Bicarbonate-dependent and -independent intracellular pH regulatory mechanisms in rat hepatocytes. Evidence for Na+-HCO3- cotransport.

Abstract

Using the pH-sensitive dye 2,7-bis(carboxyethyl)-5(6)-carboxy-fluorescein and a continuously perfused subconfluent hepatocyte monolayer cell culture system, we studied rat hepatocyte intracellular pH (pH,) regulation in the presence (+HCO3) and absence (-HCO3) of bicarbonate. Baseline pH, was higher (7.28±09) in +HCO3 than in -HCO3 (7.16±0.14). Blocking Na+/H' exchange with amiloride had no effect on pH1 in +HCO3 but caused reversible 0.1-0.2-U acidification in -HCO3 or in +HCO3 after preincubation in the anion transport inhibitor 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene (DIDS). Acute Na' replacement in +HCO3 also caused acidification which was amiloride independent but DIDS inhibitible.The recovery of pH, from an intracellular acid load (maximum H+ efilux rate) was 50% higher in +HCO3 than in -HCO3. Amiloride inhibited H+ efflux..M by 75% in -HCOj but by only 27% in +HCO3. The amiloride-independent pH, recovery in +HCO3 was inhibited 50-63% by DIDS and 79% by Na+ replacement but was unaffected by depletion of intracellular Cl-, suggesting that Cl-/HCO3 exchange is not involved. Depolarization of hepatocytes (raising external K+ from 5 to 25 mM) caused reversible 0.05-0.1-U alkalinization, which, however, was neither Na+ nor HCO3 dependent, nor DIDS inhibitible, findings consistent with electroneutral HCO3 transport.We conclude that Na'-HCO3 cotransport, in addition to Na+/H+ exchange, is an important regulator of pH1 in rat hepatocytes.

Sodium, Hydrogen exchange type 1 and bile ductular secretory activity in the guinea pig

“…12 Interestingly, the activity of Na ϩ , H ϩ exchanger at low pH i in biliary epithelial cells isolated from bile duct-ligated rats was double than in hepatocytes, but antiporter activity became zero at similar pH i values in both cell types. [13][14][15] This is compatible with the presence of a higher number, but similar pH i dependence, of the antiporter in BEC as compared with hepatocytes. And indeed, steadystate levels of Na ϩ , H ϩ exchange type I (NHE1) mRNA were several-fold higher in BEC than in hepatocytes 15 ; however, the role of Na ϩ , H ϩ exchange in biliary secretory function is not yet clear.…”

Acid‐base transport systems in a polarized human intestinal cell monolayer: Caco‐2

“…A higher concentration of DIDS was required than is usually used to inhibit HCO3-transport systems. However, some cells, such as hepatocytes (Gleeson, Smith & Boyer, 1989) are relatively resistant to DIDS. Furthermore, cells were pre-incubated rather than continuously perfused with DIDS and its inhibitory effects may be partly reversible.…”

Effect of glucagon on intracellular pH regulation in isolated rat hepatocyte couplets.

“…The ion transport processes underlying basal or hormonal-stimulated bicarbonate secretion in the hepatocyte are currently under investigation. Recent progress has been made in studies exploring intracellular pH (pHi) regulation in isolated hepatocytes (6)(7)(8). These studies identify the presence of an electroneutral Na+ -independent C1-/ HCO3-exchanger (7,8) which functions as a counterpoint to the acid extruding systems (NaI/H' exchanger and Na'-HCO-symport) (6) and which has been detected recently in the hepatocyte apical domain of human liver (9).…”

“…Recent progress has been made in studies exploring intracellular pH (pHi) regulation in isolated hepatocytes (6)(7)(8). These studies identify the presence of an electroneutral Na+ -independent C1-/ HCO3-exchanger (7,8) which functions as a counterpoint to the acid extruding systems (NaI/H' exchanger and Na'-HCO-symport) (6) and which has been detected recently in the hepatocyte apical domain of human liver (9). The Cl-/ HCO3-exchanger is present in all the bicarbonate-secreting epithelia (10)(11)(12)(13)(14)(15)(16)(17)(18)(19).…”