LOCALIZATION OF CYTOPLASMIC FREE CALCIUM-IONS IN PC12 CELLS WITH VARICOSE FIBERS

Differentiated PC12 cells with varicose fibers were used as a model of sympathetic neurons to demonstrate the intracellular localization of cytoplasmic free calcium ions. Changes in the concentration of cytopla smic free calcium ions were analyzed at individual varicosities upon s timulation with acetylcholine by laser scanning confocal microscopy, T ransient increases in cytoplasmic free calcium ion concentration were localized in the varicosities and recognized in both the absence and p resence of extracellular Ca2+. Immunocytochemical analysis of intracel lular calcium channels, 1, 4, 5-trisphosphate receptors and ryanodine receptors, by electron microscopy demonstrated that immunoreactive sit es were mainly localized in large dense core vesicles in the varicosit ies and neurites. These results suggest that the exocytosis of large d ense core vesicles is regulated by an increase in cytoplasmic free cal cium ion concentration from an intracelluler Ca2+ store.