Effects of calcium antagonists, calmodulin antagonists, and methylated xanthines on polar lobe formation and cytokinesis in fertilized eggs of Ilyanassa obsoleta

Abstract

We have studied the ability of fertilized eggs of Ilyanassa obsoleta to undergo polar lobe formation and cytokinesis in the presence of Ca2+ antagonists (Ca2+ channel blockers, Ca2+ uptake inhibitors). Earlier work had suggested little need for exogenous Ca2+ during these cellular shape changes. Again it appears that exogenous Ca2+ probably is not required, based on cell ability to undergo the shape changes with no, or only minor, delay in the presence of 50 mM La3+ at pH 6.5, 10 mM concentrations of Ni2+ or Co2+, 1 mM Cd2+, and 100 microM concentrations of Mn2+, papaverine, verapamil, D600, or diltiazem. In nominally Ca2+-free seawater (containing approximately 10 microM Ca2+) (CFSW), there still is no effect of Cd2+ (up to 100 microM), Ni2+, Co2+, Mn2+, or diltiazem; however, papaverine, verapamil, and D600 in CFSW cause longer delays in the shape changes than they do in the presence of normal levels of Ca2+ (SW). In 10-50 microM nifedipine, shape changes are progressively delayed to the same extent in both SW and CFSW, but more so in CFSW at concentrations above 50 microM nifedipine. Among calmodulin antagonists, trifluoperazine up to 100 microM was without effect, but chlorpromazine at 25-100 microM and calmidazolium at 50-100 microM caused substantial, concentration-dependent delays in the starting times for the shape changes. Methylxanthines caused a substantial speed-up in the starting times for both polar lobe formation and cytokinesis. The most effective of these, caffeine, at optimal concentrations of 0.7-10 mM in SW or CFSW caused shape changes to occur 12-15 min earlier than in controls undergoing a normal 50-min cycle. Caffeine is known to cause release of Ca2+ from muscle sarcoplasmic reticulum. A putative antagonist of intracellular Ca2+ mobilization, TMB-8, significantly inhibited the shape changes of the Ilyanassa cells, whereas a variety of inhibitors of exogenous Ca2+ uptake noted above did not inhibit. We conclude that Ca2+ may be necessary for polar lobe formation and cytokinesis in Ilyanassa cells, but that it may be released from intracellular, sequestered stores rather than derived from exogenous sources.

Polar effects of concanavalin A on the cortical cytoskeleton of a molluscan egg (Nassarius reticulatus, Gastropoda)

“…This inhibitor does not affect polar lobe formation and cytokinesis, as was found previously for Ilyanassa (Conrad et al 1987). In addition, D600…”

“…The possible role of calcium released from intracellular stores was investigated using TMB-8, which is an antagonist of intracellular calcium mobilization and is known to affect normal polar lobe formation in Ilyanassa (Conrad et al 1987). We found that continuous incubation with 200 ~tM TMB-8 concomitantly with Con A treatment prevents the occurrence of typical Con A-induced effects.…”

Calcium buffer injections delay cleavage in Xenopus laevis blastomeres

“…How are these suspected [Ca2+]i gradients generated and what are their likely targets? Our earlier work (10) suggested that, in addition to [Ca2+]i gradients, PIP2 turnover and Ins (1,4,5)P3-induced Ca 2+ release were required for normal cleavage. Thus, components of the inositol cascade may well be involved in this [Ca2+]~ gradient generation.…”

“…Imposed increases in [Ca2+]~ have been shown to induce nuclear envelope breakdown in both sea urchin eggs and mammalian 3T3 fibroblasts, while suppressing [Ca2+]t changes with the calcium buffer, BAPTA, inhibited nuclear envelope breakdown in sea urchin eggs and Swiss 3T3 cells (15,28). Elevation of [Ca2+]~ in squid eggs by treatment with the calcium ionophore, A23187 (1), and in llyanassa eggs by treatment with caffeine (5), has been shown to speed the onset of cleavage. Cell division was retarded in sea ur-chin (33), medaka (8), and Xenopus (10,19) embryos in which [Ca2+]i changes were suppressed with the BAPTA family of buffers and earlier work with much higher concentrations of EGTA also resulted in some cleavage inhibition (2,31).…”

Effects of silver ions (Ag+) on contractile ring function and microtubule dynamics during first cleavage in Ilyanassa obsoleta

“…During phase I, a period lasting 50 ? 5 min (timing of phases I-IV: Conrad et al [1987]), the PLC constricts at a constant slow rate to deform the egg into a pear shape ( Fig. 1: Cont 30 min), while the mitotic apparatus forms, expands, and then disassembles in the animal hemisphere.…”