Protonmotive force and catecholamine transport in isolated chromaffin granules

The effect of the transmembrane potential (delta psi) and the proton concentration gradient (delta pH) across the chromaffin granule membrane upon the rate and extent of catecholamine accumulation was studied in isolated bovine chromaffin granules. Freshly isolated chromaffin granules had an intragranular pH of 5.5 as measured by [14C]methylamine distribution. The addition of ATP to a suspension of granules resulted in the generation of a membrane potential, positive inside, as measured by [14C]thiocyanate (SCN-) distribution. The addition of carboxyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP), a proton translocator, resulted in a reversal of the potential to negative values (measured by [3H]tetramethylphenylphosphonium (TPMP+)) approaching -90 mV. Changing the external pH of a granular suspension incubated with FCCP produced a linear perturbation in the measured potential from positive to negative values, which can be explained by the distribution of protons according to their electrochemical gradient. When ammonia (1 to 50 mM) was added to highly buffered suspensions of chromaffin granules there was a dose-dependent decrease in the transmembrane proton gradient (delta pH) and an increase in the membrane potential (delta...Continue Reading