Ca2+ dynamics in synaptosomes isolated from the squid optic lobe

Synaptosomes from the optic lobes of squid (Loligo forbesi) were prepared b y homogenization and allowed to settle onto glass coverslips. Synaptosomes were loaded with Ca2+ sensitive dyes (Fura-2 AM, Calcium Green-1 AM and Cal cium Green-5N AM), visualized by light microscopy and Ca2+ sensitive fluore scence signals recorded and analyzed. With Fura-2, resting Ca2+ was found t o be 80 nM (n = 10, SEM 5.7). Addition of K+ (30 mM), caffeine (3 mM) and t hapsigargin (10 muM) evoked transient increases in cytoplasmic Ca2+. Additi on of BAPTA-AM (20 muM) decreased intrasynaptosomal free Ca2+. Similar resu lts were obtained with Calcium Green-1 AM but not with Calcium Green-5N AM. We conclude that synaptosomes from the squid optic lobe posses intact memb ranes and mechanisms to regulate intrasynaptosomal free [Ca2+], as well as caffeine sensitive Ca2+ stores. The results of this study are discussed wit h respect to the role of Ca2+ in presynaptic protein synthesis. (C) 2000 Wi ley-Liss, Inc.