Ionic basis of methacholine-induced shrinkage of dissociated eccrine clear cells

Abstract

The goal of the present study was to elucidate the ionic mechanisms by which cholinergic stimulation induces cell shrinkage in eccrine clear cells. Dissociated Rhesus monkey eccrine sweat clear cells were prepared by collagenase digestion of freshly isolated secretory coils and immobilized on a glass slide in a perfusion chamber at 30 degrees C. The cell was visualized by light microscopy with differential interference contract (DIC) and was recorded with a video system (15,000 x total magnification). The cell volume was calculated from the maximal cross section of the cell. Methacholine (MCh)-induced cell shrinkage, which was as much as 30% of resting cell volume, was dose dependent and pharmacologically specific. MCh-induced cell shrinkage was persistent in some cells but tended to partially wane with time in others. MCh-induced cell shrinkage was dependent on the chemical potential gradient for KCl, i.e., increasing [K] in the bath ([K]o) from 5 to 120 mM caused MCh to induce cell swelling, whereas removing [Cl]0 at 120 mM K partially restored the MCh-induced cell shrinkage. The interpolated null [K]o (medium [K] where the cell volume did not change by MCh) of 71 mM agreed with the predicted [K]o,null. MCh-induced cell shrinkage was inhibited completely by 1 mM quinidine (K-channel blocker) and partially by 1 mM diphenylamine-2-carboxylic acid (DPC, a Cl-channel blocker), but not by 0.1 mM ouabain or 0.1 mM bumetanide, suggesting that MCh-induced cell shrinkage may be due to activation of both K and Cl channels with the resultant net KCl efflux down the chemical potential gradient.(ABSTRACT TRUNCATED AT 250 WORDS)