Electrophysiological evidence for Na+-coupled bicarbonate transport in cultured rat hepatocytes.
Recent observations suggest that hepatocytes exhibit basolateral electrogenic Na+-coupled HCO3- transport. In these studies, we have further investigated this transport mechanism in primary culture of rat hepatocytes using intracellular microelectrodes to measure membrane potential difference (PD) and the pH-sensitive fluorochrome 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein to measure intracellular pH (pHi). In balanced media containing 25 mM HCO3-, PD averaged -32.1 +/- 0.6 (SE) mV and pHi averaged 7.22 +/- 0.03. PD became more negative (hyperpolarized) when extracellular [HCO3-] was increased and less negative (depolarized) when extracellular HCO3- was decreased. Acute replacement of extracellular Na+ by choline also resulted in membrane depolarization of 18.0 +/- 1.6 mV, suggesting net transfer of negative charge. This decrease in PD upon Na+ removal was HCO3- -dependent, amiloride insensitive, and inhibited by the disulfonic stilbene 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (SITS). PD also decreased upon acute exposure to SITS. The degree of depolarization seen with removal of Na+ or HCO3- correlated directly with resting PD (r = 0.81 and 0.95, respectively), suggesting a voltage-dependent mechanism. Removal of extracellular Na+ also decreased pHi to 7.06 +/- 0.02, and this acidification was decreased in the absence of HCO3- or in the presence of SITS or amiloride. These studies provide direct evidence for electrogenic Na+-coupled HCO3- transport in rat hepatocytes. Further, they suggest that it represents a major pathway for conductive movement of Na+ across the membrane and that it contributes, along with Na+-H+ exchange, to the intracellular acidification observed upon removal of extracellular Na+.(ABSTRACT TRUNCATED AT 250 WORDS)
Fitz
J G
JG
Department of Medicine, University of California School of Medicine, San Francisco 94143.
Persico
M
M
Scharschmidt
B F
BF
D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000494:pharmacology; D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D001639:Bicarbonates / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008099:Liver / Q000502:physiology*; D008297:Male; D008564:Membrane Potentials; D008839:Microelectrodes; D051381:Rats; D011919:Rats, Inbred Strains; D027982:Sodium-Bicarbonate Symporters
D001639:Bicarbonates; D002352:Carrier Proteins; D027982:Sodium-Bicarbonate Symporters; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D000584:Amiloride