OCTANOL, A GAP JUNCTION UNCOUPLING AGENT, CHANGES INTRACELLULAR [H+] IN RAT ASTROCYTES

Octanol rapidly closes gap junction channels but its mechanism of acti on is not known. Because intracellular [H+], pH(i), also affects the c onductance of gap junctions, we studied octanol's effects on pH(i) in cultured rat astrocytes, which are highly coupled cells. Octanol (1 mM ) caused an acid shift in the pH(i) of 90% of rat hippocampal astrocyt es which averaged -0.19 +/- 0.09 pH units in magnitude. In 58% of the cells tested, a biphasic change in pH(i) was seen; octanol produced an initial acidification lasting similar to 10 min that was followed by a persistent alkalinization. The related gap junction uncoupling agent , heptanol, had similar effects on pH(i). Octanol-induced changes in p H(i) were similar in nominally HCO3--free and HCO3--containing solutio ns, although the rate of initial acidification was significantly great er in the presence of HCO3-. The initial acidification was inhibited i n the presence of the stilbene DIDS, an inhibitor of Na+/HCO3- cotrans port, indicating that octanol caused acidification by blocking this po werful acid extruder. The alkalinization was inhibited by amiloride wh ich blocks the Na+/H+ exchanger (NHE), an acid extruder, suggesting th at the alkaline shift induced by octanol was caused by stimulation of NHE. As expected, octanol's effects on astrocytic pH(i) were prevented by removal of external Na+, which blocks both Na+/HCO3- cotransport a nd NHE. Octanol had only small effects on intracellular Ca2+ (Ca-i(2+) ) in astrocytes. Hepatocytes which, like astrocytes, are strongly coup led to one another, showed no change in pH(i) with octanol application . Fluorescence recovery after photobleaching (FRAP) was used to study the effect of changes in astrocyte pH(i) on degree of coupling in hipp ocampal astrocytes. Coupling was decreased by intracellular acid shift s similar to-0.2 pH units in size. Octanol's effects on astrocyte pH(i ) were complex but a prompt initial acidification was nearly always se en and could contribute to the uncoupling action of this drug in astro cytes. Because octanol uncouples hepatocytes without changing their pH (i), this compound clearly can influence gap junctional conductance in dependent of changes in pH(i). (C) 1996 Wiley-Liss, Inc.