REGULATION OF THE FERTILIZATION CURRENT IN ASCIDIAN OOCYTES BY INTRACELLULAR 2ND MESSENGERS

Neomycin, injected into ascidian oocytes to a final concentration of 1 0-50 mM, inhibits both the fertilization current and the surface contr action, showing that phosphoinositide hydrolysis is required for these early activation events. Sperm-activated fertilization currents are n ot inhibited in the presence of 100 mug/ml intracellular heparin, sugg esting that these currents are not directly gated by InsP3. The sulfhy dryl reagent thimerosal at 100 muM, in contrast, significantly increas es the fertilization current presumably by sensitizing the channel rec eptor. Since heparin inhibits the surface contraction, InsP3 receptors are shown to play a role in the propagation of the activation respons e in ascidian oocyte. Depleting intracellular calcium stores by microi njecting 50 mM EGTA into oocytes does not activate fertilization chann els; however, subsequent fertilization of these EGTA loaded oocytes le ads to a significantly larger and faster fertilization current. Thus i n contrast to somatic cells studied to date, second messenger operated plasma membrane channels in ascidian oocytes are not gated by calcium released from intracellular stores. (C) 1994 Wiley-Liss, Inc.