REPETITIVE CALCIUM WAVES INDUCED BY FERTILIZATION IN THE NEMERTEAN WORM CEREBRATULUS-LACTEUS

To analyze fertilization-induced calcium dynamics in a protostome worm , unfertilized oocytes of the nemertean Cerebratulus lacteus were co-i njected with calcium green (CG) and rhodamine (Rh) dextrans for dual-c hannel confocal imaging of early development. Based on CG/Rh ratioed i mages collected every 800 msec, fertilization elicits a ''cortical fla sh'' of elevated free calcium that spreads rapidly around the oocyte w ithout propagating as a point-source wave. A similar calcium transient occurs in unfertilized oocytes treated with KCI to depolarize the ool emma, and the fertilization-induced cortical hash is eliminated if cob alt is used to block calcium channels, collectively indicating that fe rtilization initially triggers an influx of calcium ions through volta ge-gated calcium channels in the oolemma. However, within minutes afte r producing a cortical hash, C. lacteus oocytes begin to display a ser ies of point-source, oscillating waves of elevated free calcium that a re propagated at about 15 mu m/sec. The first two calcium waves arise at the site of sperm fusion and typically fail to reach the antipode, but after sperm incorporation, the waves spread globally throughout th e ooplasm and typically shift their origin to a pacemaker region in th e vegetal cortex. About 10 oscillations with an average duration of 3. 3 +/- 1.2 min are generated for similar to 60-100 min postfertilizatio n as meiotic maturation is completed, and such waves continue to occur in cobalt-containing seawater or calcium-free seawater. Thus, wavelik e calcium oscillations induced by fertilization are apparently depende nt upon internal calcium stores, which in turn may contain IP3-insensi tive and/or IP3-sensitive receptors based on experiments using ryanodi ne, caged IP3, and heparin. Unfertilized oocytes also display repetiti ve calcium waves following intracytoplasmic injections of whole sperm, and such oscillations are eliminated if the sperm suspensions are boi led prior to injection, suggesting the possible presence of a heat-lab ile sperm component that can elicit wavelike oscillations during ferti lization. (C) 1996 Academic Press, Inc.