No abstract

Many ascidians live in clumps and usually release sperm before the eggs. Consequently, eggs are often spawned into dense clouds of sperm. Because fertilization by more than a single sperm is lethal, ascidians have evolved at least two successive blocks to polyspermy: the rapid release of a glycosidase that inhibits sperm binding to the vitelline coat (VC) and a subsequent change in membrane potential that prevents supernumerary sperm–egg fusion. This paper shows that (1) these two blocks can be uncoupled by the use of suramin, and (2) most of the glycosidase appears to be from the follicle cells, which are accessory cells on the outside of the egg VC. Phallusia mammillata eggs initially bind numerous sperm but, after the glycosidase is released, only a few additional sperm bind. Intact eggs in 20 μM suramin release glycosidase, but the electrical response is inhibited; sperm swim actively and bind to the VC but fail to penetrate. Suramin treatment is completely reversible; intact eggs exhibit the electrical response an average of 11 minutes after the drug is washed out. Sperm must contact the follicle cells before passing through the VC; eggs with the VC removed and fertilized in the presence of 20 μM suramin show the electrical response 35% of the time, thus VC removal enhances sperm entry. Like the intact eggs, 100% of the naked eggs respond electrically to fertilization after the drug is washed out. Follicle cells that are isolated by calcium magnesium free seawater and then returned to complete seawater release N‐acetylglucosaminidase activity in response to sperm. Thus, these eggs have two blocks to polyspermy that operate in sequence: an early first block resulting from enzymatic modification of the VC by N‐acetylglucosaminidase released primarily from follicle cells and a second electrical block operating at the egg plasma membrane level and requiring sperm–egg fusion. Mol. Reprod. Dev. 48:137‐143, 1997. © 1997 Wiley‐Liss, Inc.

Many ascidians live in clumps and usually release sperm before the eggs. Consequently, eggs are often spawned into dense clouds of sperm. Because fertilization by more than a single sperm is lethal, ascidians have evolved at least two successive blocks to polyspermy: the rapid release of a glycosidase that inhibits sperm binding to the vitelline coat (VC) and a subsequent change in membrane potential that prevents supernumerary sperm-egg fusion. This paper shows that (1) these two blocks can be uncoupled by the use of suramin, and (2) most of the glycosidase appears to be from the follicle cells, which are accessory cells on the outside of the egg VC. Phallusia mammillata eggs initially bind numerous sperm but, after the glycosidase is released, only a few additional sperm bind. Intact eggs in 20 microM suramin release glycosidase, but the electrical response is inhibited; sperm swim actively and bind to the VC but fail to penetrate. Suramin treatment is completely reversible; intact eggs exhibit the electrical response an average of 11 minutes after the drug is washed out. Sperm must contact the follicle cells before passing through the VC; eggs with the VC removed and fertilized in the presence of 20 microM suramin show the electrical response 35% of the time, thus VC removal enhances sperm entry. Like the intact eggs, 100% of the naked eggs respond electrically to fertilization after the drug is washed out. Follicle cells that are isolated by calcium magnesium free seawater and then returned to complete seawater release N-acetylglucosaminidase activity in response to sperm. Thus, these eggs have two blocks to polyspermy that operate in sequence: an early first block resulting from enzymatic modification of the VC by N-acetylglucosaminidase released primarily from follicle cells and a second electrical block operating at the egg plasma membrane level and requiring sperm-egg fusion.

In the sea squirt Phallusia mammillata, fertilization occurs externally in the seawater surrounding the adult. The impact of mercuric ions, one of the most toxic environmental pollutants, was investigated on the functioning of the ionic channels of unfertilized or fertilized egg membrane and on the associated egg morphological events occurring at fertilization. Hg 2+ ions in the submicromolar range affected in a dose-dependent manner (range 0.0625 to 2 µM) the functioning of voltage-dependent Na + and Ca 2+ channels pre-existing in the unfertilized oocyte membrane. These channels are also sequentially involved in the first step of the egg electrical response to fertilization (i.e., the fertilization potential) and were impaired by Hg 2+ . Mercuric ions also strongly inhibited the further operating sperm-dependent channels of this fertilization potential. The two highest concentrations used (1 and 2 µM) greatly reduced the number of eggs that responded electrically and, as a consequence, diminished the percentage of fertilized eggs that resumed meiosis. Also as a consequence of the impairment by Hg 2+ ions of the egg electrical response to fertilization, the electrical block to polyspermy became less efficient and the proportion of polyspermic eggs increased. Finally, the percentage of eggs containing a male pronucleus over the total number of sperm-penetrated eggs statistically diminished in a dose-dependent manner under the effect of Hg 2+ ions.

No abstract