The time between injection of dibromo-BAPTA and fixation of the eggs in the present study was sufficient for the buffer to diffuse to both the animal pole and vegetal pole, to dissipate cytosolic [Ca.sup.2+] gradients near the poles, and to inhibit the formation of the blastodisc and the movement of oil droplets toward the vegetal pole (Fluck et al., 1992, 1994).
Scanning electron microscopy studies on blastodisc formation in the zebrafish, Brachydanio rerio.
Moreover, three microtubule poisons - colchicine, demecolcine, and nocodazole - inhibit the normal movement of oil droplets toward the vegetal pole, eliminate the saltatory motion of small inclusions, and slow the growth of the blastodisc: eggs treated with [Beta]-lumicolchicine, an inactive derivative of colchicine, segregated normally (Abraham et al., 1993a).
The volume of the blastodisc was measured with an image analysis program (Microcomp Planar Morphometry, Southern Micro Instruments; Abraham et al., 1993a).
Eggs treated with 1% DMSO formed a blastodisc (volume at [T.sub.n] [approximately equal to] 0.85-1.0 = 15.4 [+ or -] 2.0 nl, [Mathematical Expression Omitted], N = 7 eggs; [ILLUSTRATION FOR FIGURE 1A OMITTED]), displayed cytoplasmic streaming (speed of parcels toward the animal pole = 9.00 [+ or -] 5.52 [[micro]meter] [min.sup.-1], [Mathematical Expression Omitted], N = 7 eggs), and developed normally.
(1993a): (1) oil droplets did not segregate to the vegetal pole but instead floated to the portion of the egg that was uppermost during the experiment - equator [ILLUSTRATION FOR FIGURE 1C OMITTED], animal pole [ILLUSTRATION FOR FIGURE 2C AND D OMITTED], or vegetal pole; (2) a smaller-than-normal blastodisc formed [ILLUSTRATION FOR FIGURE 1C OMITTED]; (3) saltatory motion was absent; and (4) the eggs did not cleave.
However, in preliminary experiments, we found that higher light intensities ([greater than or equal to] 1100 [Mu]W [cm.sup.-2], 1.0 x [10.sup.10] quanta [s.sup.-1] [[[micro]meter].sup.-2]) inhibited both the growth of the blastodisc and the movements of oil droplets.
In all these eggs, the droplets of injected fluids moved toward the animal pole and came to rest in the blastodisc [ILLUSTRATION FOR FIGURE 6B-D OMITTED].
However, in one egg in which the injected droplet was not observed to fuse with endogenous droplets, the injected droplet moved up, toward the animal pole, and came to rest near the blastodisc, while nearby endogenous droplets moved by it in the opposite direction on their way to the vegetal pole.
The inhibition of cytoplasmic streaming and formation of the blastodisc by CCD in medaka eggs is consistent with data obtained from other teleost eggs (Katow, 1983; Ivanenkov et al., 1987) and from eggs of other organisms, including ascidians (Sawada and Osanai, 1981; Jeffery, 1984) and an oligochaete (Shimizu, 1982).