Biochemical and Physiological Factors Affecting Early Stages of Fertilization in the Bovine
Siegel, Mark Steven
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Permalink
https://hdl.handle.net/2142/71719
Description
Title
Biochemical and Physiological Factors Affecting Early Stages of Fertilization in the Bovine
Author(s)
Siegel, Mark Steven
Issue Date
1982
Department of Study
Dairy Science
Discipline
Dairy Science
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Animal Physiology
Abstract
The purpose of this study was to evaluate a number of physiological and biochemical factors included in sperm binding to and penetration of bovine oocytes. The first part included evaluating conditions required to maximize sperm binding to and penetration in vitro; the second involved differentiating acrosin from non-acrosin proteases in bovine acrosomes. These proteases are likely to be involved in sperm binding and penetration.
The effect of Brinster's Medium for Ova Culture (BMOC) with or without bovine serum albumin (BSA) and dialyzed or undialyzed uterine or oviduct fluids on sperm acrosomal morphology and sperm binding to and penetration of oocytes were tested. Preincubation (1 x 10('9) sperm/ml) of sperm for at least 30 min. in most media enhanced the number of acrosome reacted sperm and sperm bound to and penetrating oocytes, indicating a positive effect of high sperm concentration during preincubation. At 2 x 10('5) sperm/ml, maximal penetration required at least 60 min. of incubation; while sperm binding increased over the 240 min. incubation period. Absence of BSA from BMOC or dialysis of tract fluids againt Tris, altered the magnitude and kinetic patterns of these events. No apparent differences between undialyzed tract fluids and BMOC containing BSA were found. BSA, inorganic ions and in oviduct fluids a non-steroidal low molecular weight component increased the number of acrosome reacted sperm and sperm bound to and penetrating oocytes. Significant positive correlations were observed beween acrosome reacted sperm and sperm binding and penetrating oocytes.
Acrosin and non-acrosin proteolytic activity were distinguished by differences in biochemical properties. Using gelatin membrane slides and a newly developed agar-gelatin assay for proteolytic digestion, it was shown that calcium chloride inhibited proteolytic activity in whole sperm and acrosomal extracts. Protease inhibitors only mildly inhibited proteolytic activity. Calcium chloride stimulated, while protease inhibitors markedly inhibited acrosin activity. These observations indicate the presence of non-acrosin proteases representing at least 50% of the total proteolytic activity in bovine acrosomes.
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