PMID: 9427292Jan 14, 1998Paper

Intracellular Ca2+ signals in Dictyostelium chemotaxis are mediated exclusively by Ca2+ influx

Journal of Cell Science
T Nebl, Paul R Fisher

Abstract

We measured folate- and cAMP-induced changes in cytoplasmic free calcium concentration ([Ca2+]i) using recombinant aequorin reconstituted in living Dictyostelium cells with coelenterazine-h. The resulting semi-synthetic protein displayed increased sensitivity to Ca2+ allowing accurate measurement of chemoattractant-induced transients at low resting levels. Both folate- and cAMP-induced Ca2+ responses were developmentally regulated, exhibited remarkably similar kinetics and were dependent on the relative rather than the absolute magnitude of increases in attractant concentration. They began after a short delay of 5-10 seconds, leading to a maximum increase in cytosolic calcium concentration after approximately 25 seconds and a return to basal level within approximately 60 seconds after stimulation. Responses elicited by the two chemoattractants were dose-dependent and saturated between 4 and 20 microM. They depended on the presence of free extracellular calcium ions and were inhibited in a concentration-dependent manner between 10(-4) and 10(-5) M. In accordance with 45Ca2+-uptake measurements by Milne and Coukell (J. Cell Biol. (1991) 112, 103-110), both responses were also completely inhibited by 15 microM Ruthenium Red, 15 mi...Continue Reading

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