The objective of this proposal is to elucidate the biochemical mechanism(s) that regulate mammalian sperm motility before and during ejaculation. Bovine caudal epididymal (CE) sperm are quiescent in CE fluid; dilution of CE fluid or an elevation of its pH results in the initiation of sperm motility. The cytoplasmic modulator of this motility quiescence and initiation appears to be the intracellular pH (pHi).
The first aim of this research is to measure the change of pHi of bovine CE sperm, concomitant with motility initiation, using a fluorescent, pH-sensitive chromophore, carboxyfluorescein. The mechanism of action of pHi upon motility will be investigated in a permeabilized bovine CE sperm model. The involvement of pH and selected ionic components in motility regulation will be defined. This model is ideal for testing our working hypothesis: that the changes in pHi act directly upon the dynein ATPase to modulate motility. The effects of ionic composition and certain motility inhibitors upon the pH-dependence of the ATPase activity will also be determined. After evaluation of the model system, the dynein ATPase from bovine CE sperm will be isolated and purified. This enzyme will be characterized physically and its kinetic properties will be defined. The effects of pH, ionic composition and inhibitors upon ATPase kinetics and upon the interaction of the ATPase with microtubules and extracted flagella will be studied. The existence of cyclic AMP-dependent phosphorylation of the purified dynein ATPase and of a role for pH in this process will be investigated. The possibility that calmodulin (CaM) is a component of, or interacts with the ATPase in a calcium- and a pH-dependent manner will be tested. The interaction of pH, cyclic AMP-dependent phosphorylation and Ca2+/CaM upon the motility of the permeabilized sperm model will also be evaluated. The understanding gained from these studies will be applied to human and nonhuman primate sperm with emphasis upon male contraception and certain cases of male infertility.
| Carr, D W; Acott, T S (1990) The phosphorylation of a putative sperm microtubule-associated protein 2 (MAP2) is uniquely sensitive to regulation. Biol Reprod 43:795-805 |
| Carr, D W; Usselman, M C; Acott, T S (1985) Effects of pH, lactate, and viscoelastic drag on sperm motility: a species comparison. Biol Reprod 33:588-95 |
