EFFECTS OF CALCIUM ON GAP-JUNCTIONS BETWEEN OSTEOBLAST-LIKE CELLS IN CULTURE

The present study investigated the effects of elevated cytoplasmic fre e calcium concentrations ([Ca2+](i)) on the permeability of gap juncti ons between cultured osteoblast-like (OB) cells derived from calvarial and periosteal fragments of newborn rats. This was studied using the double whole cell patch clamp technique and intracellular dye injectio ns. To increase [Ca2+](i), patch pipette solutions contained 100 mu mo l/liter Ca2+. About 1-2 minutes after whole cell modes had been attain ed, the total number of gap junction channels was reduced from an aver age of 400 in normal Ca2+ to 20 in high Ca2+. Thereafter, remaining ga p junction channels were active for up to 8 minutes. In normal rat kid ney (NRK) cells, gap junction channels were closed by high Ca2+ within 1 minute, pointing to a similar sensitivity of Connexin43 gap junctio n channels in OB and NRK cells. To study the effects of elevated [Ca2](i) on the dye permeability of gap junctions between extended OB cell s, the spread of Lucifer Yellow to neighboring cells was evaluated. [C a2+](i) was gradually increased from 1.5- to 14-fold the normal value by application of either ouabain, Na+-free/ouabain, or A23187. Reduced dye spread correlated with the increase of [Ca2+](i) measured by anal yzing the fluorescence of fura-2. These data show that gap junctions i n OB cells are sensitive to Ca2+.