ニシン卵の受精の研究-V

Abstract

(1) Ca ions play an extremely important role in the fertilization of Clupea eggs. When the eggs are repeatedly washed in an isotonic Ca-free Ringer's solution (M/4.5 NaCl 100 parts+M/4.5 KCl 3.5 parts+M/6.7 MgCl2 2.4 parts, pH 7.6 by NaHCO3) and inseminated in such a solution, not a single one is fertilized. However, if a few drops of CaCl2 solution are added to this solution, all the eggs are fertilized at once, In one experiment, the eggs and sperm were kept overnight in the Ca-free Ringer's solution. The next morning all the eggs were intact and not a single one was fertilized. At that time a few drops of CaCl2 Solution were added to this solution and instantly all the eggs were fertilized and began to cleave at the proper time. Thus the fertilization of the Clupea egg is closely dependent on Ca ions in the surrounding medium. It has been found that the failure of fertilization in the Ca-free Ringer's solution results from the fact that the spermatozoa fail to enter the micropyle. As pointed out previously (YANAGIMACHI'57: Zool. Mag. 66; Bull. Jap. Soc. Sci. Fish. 23), the Clupea spermatozoa are rendered extremely active in the micropyle area of the mature egg. When Ca ions are lacking in the surrounding medium, the spermatozoa are not activated: the spermatozoa are quiescent even if they are brought to the micropyle area of the egg, so that not a single spermatozoon enters the micropyle. It is only when a sufficient amount of Ca ions is present in the medium, that the spermatozoa are activated and enter the micropyle. It has been shown that Mg ions can be substituted for Ca ions, though the former must be in higher concentration than the latter. Data on fertilization of the eggs in the solutions containing various amounts of Ca ions and Mg ions are shown in Tables 1 and 2. (2) Ca ions are necessary for parthenogenetic activation of the eggs. When the eggs are repeatedly washed in the Ca-free Ringer's solution and pricked with a fine glass needle, not a single egg is activated. The same is true when the eggs are treated with an isotonic solution of sodium oxalate (a Ca precipitant) for 15 minutes or longer. When such eggs are transferred into the isotonic Ringer's solution containing Ca ions, all the eggs are instantly activated without further pricking. Thus, the activation by pricking seems to depend on the presence of Ca ions. The eggs are also activated by chemical agents, such as saponin. In this case, too, the presence of Ca ions in the surrounding medium is of essential importance. (3) The eggs once fertilized are able to cleave in the Ca-free Ringer's solution. However, blastomeres tend to pull apart from each other (Fig. 1). This is probably due to the dissolution. of the so-called “surface gel layer”.