Monitoring cytosolic calcium in the dinoflagellate Crypthecodinium cohnii with calcium orange-AM.
Calcium plays several important roles in the signal transduction pathways of dinoflagellates. We describe here the development of calcium orange-AM as an intracellular calcium reporter for the heterotrophic dinoflagellate Crypthecodinium cohnii. We demonstrated with confocal microscopy that by restricting the incubation period to 30-45 min, no compartmentalization of the dye occurs in the mitochondria or endoplasmic reticulum. The dye fluorescence responded well to the effects of calcium ionophores and calcium chelators. By calibrating the dye with known calcium concentrations, we determined the intracellular calcium concentration of C. cohnii to be 158 +/- 56 nM, which rose to about 550 nM upon mechanical stimulation.
Lam
Connie M C
CM
Biology Department, Hong Kong University of Science and Technology, Clearwater Bay, Kowloon, Hong Kong SAR, PR China.
Yeung
Patrick K K
PK
Wong
Joseph T Y
JT
D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002614:Chelating Agents / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D004141:Dinoflagellida / Q000187:drug effects / Q000378:metabolism*; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D005456:Fluorescent Dyes; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D018613:Microscopy, Confocal; D009930:Organic Chemicals; D013314:Stress, Mechanical
D002614:Chelating Agents; D005456:Fluorescent Dyes; D007476:Ionophores; D009930:Organic Chemicals; C071335:calcium orange; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D000001:Calcimycin; D004533:Egtazic Acid; D015759:Ionomycin; D002118:Calcium