Basolateral impalement of intestinal villus cells: electrophysiology of Cl- transport

Abstract

A method of dissecting the serosal muscle layers is described that transforms the villus of isolated Amphiuma small intestine into a flat sheet one cell layer thick, allowing rapid equilibration of the serosal medium with the basolateral membrane of the villus tip cells and direct access of the basal membrane to microelectrodes. The "villus sheet" preparation was used to examine the luminal and basolateral mechanisms of Cl- transport. The serosal membrane potential (Vs), measured with conventional microelectrodes, averaged -79.7 mV in tissues bathed in Cl- -free medium; the mucosal membrane potential (Vm) averaged -80.9 mV. Fractional resistance measured directly was 0.82 and 0.14 for the mucosal and serosal membranes, respectively. Elevation of bath [K] reduced Vm and Vs by 30.3 and 44.5 mV, respectively. Cl- (20 mM) added to the luminal medium reduced Vm by 23.9 mV and stimulated Cl- transport; luminal addition of furosemide then increased Vm by 5.6 mV and reduced Cl- transport. Addition of Cl- (20 mM) to the Cl- -free serosal fluid increased Vs 2.0 +/- 1.9 mV. On reducing the serosal [Cl] 10-fold Vs decreased 2.0 +/- 2.2 mV. These and other results indicate that basolateral Cl- exit is not over a conductive pathway. The villus sheet affords new opportunities for studying enterocyte function in the intact mucosa.