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Keywords:

  • Ca2+ Uptake;
  • IP 3 ;
  • Liver;
  • Microsomes;
  • Glucose 6-Phosphatase;
  • Chloromethiazole;
  • 4-Methyl-Pyrazole;
  • EtOH

Background

Hydrolysis of glucose 6-phosphate (G6P) via glucose 6-phosphatase (G6Pase) enlarges the reticular Ca2+ pool of the hepatocyte. Exposure of liver cells to ethanol (EtOH) impairs reticular Ca2+ homeostasis. The present study investigated the effect of acute EtOH administration on G6P-supported Ca2+ accumulation in liver cells.

Methods

Total microsomes were isolated from rat livers acutely perfused with varying doses of EtOH (0.01, 0.1, or 1% v/v) for 8 minutes. Calcium uptake was assessed by 45Ca redistribution. Inorganic phosphate (Pi) formation was measured as an indicator of G6Pase hydrolytic activity.

Results

G6P-supported Ca2+ uptake decreased in a manner directly proportional to the dose of EtOH infused in the liver, whereas Ca2+ uptake via SERCA pumps was decreased by ~25% only at the highest dose of alcohol administered. The reduced accumulation of Ca2+ within the microsomes resulted in a smaller inositol 1,4,5-trisphosphate (IP3)-induced Ca2+ release. Kinetic assessment of IP3 and passive Ca2+ release indicated a faster mobilization in microsomes from EtOH-treated livers, suggesting alcohol-induced alteration of Ca2+ releasing mechanisms. Pretreatment of livers with chloromethiazole (CMZ) or dithiothreitol (DTT), but not 4-methyl-pyrazole prevented the inhibitory effect of EtOH on G6Pase activity and Ca2+ homeostasis.

Conclusions

Liver G6Pase activity and IP3-mediated Ca2+ release are rapidly inhibited following acute (8 minutes) exposure to EtOH, thus compromising the ability of the endoplasmic reticulum to dynamically modulate Ca2+ homeostasis in the hepatocyte. The protective effect of CMZ and DTT suggests that the inhibitory effect of EtOH is mediated through its metabolism via reticular cyP4502E1 and consequent free radicals formation.