Acute and chronic changes in cholesterol modulate Na-P_i cotransport activity in OK cells

We previously showed an inverse correlation between membrane cholesterol content and Na-P_i cotransport activity during the aging process and adaptation to alterations in dietary P_i in the rat (Levi M, Jameson DM, and van der Meer BW. Am J Physiol Renal Fluid Electrolyte Physiol 256: F85-F94, 1989). The purpose of the present study was to determine whether alterations in cholesterol content per se modulate Na-P_i cotransport activity and apical membrane Na-P_i protein expression in opossum kidney (OK) cells. Acute cholesterol depletion achieved with β-methyl cyclodextrin (β-MCD) resulted in a significant increase in Na-P_i cotransport activity accompanied by a moderate increase in apical membrane Na-P_i protein abundance and no alteration of total cellular Na-P _i protein abundance. Conversely, acute cholesterol enrichment achieved with β-MCD/cholesterol resulted in a significant decrease in Na-P_i cotransport activity with a moderate decrease in apical membrane Na-Pi protein abundance and no change of the total cellular Na-P _i protein abundance. In contrast, chronic cholesterol depletion, achieved by growing cells in lipoprotein-deficient serum (LPDS), resulted in parallel and significant increases in Na-P_i cotransport activity and apical membrane and total cellular Na-P_i protein abundance. Cholesterol depletion also resulted in a significant increase in membrane lipid fluidity and alterations in lipid microdomains as determined by laurdan fluorescence spectroscopy and imaging. Chronic cholesterol enrichment, achieved by growing cells in LPDS followed by loading with low-density lipoprotein, resulted in parallel and significant decreases in Na-P_i cotransport activity and apical membrane and total cellular Na-P_i protein abundance. Our results indicate that in OK cells acute and chronic alterations in cholesterol content per se modulate Na-P_i cotransport activity by diverse mechanisms that also include significant interactions of Na-P _i protein with lipid microdomains.