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PubMed1982

Mechanisms of p-aminohippurate transport by brush-border and basolateral membrane vesicles isolated from rat kidney cortex.

The uptake of [3H]-labeled p-aminohippurate by brush-border and basolateral membrane vesicles isolated from rat renal cortex has been studied by a rapid filtration technique. Some characteristics of carrier-mediated transport for p-aminohippurate were demonstrated in basolateral membrane vesicles: the uptake was inhibited by probenecid or 4,4'-diisothiocyano-2,2'-disulfonic stilbene (DIDS), was saturable, was stimulated by the countertransport effect, and showed discontinuity in the Arrhenius plot. In contrast, brush-border membrane vesicles failed to display saturability of p-aminohippurate uptake and stimulation by the countertransport effect, although probenecid and DIDS reduced the uptake. Furthermore, p-aminohippurate uptake by brush-border membrane vesicles was influenced more sensitively by alteration in the membrane potential compared with that by basolateral membrane vesicles.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1981

Oxalate secretion in the rat proximal tubule.

Simultaneous capillary and luminal microperfusion studies in the proximal convoluted tubule of the rat were performed to examine the transepithelial secretory flux of [14C]oxalate. Increases in the concentration of oxalate in the capillary solution from 0.096 to 4.3 mM resulted in progressively higher rates of oxalate secretion into the lumen. Further increases in the capillary concentration of oxalate indicated a tendency toward a plateau. The inclusion of para-chloromercuribenzoate, sodium cyanide, indanyloxyacetic acid, furosemide, or para-aminohippurate in the capillary solution significantly lowered the secretory flux of oxalate. the addition of probenecid in a concentration of 10(-4) M inhibited oxalate secretion when the oxalate concentration in the capillary solution ranged between 1.1 and 4.3 mM, but did not affect oxalate secretion at higher capillary concentrations of oxalate. These results indicate that oxalate secretion in the rat proximal tubule is an active carrier-mediated process. When considered in conjunction with prior studies, the present investigations suggest the possibility that more than one oxalate secretory system exists in the rat proximal tubule.

Animals↗
PubMed1983

Nature of Na+-independent stimulation of renal transport of p-aminohippurate by exogenous metabolites.

The relationship between p-aminohippurate (PAH) and Na+ transport in rabbit kidney cortical slices was examined under optimal metabolic conditions. Addition of lactate, pyruvate and acetate to the incubation medium stimulated PAH transport and accumulation, but had no effect on active Na+ efflux from the slices. Conversely, small concentration of F-, in the presence of acetate, decreased PAH transport and accumulation, but had no effect on Na+ efflux. These observations constitute evidence that, in addition to Na+ cotransport [M. I. Sheikh and J. V. Møller Biochem. J. 208, 243 (1982)], PAH uptake is metabolically stimulated by a Na+-independent pathway. This could occur either by a direct metabolic effect on the PAH carrier, or by exchange with intracellular anions that are generated in the presence of exogenous substrate. In support of the latter possibility the PAH carrier is demonstrated to function as an anion exchanger of PAH and fumurate after preloading of the slices with fumurate under anaerobic conditions.

Acetates↗
PubMed1983

Effects of divalent cations and sulfhydryl reagents on the p-aminohippurate (PAH) transporter of renal basal-lateral membranes.

A specific system of transport for p-aminohippurate (PAH) is demonstrated in rabbit renal basal-lateral membrane vesicles. The PAH uptake into an intravesicular space is inhibited by probenecid in concentrations above 0.2 mM. The transport is saturable and is also temperature-dependent with an optimum between 37 and 45 degrees C. Divalent cations are able to enhance the uptake 2- to 3-fold. The stimulatory effect of the divalent cations diminishes in the following order: Mg++ = Mn++, Ba++, Ca++ and Sr++. Maximum stimulation occurs between 2.5 and 5 mM Mg++. The divalent cation stimulatory effect is not the result of changes in the size of the vesicles, in the degree of vesiculation, in the net charge of the membrane or of a transient potential difference across the membrane. Several inhibitors, more inhibitory than probenecid, were found. These are: lithium diiodosalicylate; 4-acetamido-4'-isothiocyano 2,2'-disulfonic acid stilbene; the mercurials, mersalyl acid, p-chloromercuriphenyl sulfonate and Hg++; and 5,5'-dithiobis(nitrobenzoate). Among these, mersalyl acid is the most potent inhibitor for PAH uptake. Its inhibitory effect is probably a combination of its reactivity toward sulfhydryl groups and its anionic character. The results with sulfhydryl reagents indicate that the PAH transport system contains sulfhydryl groups which are essential for the uptake activity. These sulfhydryl groups are probably buried in a hydrophobic region within the lipoprotein matrix of the basal-lateral membrane.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗
PubMed1980

Effect of DIDS on renal tubular transport.

Two stilbene derivates that had been used to covalently label the Cl- carrier in the erythrocyte were investigated for reactivity with the renal organic anion system. These compounds, 4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS) and (4,4'-diisothiocyano)-dihydrostilbene-2,2'-disulfonate (H2DIDS), were found to be potent inhibitors (Ki congruent to 35 microM) of p-aminohippurate (PAH) transport in the renal cortical slice without affecting tetraethylammonium (TEA) transport or tissue viability. During renal clearance studies performed in the perfused kidney, DIDS decreased the PAH/inulin clearance ratio to congruent to 1. When the possible renal transport of [3H]H2DIDS was investigated, the renal slice transport or binding of [3H]H2DIDS reached a slice-to-medium ratio of congruent to 6, and this accumulation was decreased by probenecid. In perfused kidney experiments, the [3H]H2DIDS/inulin clearance ratio was congruent to 0.8. Since probenecid reduced this clearance ratio to congruent to 0.5, there was the possibility that H2DIDS underwent tubular secretion. In conclusion, DIDS and H2DIDS interacted with the renal organic anion transport system, which indicated that these compounds were possible probes for this transport system.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1980

Bath and lumen effects of SITS on PAH transport by isolated perfused renal tubules.

Effects of 4-acetamido-4'-isothiocyano-2,2'-disulfonic stilbene (SITS) in bath or lumen on p-aminohippurate (PAH) transport by isolated perfused snake (Thamnophis spp.) distal-proximal renal tubules were studied. Addition of SITS to the bath in concentrations as low as 10(-10) M irreversibly depressed net PAH secretion. Addition of 10(-4) M SITS in the bath depressed net PAH secretion by about 60% without affecting apparent permeability of luminal membrane to PAH (PL) or net fluid absorption. Cellular PAH concentration was reduced by about 70% at this time but was still greater than that in the bath. Apparent permeability of peritubular membrane, determined from PAH efflux from tubules with oil-filled lumens, was unchanged even with 10(-3) M SITS in bath. These data indicate that SITS in the bath inhibits PAH transport from bath into cells without enhancing passive efflux from cells to bath. Addition of 10(-4) M SITS to the lumen alone depressed net PAH secretion and PL by about 60%, but this depression was reversed when SITS was removed from the lumen. These data suggest that PAH moves from cells to lumen by a mediated process distinctly different from that fro transport into cells from bath.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗