Effects of buffer concentration on voltage-gated H+ currents: does diffusion limit the conductance?
The single-channel proton conductance of the voltage-gated H(+)-selective channel, like that of the F0 component of the H(+)-ATPase, is nearly constant over a wide range of pH encompassing the physiological range. To examine the possible contributions of buffer diffusion and buffer-channel proton transfer reactions to this phenomenon, the effects of buffer concentration on voltage-activated H+ currents were explored in voltage-clamped rat alveolar epithelial cells. Changes in the external buffer concentration ([B]o), evaluated using the whole-cell configuration, had only small effects on H+ currents (IH). Lowering [B]o from 100 to 1 mM did not alter the voltage-activation curve or reversal potential (Vrev) but reduced IH, typically by 10-30%. Changes in internal buffer concentration ([B]i), examined in inside-out patches, usually altered IH more distinctly and subtly changed the kinetics. Overall, the effects of changing buffer concentration were small and subtle. The maximum attenuation of the single-channel H+ current at 1 mM buffer was estimated to be approximately 20% at either mouth of the H+ channel. Therefore, the rate-determining step in H+ permeation is neither deprotonation of buffer at the inner mouth of the channel nor protonation of buffer at the external surface. Evidently the rate of H+ permeation through the channel is itself small enough that diffusion of buffer in bulk solution does not directly limit the conductance significantly.
DeCoursey
T E
TE
Department of Molecular Biophysics and Physiology, Rush Presbyterian St. Luke's Medical Center, Chicago, illinois 60612, USA. tdecours@rpslmc.edu
Cherny
V V
VV
D000818:Animals; D055592:Biophysical Phenomena; D001703:Biophysics; D002021:Buffers; D004058:Diffusion; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D015640:Ion Channel Gating; D007473:Ion Channels / Q000378:metabolism*; D008297:Male; D008564:Membrane Potentials; D018408:Patch-Clamp Techniques; D011522:Protons; D011650:Pulmonary Alveoli / Q000166:cytology / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
D002021:Buffers; D007473:Ion Channels; D011522:Protons