Measurement of intracellular Ca2+ activity in Necturus gallbladder.

Abstract

To clarify the effects of Ca2+-free solutions on the electrophysiological properties of leaky epithelia, Necturus gallbladder was mounted in an Ussing-type chamber and its mucosal surface exposed to Ca2+-free EGTA (2 mM) Ringer. Lateral-space width was controlled by a -3-cmH2O pressure gradient on the serosal outflow. Transepithelial potential difference and resistance were monitored while cell membrane potential and intracellular Ca2+ activity (Ca2+i) were determined with conventional and Ca2+-sensitive microelectrodes. Ca2+i averaged 183 +/- 27 nM (n = 15). Reduction of mucosal Ca2+ activity to approximately 500 nM reversibly lowered transepithelial resistance while cell membrane potential remained unaltered and fractional membrane resistance increased from 0.77 +/- 0.01 to 0.83 +/- 0.02 (P less than 0.01, n = 5). In five gallbladders mucosal Ca2+ reduction induced a significant drop in Ca2+i from 133 +/- 26 to 77 +/- 20 nM (P less than 0.01, n = 5) while transepithelial resistance fell from 125 +/- 27 to 107 +/- 24 omega X cm2 (P less than 0.01). These results indicate that transepithelial resistance decrements observed during exposure to Ca2+-free solutions stem from a reversible increase in tight-junctional but not cell membrane permeability and that this effect is associated with a fall in intracellular Ca2+ activity.