TRANSIENT NA-HOST CELLS AND SUBSEQUENT IMMERSION IN SEAWATER( STRESS IN SYMBIOTIC DINOFLAGELLATES AFTER ISOLATION FROM CORAL)

Dinoflagellates which live in intracellular symbiosis with corals (zoo xanthellae), probably share the ionic conditions of their host cells, i.e. are subjected to lower sodium and calcium concentrations than amb ient seawater. Although free-living zooxanthellae are not generally fo und in waters of reef ecosystems, they can be released in either a con trolled diurnal regulation or an uncontrolled (coral bleaching) reacti on by their animal hosts. Upon release, zooxanthellae experience new e xternal ionic conditions. The aims of this study were to (1) examine t he ionic conditions experienced by zooxanthellae in hospite, (2) deter mine changes in the intracellular Na+ concentration of dinoflagellates (Symbiodinium sp.) following isolation from the scleractinian coral G alaxea fascicularis (Linnaeus, 1767), and (3) characterize the mechani sm of Na+ regulation and control. On the basis of equilibrium studies, it has been suggested that zooxanthellae in coral-host cells experien ce a Na+ concentration of similar or equal to 60mM. The intracellular concentration of Na+ in zooxanthellae, as determined by flame photomet ry, was found to be similar or equal to 0.300 mu equiv mg(-1) protein, or 30 to 35 mM when the water content of cells was taken into account . Half the cell Na+ seems to be compartmentalized (i.e. non-exchangeab le) in freshly isolated zooxanthellae (FIZ), while in cultured zooxant hellae (CZ) all the Na+ was exchangeable. Isolation of zooxanthellae i nto seawater from their intracellular environment caused a transient t wo-fold increase in Na+ concentration within the first 30 min. This in crease was directly proportional to extracellular Na+ concentration, s uggesting passive influx. After 30 min the Na+ concentration decreased , reaching its initial level within 1 to 3 h. Following isolation for up to 3 h, the Na+ influx rate measured during short incubations (5 mi n) was constant, suggesting that some Na+-regulation mechanism, probab ly a Na+ efflux system, was created or-stimulated within the first 30 min. Since this mechanism is independent of DNA transcription (as prov ed by its insensitivity to 100 mu M actinomycin D) and protein synthes is (insensitivity to 100 mu M cycloheximide or emetin), we conclude th at activation of an efflux system already present in the zooxanthella membrane occurred during the first 30 min after isolation in seawater. This mechanism was cyanide (CN)- and 3-(3,4-dichlorophenyl)-1, 1-dime thylurea (DCMU)-sensitive and dependent on extracellular K+. However, while we found an ouabain sensitive ATPase activity in zooxanthella ho mogenates, suggesting the involvement of a Na+/K+ ATPase, no correlati on was found between Na+ efflux and K+ influx [measured using Rb-86 (r ubidium chloride) as a tracer]. The results presented show that isolat ion of symbiotic dinoflagellates into seawater causes major ionic stre ss, resulting in a two-fold increase in intracellular Na+ concentratio n. Within 30 min, a Na+-efflux mechanism (putatively involving Na+-ATP ase), in the membrane of zooxanthellae ir? hospite is stimulated. The Na+ concentration of the cell consequently returns to its initial leve l 3 h after isolation.