Pfltigers Arch (1993) 423:406-410 Et i j hfin Journal of Physiology 9 Springer-Verlag 1993 Co-expression of an anion conductance pathway with Na+-glucose cotransport in rat renal brush-border membrane vesicles C. D. A. Brown, N. King, N. L. Simmons Department of Physiological Sciences, The Medical School, Framlington Place, The University, Newcastle upon Tyne, NE2 4HH, UK Received October 30, 1992/Received after revision December 21, 1992/Accepted January 4, 1993 Abstract. Brush-border membrane vesicles were pre- pared from superficial rat renal cortex by a Mg2+-pre - cipitation technique. The initial (20 s) [14C]glucose up- take rate from solutions containing 100 mmol/1 Na (salt) was found to be dependent upon the anion composition of the medium; in comparison to gluconate-containing medium (0.46 _+ 0.05 nmol/mg protein), C1- accelerated the initial rate to 1.47 _+ 0.21 nmol/mg protein (n = 4 preparations, _+ SEM). This enhancement was reduced by 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB, 0.5 mmol/1), but was unaffected by 4,4'-diisothi- ocyanatostilbene 2,2'-disulphonate (DIDS, 0.5 mmol/1). When membrane vesicles were pre-equilibrated with 100 mmol/1 K (salt) and 100 mmol/1 mannitol and glu- cose uptake was measured from a solution containing 100 mmol/1 Na gluconate and 100 mmol/1 mannitol in the presence of 80 txmol/1 valinomycin (to generate an outward K + diffusion electrical p. d.), it was found that intravesicular KC1 depressed the initial glucose uptake compared to K gluconate. NPPB (0.5 mmol/1) increased the initial glucose uptake with intravesicular KC1 towards values seen in K gluconate vesicles. In con- ditions where the only driving force for glucose uptake was established by an inward anion gradient (Nao = Na~) it was found that inward C1- gradients could drive uphill glucose transport and that this was sensitive to NPPB (0.5 mmol/1), but insensitive to DIDS. We conclude that a C1 conductance co-exists with Na-cotransport in rat renal brush-border membrane vesicles prepared from superficial renal cortex and this may function to regulate the activity of electrogenic transport systems at this membrane. Key words: Brush border - Membrane vesicle - C1 conductance - Cystic fibrosis transmembrane regu- lator - Na-glucose cotransport Correspondence to: C. D. A. Brown Introduction The existence of a conductance pathway for anions in the proximal tubules is the subject of controversy. A C1- conductive pathway sensitive to an imposed K-diffusion potential was identified in brush-border membrane ves- icles (BBMVs) isolated from rabbit renal cortex as early as 1981 [29]. The existence of a diphenylamine-car- boxylate-inhibitable anion conductance has subsequently been confirmed in rabbit BBMVs using a fluorescent indicator technique [8]. Recently Lipkowitz et al. [18] have shown that parathyroid hormone (PTH) may in- crease CI- permeability in rat renal BBMV. Similarly Suzuki et al. [24] have identified an apical CI- channel activated by PTH via the protein kinase A and C path- ways in primary cultures of isolated rabbit proximal tu- bules. The CI- channel was outwardly rectifying pos- sessing a conductance of 33 pS at positive and 22.5 pS at negative potentials. It was sensitive to inhibition by the stilbene 4,4'-diisothiocyanatostilbene 2,2'-disulphon- ate (DIDS, 0.1 mmol/1). In contrast to these positive data, measurements made with intact preparations argue for a minimal C1- conductance pathway, e.g. Cassola et al. [7]. More re- cently Ishibashi et al. [16] have shown that perfusion of low-C1- medium across the luminal surface has a minor effect on intracellular C1- activity when compared to basal-lateral perfusion. The effect from the luminal sur- face was blocked by the stillbene 4-acetamido-4-iso- thiocyanostilbene 2,2-disulphonic acid (SITS) [16]. Recently renal proximal tubule expression of both P- glycoprotein [multi-drug resistance (MDR)] [12, 13, 27] and the cystic fibrosis transmembrane conductance regu- lator (CFTR) [10, 11, 20, 26] have been reported. Since both P-glycoprotein [28] and CFTR function as C1- channels [1, 30] the debate concerning the expression and functional significance of C1- channels in the apical membrane of renal proximal tubule cells has now been reopened. In order to examine the C1 conductance of BBMVs and to overcome the possible contamination of this yes-