Acid/base transport in glial cells of Necturus optic nerve

Membrane properties of glial cells of Necturus maculosus optic nerve were studied using conventional microelectrodes and double-barreled, pH-sensitive microelectrodes. The effects of Ba{dollar}sp{lcub}++{rcub}{dollar}, a known K{dollar}sp+{dollar}-channel blocker, were studied with conventional microelectrodes. The addition of Ba{dollar}sp{lcub}++{rcub}{dollar} (2-5 mM) reversibly depolarized glial cells by 20 to 50 mV and reduced the sensitivity of the membrane to changes in bath (K{dollar}sp+{dollar}). These effects were accompanied by an increase in the input resistance of the membrane, strongly suggesting that Ba{dollar}sp{lcub}++{rcub}{dollar} decreases the K{dollar}sp+{dollar} conductance of the glial cells. With the K{dollar}sp+{dollar} conductance decreased by Ba{dollar}sp{lcub}++{rcub}{dollar}, the membrane response to HCO{dollar}sb3sp-{dollar} was investigated. Addition of HCO{dollar}sb3sp-{dollar} at constant pH caused a hyperpolarization which was Na{dollar}sp+{dollar}-dependent, SITS (4-acetamido-4{dollar}spprime{dollar}-isothiocyanato-stilbene-2,2{dollar}spprime{dollar}-disulfonic acid)-sensitive, Cl{dollar}sp-{dollar}-independent, and strophanthidin-insensitive. These results strongly suggest the presence in the glial membrane of an electrogenic Na{dollar}sp+{dollar}/HCO{dollar}sb3sp-{dollar} cotransporter that transports Na{dollar}sp+{dollar}, HCO{dollar}sb3sp-{dollar}, and net negative charge in the same direction across the cell membrane.; To determine the relation between intracellular pH (pH{dollar}sb{lcub}rm i{rcub}{dollar}) and acid/base transport mechanisms like electrogenic Na{dollar}sp{lcub}+{rcub}{dollar}/HCO{dollar}sb3sp{lcub}-{rcub}{dollar} cotransport, glial cells were studied with double-barreled, pH-sensitive microelectrodes. At a bath pH of 7.5, the mean initial pH{dollar}sb{lcub}rm i{rcub}{dollar} was 7.32 (S.D. 0.03, n = 6) in HEPES-buffered Ringer's solution and 7.39 (S.D. 0.1, n = 6) in HCO{dollar}sb3sp{lcub}-{rcub}{dollar}/CO{dollar}sb2{dollar}-buffered solution. These values for pH{dollar}sb{lcub}rm i{rcub}{dollar} are more than 1 pH unit alkaline to the pH{dollar}sb{lcub}rm i{rcub}{dollar} predicted from a passive distribution of protons; thus, glial cells actively extrude acid. Acid extrusion mechanisms were determined by inducing acidifications from the steady-state pH{dollar}sb{lcub}rm i{rcub}{dollar} and analyzing the ionic dependence and pharmacology of the pH{dollar}sb{lcub}rm i{rcub}{dollar} recovery. Superfusion followed by withdrawal of 15 mM NH{dollar}sb4sp{lcub}+{rcub}{dollar} induced an acidification of 0.1 to 0.3 pH unit after which the pH{dollar}sb{lcub}rm i{rcub}{dollar} recovered toward the original steady-state over the next 7 to 16 min. In the absence of HCO{dollar}sb3sp{lcub}-{rcub}{dollar}/CO{dollar}sb2{dollar}, the recovery from acidification was Na{dollar}sp{lcub}+{rcub}{dollar}-dependent, and amiloride-sensitive. Recovery from acidification was stimulated by adding HCO{dollar}sb3sp{lcub}-{rcub}{dollar}/CO{dollar}sb2{dollar} at constant pH. In HCO{dollar}sb3sp{lcub}-{rcub}{dollar}/CO{dollar}sb2{dollar}-buffered solution, the recovery was Na{dollar}sp{lcub}+{rcub}{dollar}-dependent, SITS-sensitive, and associated with a membrane hyperpolarization. The data strongly suggest that the relatively alkaline pH{dollar}sb{lcub}rm i{rcub}{dollar} of glial cells is due to the activity of at least two mechanisms, Na{dollar}sp{lcub}+{rcub}{dollar}/H{dollar}sp{lcub}+{rcub}{dollar} exchange and electrogenic Na{dollar}sp{lcub}+{rcub}{dollar}/HCO{dollar}sb3sp{lcub}-{rcub}{dollar} cotransport.