Ca2+ regulation of gap junctional coupling in lens epithelial cells

The quantitative effects of Ca2+ signaling on gap junctional coupling in le ns epithelial cells have been determined using either the spread of Mn2+ th at is imaged by its ability to quench the fluorescence of fara 2 or the spr ead of the fluorescent dye Alexa Fluor 594. Gap junctional coupling was una ffected by a mechanically stimulated cell-to-cell Ca2+ wave. Furthermore, w hen cytosolic Ca2+ concentration (Ca-i(2+)) increased after the addition of the agonist ATP, coupling was unaffected during the period that Ca-i(2+) w as maximal. However, coupling decreased transiently similar to5-10 min afte r agonist addition when Ca-i(2+) returned to resting levels, indicating tha t this transient decrease in coupling was unlikely due to a direct action o f Ca-i(2+) on gap junctions. An increase in Ca-i(2+) mediated by the ionoph ore ionomycin that was sustained for several minutes resulted in a more rap id and sustained decrease in coupling (IC50 similar to 300 nM Ca2+, Hill co efficient of 4), indicating that an increase in Ca-i(2+) alone could regula te gap junctions. Thus Ca-i(2+) increases that occurred during agonist stim ulation and cell-to-cell Ca2+ waves were too transient to mediate a sustain ed uncoupling of lens epithelial cells.