RHEOGENIC CL- CONDUCTANCE AND CL(-)-CL(-)-EXCHANGE ACTIVITIES IN GUINEA-PIG JEJUNAL BASOLATERAL MEMBRANE-VESICLES

We describe a method for the simultaneous purification of apical (brus h-border membrane) and basolateral membrane (BLM) vesicles from the sa me sample of guinea pig jejunum. We applied functional tests to demons trate the absence of reciprocal cross contamination between the two ve sicle preparations. By using the BLM vesicles and a rapid filtration t echnique, we quantified Cl-36 uptake under conditions of equilibrated pH (pH(out) = pH(in) = 7.5). The presence of 200 mM cis of either Na- or K-+, or an equimolar mixture of both, significantly increased the initial Cl- entry rate. In the presence of K+, valinomycin further inc reased Cl- uptake, but no Cl- uphill transport was ever observed under any of the conditions. Ah the increases were abolished by voltage cla mping, indicating that the alkali-metal ions act by creating an inside -positive membrane potential capable of stimulating a Cl--conductance pathway. In the absence of K+, BLM vesicle preloading to obtain a [Cl- ](out)/[Cl-](in) = 16/200 mM gradient (Delta Cl-) resulted in a 500% i ncrease in the initial Cl-36(-) entry rate, accompanied by a transient Cl- accumulation, with an overshoot at similar to 5 min. In the prese nce of both a positive-inside electrical gradient (Delta Psi) and a De lta Cl-, the initial Cl- uptake rate was increased by 800%, indicating that the effects of Delta Psi, and of Delta Cl- are additive. The Del ta Cl- effect was blocked, but only partially, by short-circuiting the membrane potential with equilibrated K+ and valinomycin, thus indicat ing that it has both rheogenic and electroneutral components. We concl ude that Cl- influx across the guinea pig intestinal BLM involves a Cl --conductance pathway plus a distinct Cl--Cl--exchange system, exhibit ing both electroneutral and rheogenic components. Alternatively, the p ossibility can also be entertained that the conductance and the exchan ge pathways share a common molecular basis, e.g., a nonobligatory Cl-- Cl- exchanger or rheogenic uniport.