Phosphodiesterase activity of mouse sperm incubated under conditions that modulate fertilizing potential in vitro

Abstract

Cyclic AMP has been implicated as a regulator of capacitation, but the control of its metabolism in sperm remains obscure. A recent study of mouse sperm has shown capacitation-related changes in the activities of both adenylate cyclase, which increased during incubation, and cyclic nucleotide phosphodiesterase, which decreased. The present study was conducted to extend these observations by measuring phosphodiesterase activity in sperm incubated in media with modified calcium and/or glucose content, conditions known to modulate fertilizing ability. Phosphodiesterase activity of sequential sperm samples, taken first when sperm are essentially uncapacitated and then when they are either partially or completely capacitated, decreased with time under all conditions, and in each case the greater fall in activity was seen in the medium that would support the greater change in fertilizing ability of the sperm population. Sperm washed by centrifugation to remove epididymal fluid also displayed a reduction in phosphodiesterase activity with time. The medium surrounding the sperm contained about half of the total phosphodiesterase activity, as well as 5'-nucleotidase and adenosine deaminase. The crude enzyme preparation showed complex kinetic behavior when assayed over a range of cAMP concentrations, but the reduction in activity with time was seen at all substrate levels. The observed changes in phosphodiesterase activity, together with the increased adenylate cyclase activity seen under these sperm incubation conditions, would increase cAMP availability with time, thus providing further evidence for a fundamental role for cAMP in controlling the events of capacitation.

Enzymes of adenosine metabolism in mouse sperm suspensions

“…Adenosine was the only product of 5'-AMP metabolism in washed spermatozoa, extending our previous observations of the products of cAMP metabolism in such suspensions, in which a limited capacity for adenosine metabolism was noted (Monks & Fraser, 1987). In the present study, a considerable difference in the ability to metabolize adenosine was found between unwashed and washed sperm suspensions.…”

“…To investigate whether 5'-nucleotidase is affected by the functional state of the sperm population, its activity was measured in sperm populations incubated under conditions known to modulate both capacitation and the activities of the enzymes adenylate cyclase and phosphodiesterase (Stein & Fraser, 1984;Monks et al, 1986; Monks & Fraser, 1987). Initially, a single sperm suspension was prepared from 6 3) and the data were subjected to an analysis of variance using the statistical computer package GENSTAT (Lawes Agricultural Trust, Rothamsted Experimental Station; 1980).…”

“…(Stein & Fraser, 1984;Monks et al, 1986;Monks & Fraser, 1987 Levin & Bodansky, 1966;Edwards et al, 1982;Camici et ai, 1985). The high levels of these enzymes seen in the medium surrounding spermatozoa are probably derived from epididymal plasma; 5'-nucleotidase has been reported both in epididymal (Jones, 1978) and seminal (Levin & Bodansky, 1966) fluids.…”

“…A well-defined system for capacitation of epididymal mouse spermatozoa and fertilization of homologous eggs in vitro has allowed investi¬ gation of a number of compounds which influence these processes (Fraser, 1983). Cyclic AMP (cAMP) appears to play a key role in modulating many capacitation-related events (Tash & Means, 1983); observation of increased adenylate cyclase and decreased phosphodiesterase activities during capacitation of mouse spermatozoa in vitro support this view (Stein & Fraser, 1984;Monks et ai, 1986;Monks & Fraser, 1987). However, the control of sperm cAMP production remains obscure since, unlike the enzyme of somatic cells, sperm adenylate cyclase is not hormonecontrolled (Hildebrandt et ai, 1985).…”

“…In contrast, calcium-free medium supports only partial capacitation. Such modified incubation conditions have been shown to modulate both capacitation and the activities of the enzymes of cAMP metabolism, adenylate cyclase and phosphodiesterase (Stein & Fraser, 1984;Monks et al, 1986;Monks & Fraser, 1987). Since epididymal mouse sperm suspensions contain derivatives of epididymal fluid in addition to intrinsic sperm components, enzyme activity was determined in unwashed and washed sperm suspensions.…”