INTRACELLULAR FREE CALCIUM LEVEL AND ITS RESPONSE TO CAMP STIMULATION IN DEVELOPING DICTYOSTELIUM CELLS TRANSFORMED WITH JELLYFISH APOAEQUORIN CDNA

A new method is described for measuring intracellular free calcium concentrations, [(Ca2+)(i)], in the cells of Dictyostelium discoideum transformed with apoaequorin cDNA of the jellyfish, Aequorea victoria. Aequorin, a calcium-specific indicator, was regenerated in vivo from apoaequorin produced in the cells by incubation with coelenterazine. The results showed that [(Ca2+)(i)] in developing cells markedly increases at the aggregation stage and again at the culmination stage after a temporary drop at the migration stage. Except for the vegetative stage, the cells al all stages of development exhibit a sharp transient increase in [(Ca2+)(i)] upon stimulation with a cAMP (50 nM) pulse, high responses being observed at the migration and culmination stages. Separated prestalk cells of migrating slugs contain more than twice as much [(Ca2+)(i)] and show three times as large a response to cAMP stimulation as prespore cells.