COVI, J.A.*; HAND, S.C.; Louisiana State University; Louisiana State University: Physiological effects of V-ATPase inhibition on whole embryos of the brine shrimp, Artemia franciscana

Upon exposure to anoxia, encysted embryos of the brine shrimp, Artemia franciscana, enter a potentially ametabolic state, the transition into which is facilitated in large part by an intracellular acidification exceeding 1.5 pH units. However, while the critical role of this pH shift in metabolic downregulation is now clear, its endogenous origin remains largely unexplained. We hypothesize that the yet unidentified proton equivalents involved in this event originate from the dissipation of proton chemical gradients between acidic compartments and the cytoplasm. These gradients are likely to be established during aerobic development by an ATP-dependent proton pump like the V-ATPase, and subsequently dissipated in response to the severe energetic constraints imposed by anoxia. Here we report that exposure of anoxia tolerant embryos to nanomolar concentrations of bafilomycin (a specific inhibitor of the V-ATPase) decreased oxygen consumption rates in these intact embryos by almost 80% over 3.75 h, but did not affect respiration rates of their isolated mitochondria. Incubation with bafilomycin also decreased hatching success of embryos in a dose dependent manner. These data support an argument for the involvement of proton gradient dissipation in the metabolic downregulation induced by anoxia in this crustacean. [Supported by DARPA grant N00173-01-1-G011 and NIH grant 1-R01-GM071345-01]