Effects of osmotic stresses on isolated rat hepatocytes. II. Modulation of intracellular pH.
To assess the roles of acid-base transport systems in cell volume regulation in rat hepatocytes, intracellular pH (pHi) was measured in subconfluent monolayers loaded with 2'-7'-bis(carboxyethyl)-5,6-carboxyfluorescein (BCECF) after exposure to hypotonic and relative hypertonic media, interventions that stimulate regulatory volume decrease (RVD) and increase (RVI), respectively. During RVD, pHi decreased from 6.98 +/- 0.11 to 6.85 +/- 0.08 in the absence of HCO3- and from 7.26 +/- 0.10 to 7.19 +/- 0.06 in its presence. Omission of Na+ or addition of 1 mM amiloride prevented the decline in pHi. Acute withdrawal or replacement of Na+ in hypotonic medium resulted in a slower rate of fall or recovery in pHi, respectively, than when the same maneuvers were carried out in isotonic medium. In contrast, during RVI, pHi increased from 6.86 +/- 0.11 to 7.15 +/- 0.15 in the absence of HCO3-, a rise in pHi that was also completely abolished by Na+ removal or by 1 mM amiloride. In the presence of HCO3-, the rise in pHi was less marked than in its absence, although net acid efflux was greater because of a greater intracellular buffering capacity. Cl- removal in the presence of HCO3- had no effect on the change in pHi during either RVD or RVI. Perfusion with 0.5 mM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) during RVD lowered pHi further and accentuated the subsequent pHi rise seen after the return to isotonic medium. These data suggest that Na(+)-H+ exchange in rat hepatocytes is downregulated during RVD and activated during RVI. Cl(-)-HCO3- exchange does not appear to be involved in hepatocyte volume regulation.
Gleeson
D
D
Department of Medicine and Liver Center, Yale University School of Medicine, New Haven, Connecticut 06510.
Corasanti
J G
JG
Boyer
J L
JL
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001639:Bicarbonates / Q000494:pharmacology; D002021:Buffers; D002478:Cells, Cultured; D002794:Choline / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D006982:Hypertonic Solutions; D007038:Hypotonic Solutions; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D009994:Osmolar Concentration; D051381:Rats; D011919:Rats, Inbred Strains; D012964:Sodium / Q000494:pharmacology
D001639:Bicarbonates; D002021:Buffers; D006982:Hypertonic Solutions; D007038:Hypotonic Solutions; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D012964:Sodium; D002794:Choline; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid