High and low affinity binding of [3H]cholate to rat liver plasma membranes.

The transport of bile acids across sinusoidal and canalicular membranes of hepatocytes is characterized as carrier mediated. Such a carrier should specifically bind bile acids at physiological concentrations. We examined the binding of [3H]cholate to rat liver plasma membranes using a microcentrifugation technique and detected high (KD = 1.23 +/- 0.44 microM, Bmax = 21.8 +/- 3.3 pmol/mg protein) and low (KD = 1.97 +/- 1.33 mM, Bmax = 41.5 +/- 25.3 nmol/mg protein) affinity binding sites. Maximal binding was achieved within 15-45 sec and was stable for 2 min at 37 degrees. Binding to the high affinity site was reversible, was not Na+ dependent or attributable to vesicular uptake, and exhibited a broad pH optimum. Binding to this site was negligible or not detected in liver mitochondrial and microsomal fractions, was saturable, and was inhibited by other bile acids. The IC50 values for bile acids as inhibitors of [3H]cholate binding at the high affinity site were: taurocholate, 1.9 nM; glycodeoxycholate, 3.1 nM; chenodeoxycholate, 5.6 nM; taurochenodeoxycholate, 7.3 nM; glycochenodeoxycholate, 11 nM; lithocholate, 13 nM; taurodeoxycholate, 20 nM; glycocholate, 3.6 microM; and deoxycholate, 5.6 microM. [3H]Cholate specific binding was inhibited by 10(-5) M bromosulfophthalein, bilirubin and indocyanin green. These data support the hypothesis that the high affinity binding site represents a carrier which is shared by bile acids and nonbile acid organic anions.