Voltage clamp studies of fertilization in sea urchin eggs. I. Effect of clamped membrane potential on sperm entry, activation, and development.
To analyze the role of the activation potential (a positive shift of the membrane potential which occurs following sperm attachment) in fertilization and development of the sea urchin egg, unfertilized Lytechinus variegatus eggs were voltage clamped at membrane potentials (Em) from +20 to -90 mV, and then inseminated. Either a fast two electrode voltage clamp, or a single electrode switched voltage clamp was used. The clamp was maintained for 3 to 15 min after initiation of a conductance increase. At Em more positive than +18 mV, even though many sperm may attach, the egg remains completely inert (Jaffe, Nature (London) 261, 68-71, 1976). At Em from +17 to -90 mV, all inseminated eggs elevate normal fertilization envelopes, although substantially increased concentrations of sperm are required at Em from +17 to +12 mV. Whether cleavage occurs depends on the clamped Em. When clamped at Em from +17 to -25 mV, 100% of activated eggs cleave. However, when clamped at Em from -26 to -75 mV the percentage of activated eggs which cleave progressively decreases. At clamped Em between -76 and -90 mV, none of the activated eggs cleave. All monospermic voltage clamped eggs that cleave develop to normal swimming blastulae. In all eggs that fail to cleave (clamped at Em more negative than -30 mV), sperm penetration is blocked, the sperm is lifted off the egg surface as the fertilization envelope rises, and a sperm aster never forms. Preventing formation of the fertilization envelope by prior disruption of the vitelline layer with dithiothreitol does not promote entry of the sperm. In conclusion, preventing the depolarization normally associated with fertilization suppresses sperm entry in the sea urchin egg, yet activation proceeds. Present evidence suggests an effect of the electrical field across the plasma membrane in suppressing sperm entry.
D000200:Action Potentials; D000818:Animals; D001755:Blastocyst / Q000502:physiology; D002462:Cell Membrane / Q000502:physiology; D004553:Electric Conductivity; D005260:Female; D005306:Fertilization; D008297:Male; D008564:Membrane Potentials; D010063:Ovum / Q000502:physiology*; D012617:Sea Urchins; D013084:Sperm-Ovum Interactions; D013094:Spermatozoa / Q000502:physiology*