CA2-URCHIN EGGS - RELATIONSHIP TO LARVAL DEVELOPMENT AND RELEVANCE INEGG ACTIVATION( RELEASE FROM INTRACELLULAR STORES BY THAPSIGARGIN IN SEA)

Thapsigargin (Tg), an inhibitor of microsomal Ca2+ ATPase, is used as a tool to study the changes in Ca2+ sequestration in sea urchin eggs a nd their relationship to embryonic development. Micromolar amounts of Tg inhibit ATP-dependent Ca2+ sequestration in a dose-dependent and no n-reversible manner, depending on the bulk of biological material used . IC50 values are 1 nmol/L and 1-10 mu mol/L, respectively, in the cor tical Ca2+ stores (isolated cortices preparation) and in digitonin-per meabilized eggs, a preparation giving access to the deeper reticulum c ompartment. Micromolar Tg does not induce Ca2+ release from Ca-45 pre- loaded cortices but leads to a loss of 25% of the total Ca2+ content f rom the cortical area. Using microspectrofluorimetry of fura-2-loaded eggs, we found that 10 mu mol/L Tg induced a moderate rise in cytosoli c Ca2+ activity as compared with the fertilization-induced Ca2+ transi ent whether eggs were fertilized or not. Early events related to ferti lization as, far example, elevation of the fertilization envelope, pro ton excretion and sustained increase of amino acid uptake, are trigger ed by 10 mu mol/L Tg but with a delayed onset relative to sperm-induce d effects. The present findings indicate that although it triggers mos t fertilization-related events, Tg cannot be considered as a true mito tic agent in sea urchin eggs. When added after fertilization, Tg affec ts cleavage and the further embryonic development giving rise to abnor malities comparable to the animalized larvae obtained with other compo unds responsible for the inhibition of reticular Ca2+ sequestration.