Fall in intracellular pH and increase in resting tension induced by a mitochondrial uncoupling agent in crayfish muscle.

Abstract

1. The influence of the mitochondrial uncoupling agent carbonylcyanide-m-chlorophenylhydrazone (CCCP) upon resting tension and intracellular pH (pHi) was studied in the dactyl opener muscle of the crayfish. pHi was measured with liquid sensor H+-selective microelectrodes. 2. CCCP (10(-6)-10(-5) mol l-1) induced a reversible, tonic contracture which was associated with a depolarization of the membrane potential. Both effects were augmented by a fall and inhibited by a rise in extracellular pH. The action of CCCP on tension was not mimicked by cyanide + oligomycin or by cyanide + dicyclohexylcarbodiimide nor was it inhibited by pre-exposure to these agents. 3. CCCP produced an initial alkalosis of less than 0.1 units and thereafter a fall in pHi of 0.4-0.6 units during which the sarcolemmal H+ driving force decreased from 61 to 15 mV. The apparent influx of H+ due to CCCP had a maximum of 2.7 mequiv l-1 min-1. The CCCP-induced acidosis was unaffected by iodacetate (0.5 mmol l-1) but it was inhibited by a depolarization of the membrane potential. 4. The contraction caused by CCCP was not due to the simultaneous fall in pHi since an intracellular acidosis of equal magnitude, produced by propionate (50 mmol l-1), did not lead to force generation. In addition, propionate had an inhibitory effect on the depolarization and contracture caused by CCCP. 5. Both the depolarization and the contracture caused by CCCP were inhibited by gamma-aminobutyric acid (GABA). The contracture was blocked by Cd2+, Mn2+ and by a nominally Ca2+ -free medium but not by a pre-exposure to caffeine (20 mmol l-1). Cd2+ and Mn2+ had no influence on the fall of pHi caused by CCCP. 6. It is concluded that CCCP induces a sarcolemmal H+ conductance which leads to a fall in pHi and to a depolarization of the membrane potential. This depolarization activates sarcolemmal, voltage-dependent calcium channels and thereby induces an increase in tension. The initial alkalosis produced by CCCP may be due to a transient uptake of H+ by mitochondria.