnitude higher (3.7 mM) than that of the epithelial H+/K+-ATPase. This is a first report of an H+/K+ transporter in microglial cells with Kd in the physio-logical range of [K+]out. Implications of the H+/K+-ATPase on potassium homeostasis in microglia under high extracellular potassium and low pH, as found at the site of brain injury, are discussed.