Open AccessEvidence that catecholamine transport into chromaffin vesicles is coupled to vesicle membrane potential
Author(s) -
Ronald W. Holz
Publication year - 1978
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.75.10.5190
Subject(s) - vesicle , chemistry , electrochemical gradient , membrane potential , valinomycin , membrane , biophysics , methylamine , thiocyanate , atpase , catecholamine , biochemistry , membrane transport , biology , enzyme , endocrinology
The effects of ATP, Mg2+ , and various agents on pH gradient, membrane potential, and catecholamine transport across membranes of intact bovine chromaffin vesicles were investigated. Methylamine and thiocyanate (SCN- ) distributions across the vesicle membrane were used to estimate the H+ concentration gradient and membrane potential, respectively. The H+ concentration ratio (intravesiculanmedium) equals 16 when the medium pH is 6.9 and is unaltered by ATP and Mg2+ . In the absence of ATP and Mg2+ , the steady-state intravesicular S14 CN- concentration is lower than the medium concentration. ATP and Mg2+ cause an increased influx and a decreased efflux of SCN- that results in SCN- being concentrated in the vesicles 6- to 8-fold over the medium. The findings are consistent with an ATP,Mg2+ -induced potential of approximately 50 mV (intravesicular side positive). Carbonyl cyanidep -trifluoromethoxyphenylhydrazone (FCCP), a H+ translocater, andN -ethylmaleimide (NEM), a sulfhydryl reagent, decrease the SCN- ratio and, thus, the membrane potential in the presence of ATP and Mg2+ . They have no effect on the H+ concentration gradient. The rate of catecholamine uptake into vesicles is increased 4- to 6-fold by ATP and Mg2+ . The ATP,Mg2+ -stimulated uptake is inhibited by FCCP and NEM over the same concentration ranges that reduce the SCN- distribution (membrane potential). FCCP increases and NEM decreases vesicular membrane ATPase activity. Thus, catecholamine uptake is correlated to an inside-positive membrane potential, and not to ATPase activity. If catecholamine uptake is coupled to membrane potential, then a charged species must be involved in the transport mechanism. Reserpine and rotenone inhibit catecholamine influx but have no effect on the H+ electrochemical gradient; they probably act at a step before coupling to the membrane potential (or the H+ electrochemical gradient). Atractyloside, an inhibitor of nucleotide transport, has no effects on catecholamine transport or the H+ electrochemical gradient.