Ca(2+) dynamics in synaptosomes isolated from the squid optic lobe.
Synaptosomes from the optic lobes of squid (Loligo forbesi) were prepared by homogenization and allowed to settle onto glass coverslips. Synaptosomes were loaded with Ca(2+) sensitive dyes (Fura-2 AM, Calcium Green-1 AM and Calcium Green-5N AM), visualized by light microscopy and Ca(2+) sensitive fluorescence signals recorded and analyzed. With Fura-2, resting Ca(2+) was found to be 80 nM (n = 10, SEM 5.7). Addition of K(+) (30 mM), caffeine (3 mM) and thapsigargin (10 microM) evoked transient increases in cytoplasmic Ca(2+). Addition of BAPTA-AM (20 microM) decreased intrasynaptosomal free Ca(2+). Similar results 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 membranes and mechanisms to regulate intrasynaptosomal free [Ca(2+)], as well as caffeine sensitive Ca(2+) stores. The results of this study are discussed with respect to the role of Ca(2+) in presynaptic protein synthesis.
Benech
J C
JC
Departmento de Biofísica, Instituto de Investigaciones Biológicas Clemente Estable, Montevideo, Uruguay.
Lima
P A
PA
Sotelo
J R
JR
Brown
E R
ER
D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D000697:Central Nervous System Stimulants / Q000494:pharmacology; D002614:Chelating Agents / Q000494:pharmacology; D049832:Decapodiformes; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005456:Fluorescent Dyes / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009899:Optic Lobe, Nonmammalian / Q000187:drug effects / Q000378:metabolism*; D009930:Organic Chemicals; D011189:Potassium Chloride / Q000494:pharmacology; D013574:Synaptosomes / Q000187:drug effects / Q000378:metabolism*; D019284:Thapsigargin / Q000494:pharmacology
D000697:Central Nervous System Stimulants; D002614:Chelating Agents; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D009930:Organic Chemicals; C071336:calcium green; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D002110:Caffeine; D004533:Egtazic Acid; D011189:Potassium Chloride; D019284:Thapsigargin; D002118:Calcium