Ca2+ regulation of gap junctional coupling in lens epithelial cells

The quantitative effects of Ca2+ signaling on gap junctional coupling in lens epithelial cells have been determined using either the spread of Mn2+ that is imaged by its ability to quench the fluorescence of fara 2 or the spread of the fluorescent dye Alexa Fluor 594. Gap junctional coupling was unaffected by a mechanically stimulated cell-to-cell Ca2+ wave. Furthermore, when cytosolic Ca2+ concentration (Ca-i(2+)) increased after the addition of the agonist ATP, coupling was unaffected during the period that Ca-i(2+) was maximal. However, coupling decreased transiently similar to5-10 min after agonist addition when Ca-i(2+) returned to resting levels, indicating that this transient decrease in coupling was unlikely due to a direct action of Ca-i(2+) on gap junctions. An increase in Ca-i(2+) mediated by the ionophore ionomycin that was sustained for several minutes resulted in a more rapid and sustained decrease in coupling (IC50 similar to 300 nM Ca2+, Hill coefficient of 4), indicating that an increase in Ca-i(2+) alone could regulate gap junctions. Thus Ca-i(2+) increases that occurred during agonist stimulation and cell-to-cell Ca2+ waves were too transient to mediate a sustained uncoupling of lens epithelial cells.