UPTAKE OF SULFATE CONJUGATES BY ISOLATED RAT HEPATOCYTES

The uptake of estrone sulfate (E(1)S; 1 to 400 mu M), harmol sulfate ( HS; 5 to 900 mu M), and 4-methylumbelliferyl sulfate (4MUS; 5 to 1000 mu M) was investigated in isolated rat hepatocytes in the presence or absence of inhibitors. Uptake of all of the sulfate conjugates was rap id and exhibited saturation kinetics, best characterized by saturable and nonsaturable (linear transmembrane clearance) transport systems. T he K-M's were: 16 +/- 6, 123 +/- 28, and 64 +/- 6 mu M for E(1)S, HS, and 4MUS, respectively, with corresponding V-max's of 0.85 +/- 0.56, 0 .48 +/- 14, and 0.42 +/- 0.07 nmol/min/10(6) cells. The nonsaturable u ptake clearances, which displayed concentration-independent uptake, we re 3 +/- 2, 1 +/- 0.1, 0.5 +/- 0.1 mu l/min/10(6) cells, respectively. Uptake of E(1)S was inhibited by ouabain (1 mM) and replacement of so dium by choline, whereas HS was insensitive to the addition or substit ution. Uptake of both E(1)S and HS was significantly reduced by metabo lic inhibitors (antimycin A, 2.7 mu M, rotenone, 30 mu M, and KCN, 2 m M) and temperature reduction (from 37 to 27 degrees C). 4,4'-Diisothio cyanostilbene-2-2'-disulfonic acid (2 mM), an inhibitor of anion trans port, reduced E(1)S and HS uptake; E(1)S uptake was also reduced by HS . HS uptake by both saturable and nonsaturable transport components wa s depressed by 4MUS (300 mu M); the apparent K-M was increased by 83% while the V-max remained unaltered, and the nonsaturable component was decreased by 48%. The data strongly suggest that multiple pathways ex ist for the uptake of E(1)S, HS, and 4MUS. E(1)S uptake is sodium-depe ndent, requires energy, and is inhibited by anions such as 4,4'-diisot hiocyanostilbene-2-2'-disulfonic acid and other sulfate conjugates. HS uptake, while being energy dependent, is not sodium dependent, and is inhibited by 4MUS in a competitive fashion. At least one of these pat hways is shared.