The purpose of these studies was to assess the intracellular signals which regulate anion channel opening during hepatocyte swelling. In rat HTC hepatoma cells and primary rat hepatocytes, anion currents (whole-cell patch clamp), cell volume (Coulter multisizer), and cell fraction protein kinase C (PKC) (Western blot) were measured. In HTC cells, we found that swelling during incubation in hypotonic buffer (20-40% less NaCl) was followed by: i) activation of Cl--selective currents (ICl-Swell) from < -4 pA/pF (-80 mV) to > 45 pA/pF, ii) regulatory volume decrease, and iii) translocation of PKCα from cytosol to membrane within 1 min. ICl-Swell was abolished by PKC downregulation with phorbol 12-myristate 13-acetate (100 nM,18 hr), chelation of intracellular Ca2+ with EGTA (2mM), or depletion of cytosolic ATP with apyrase (1U/ml). Moreover, PKC inhibition by chelerythrine (25μM) and phloretin (2.5μM) prevented activation of both ICl-Swell and volume recovery. In HTC cells and primary hepatocytes, swelling during intracellular hypertonic perfusion (50mM sucrose) activated similar Cl- currents (45.77±5.4 and 35.8±7.8 pA/pF respectively); these currents were also inhibited by chelerythrine (25μM). In conclusion, PKC represents an important component of the signaling pathways that couple hepatocyte volume to changes in membrane ion permeability.
|Original language||English (US)|
|State||Published - 1997|
ASJC Scopus subject areas
- Molecular Biology