Uptake of bromosulfophthalein via SO2-4/OH− exchange increases the K+ conductance of rat hepatocytes

In confluent primary cultures of rat hepatocytes, micromolar concentrations of bromosulfophthalein (BSP) lead to a sizeable hyperpolarization of membrane voltage. The effect is a saturable function of BSP concentration yielding an apparent value of 226 μmol/l and a V max of −10.3 mV. The BSP-induced membrane hyperpolarization is inhibited by the K+ channel blocker Ba2+, and in cable-analysis and ion-substitution experiments it becomes evident that the effect is due to a significant increase in cell membrane K+ conductance. Voltage changes were attenuated by the simultaneous administration of [Formula: see text], succinate, and cholate ( cis-inhibition) and increased after preincubation with [Formula: see text] and succinate ( trans-stimulation), suggesting that the effect occurs via BSP uptake through the known[Formula: see text]/OH− exchanger. Microfluorometric measurements reveal that BSP-induced activation of K+ conductance is not mediated by changes in cell pH, cell Ca2+, or cell volume. It is concluded that K+channel activation by BSP (as well as by DIDS and indocyanine green) may reflect a physiological mechanism linking the sinusoidal uptake of certain anions to their electrogenic canalicular secretion.

Expression of sodium-independent organic anion uptake systems of skate liver in Xenopus laevis oocytes

The expression of the basolateral sodium-independent organic anion uptake system of the little skate (Raja erinacea) has been studied in Xenopus laevis oocytes. Injection of oocytes with skate liver poly(A)+ RNA resulted in the functional expression of chloride-dependent sulfobromophthalein (BSP) uptake and sodium-independent taurocholate uptake within 3-5 days. The expressed chloride-dependent BSP uptake activity exhibited saturation kinetics [apparent Michaelis constant (Km) 1.8 microM] and efficiently extracted BSP from its binding sites on bovine serum albumin. The chloride-sensitive portion of BSP uptake was inhibited by bilirubin (10 microM; 27%), 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (100 microM; 57%), bumetanide (100 microM; 48%), taurocholate (200 microM; 51%), and cholate (200 microM; 45%). Size fractionation of total skate liver mRNA revealed that a 1.8- to 2.9-kb size class mRNA was sufficient to express chloride-dependent BSP uptake and sodium-independent taurocholate uptake. In addition, a 1- to 1.7-kb size class mRNA expressed sodium-independent taurocholate uptake but had no effects on BSP uptake. This study confirms that an organic anion transport system for chloride-dependent BSP uptake, with characteristics similar to rat liver, is already expressed in the liver of lower vertebrates and thus represents a phylogenetically old system. Sodium-independent taurocholate uptake in skate liver may be mediated by two different transport systems.

Sex differences in the hepatic uptake of sulphobromophthalein in the rat

1. Sex difference in the hepatic uptake of sulphobromophthalein (BSP) was investigated in male and female rats in three different experimental models. 2. In the intact animal the BSP plasma disappearance rate was significantly higher (P < 0.01) in females than in males when 0.15 or 1.5 μmol/kg body wt. was injected. Comparable values were found at the highest BSP dose (15 μmol/kg body wt.) used. 3. In the perfused liver, the first-pass hepatic extraction and the uptake velocity were significantly higher (P < 0.001) in female rats at low BSP doses (0.3–750 μmol/g of liver) whereas identical values were found at higher concentrations. 4. In hepatocytes isolated by collagenase perfusion, the BSP uptake occurs via two different uptake sites in both sexes. The Km of the high affinity sites was lower in females than in males (3.67 ± 0.58 vs 7.24±0.68 Mmol/l, P < 0.001) whereas Vmax. showed comparable values (2.70 ± 0.36 vs 2.47 ± 0.45 nmol of BSP/mg of protein, NS). In contrast, no difference was found in the kinetic parameters of the low affinity sites (Km 50.6±31.1 vs 61.0±17.5 μmol/l; Vmax. 21.9 ± 13.2 vs 25.0±3.6 nmol of BSP/mg of protein, mean ± sd, NS, females and males respectively). 5. Taken together these data show that low doses of BSP are taken up by the liver more efficiently in female than in male rats and are consistent with a sex-related difference in the affinity but not in the number of the BSP high affinity uptake sites.

The Kinetics of Organic Anion Excretion by the Liver in Acute Intermittent Porphyria

1. The kinetics of disappearance from the blood of three organic anions were studied in patients with acute intermittent porphyria. Indocyanine green (ICG) and [14C]bilirubin were cleared normally in this disease, whereas bromsulphthalein (BSP) was cleared at less than the normal rate. The first exponential component of the BSP clearance curve was normal in acute intermittent porphyria, but the second exponential component declined less rapidly than in normals. The abnormality was greater in symptomatic than in asymptomatic patients, but both groups were significantly different from normals. 2. The data are discussed in terms of a previously published model of BSP metabolism. The findings suggest normal BSP uptake initially by the liver with increased regurgitation of dye into the blood and decreased excretion of BSP from the liver cell into the bile. 3. In contrast to human acute intermittent porphyria, plasma clearance of BSP was greatly increased in experimental porphyria. This was associated with an increased level of BSP conjugating enzyme in the liver in experimental porphyria. 4. The BSP retention in acute intermittent porphyria, along with certain other findings, raises the question of the existence of an ‘oestrogen effect’ in this disease, but further studies will be required to determine whether the mechanism of BSP retention is identical to that produced by oestrogen.