Determination of Δψ, ΔpH and the proton electrochemical gradient in isolated cholinergic synaptic vesicles

The electrical potential (Δψ) of intact cholinergic synaptic vesicles was measured in the presence and absence of the proton translocator carbonyl cyanide p-trifluoromethoxy-phenylhydrazone (FCCP), and the results were utilized to calculate the vesicular proton chemical gradient (ΔpH) and proton electrochemical potential (Δμ_H+). At external pH = 7.4 the vesicles maintain a proton electrochemical gradient of {reversed tilde equals}+20 mV (positive inside) which is composed of Δψ{reversed tilde equals}-80 mV (negative inside) and ΔpH{reversed tilde equals}1.6 (acidic inside). The proton chemical gradient (ΔpH) increases as a function of pH_out whereas the vesicular electrical potential (Δψ) is only slightly affected by the external pH. Consequently, Δμ_H+ is larger at basic external pH values ({reversed tilde equals}+40 mV at pH_out = 9.0) and smaller at acidic external pH values (Δμ_H+{reversed tilde equals}0 at (pH_out = 5.6). The possible physiological role of the electrochemical potentials in maintaining high concentrations of acetylcholine within the cholinergic synaptic vesicle is discussed.