Abstract
Chronic ethanol administration alters the process of receptor-mediated endocytosis in isolated rat hepatocytes. Using the asialoglycoprotein receptor (ASGP-R) as a model, we have previously shown decreased binding of asialoglycoproteins to this receptor after as early as 1 week of ethanol administration. In the present study, we further analyzed the mechanism(s) responsible for this impairment by determining the ligand and antibody binding characteristics of the ASGP-R in rats fed ethanol over a 5-week time course. The results presented here demonstrate that ethanol treatment for 4 days significantly impaired total ligand binding without affecting antibody binding. Ethanol administration for a longer period of 1-2 weeks resulted in intermediate impairments in both ligand and antibody binding. After 5 weeks of ethanol exposure, ligand and antibody binding were equally lowered. In contrast to total cellular receptor binding, surface binding of both ligand and antibody were decreased over the entire time course of ethanol administration. Our data indicate that the ASGP-R is initially inactivated during the time course of ethanol exposure and that a redistribution of surface receptors to intracellular compartments occurs. Northern blot analysis showed that there was a significant decrease in receptor mRNA content in the 5-week chronically fed animals but not in the animals fed for 1 week. In addition, after 5 weeks of ethanol feeding, biosynthetic labeling of the ASGP-R was decreased in the ethanol cells, indicating impaired synthesis of the ASGP-R. In summary, an early inactivation of the ASGP-R occurs during ethanol exposure followed by an actual decrease in protein and mRNA content for the receptor.
Highlights
Chronic ethanol administration alters the process of receptor-mediated endocytosis in isolated rat hepatocytes
Hepatocytes were treated with low levels of digitonin (0.055% final concentration) to permeabilize cells and make intracellular receptors available for ligand and antibody binding
Our results show that after 4 days of feeding, binding of ASOR to permeabilized cells was decreased significantly (22%) in ethanol-fed animals when compared to controls (Fig. 1A)
Summary
Chronic ethanol administration alters the process of receptor-mediated endocytosis in isolated rat hepatocytes. The purpose of the present study was to clarify whether the decreased ligand binding we reported after ethanol administration was attributed to an actual reduction in the number of receptors (decreased receptor content) or to an inability of the receptors to recognize ligand due to receptor inactivation. For these experiments, ligand (ASOR) and antirat ASGP-R antibody binding were performed using both intact and permeabilized hepatocytes isolated from animals fed alcohol over a time course. Ethanol-impaired Asialoglycoprotein Receptor Function messenger RNA levels after 1 and 5 weeks of feeding and in vitro biosynthesis of the ASGP-R was determined in these cells
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