Amiloride blockable sodium fluxes in toad bladder membrane vesicles.

摘要: Recently we reported a simple manual assay for the measurements of isotope fluxes through channels in heterogenous vesicle populations (Garty et al.,J. Biol. Chem.258:13094–13099 (1983)). The present paper describes application this method to assessment amiloride blockable toad bladder microsomes. When22Na+ uptake was monitored presence an opposing Na+ gradient, relatively large and transient amiloride-sensitive flux observed. Such amiloride-blockable could also be induced by KCl+valinomycin diffusion potential. effects intra- extravesicular ionic composition on rate of22Na+ were examined. It shown that occur particles permeable Li+ but impermeable K+, Tris+ Cl−. Analysis dose-response relations revealed complex “non Michaelis-Menten” behavior. data accounted assuming either strong negative cooperativity amiloride-membrane interaction, or two conducting pathways withKi values 0.06 6.4 μm. Both appear electrogenic therefore possibility electroneutral Na/H exchange excluded. Calcium ions block from inner not outer phase membrane. is suggested although substantial part the22Na+ inhibited only high concentration amiloride, represents transport apical Na-specific channels. define optimal experimental conditions study