Journal Article

Monitoring Cytosolic Calcium in the Dinoflagellate <i>Crypthecodinium cohnii</i> with Calcium Orange-AM

Connie M. C. Lam, Patrick K. K. Yeung and Joseph T. Y. Wong

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 46, issue 6, pages 1021-1027
Published in print June 2005 | ISSN: 0032-0781
Published online June 2005 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/pcp/pci102
Monitoring Cytosolic Calcium in the Dinoflagellate Crypthecodinium cohnii with Calcium Orange-AM

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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.

Keywords: Dinoflagellates; Intracellular calcium; Mechanical stimulation; AM, acetoxymethyl ester; BAPTA-AM, 1,2-bis (2-aminophenoxy) ethane-N,N,N′,N′-tetraacetic acid tetra (acetoxymethyl) ester; Ca2+i, intracellular calcium; CDCFDA, 5-(and -6)-carboxy-2′,7′-dichlorofluorescein diacetate; DiOC6(3), 3,3′-dihexyloxacarbocyanine iodide; DMSO, dimethylsulfoxide; EGTA, ethylene glycol-bis-(2-aminoethyl)-N,N,N′,N′-tetraacetic acid; ER, endoplasmic reticulum; Kd, dissociation constant

Journal Article.  4414 words.  Illustrated.

Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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