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Masashi Takami1 , Sandra L. Preston1 , Harold R. Behrman1 *
Reproductive Biology Section, Department of Ob/Gyn, Yale University School of Medicine, New Haven, CT USA 1
We showed that cell permeant antioxidants inhibit spontaneous germinal vesicle breakdown (GVBD) of denuded (DO) rat oocytes. The objective of the present studies was to determine whether generation of oxygen radicals with 2,2'-azobis(2-amidinopropane) HCl (AAPH) reverses the inhibition of GVBD produced by IBMX or db-cAMP. DO from preovulatory rat follicles were incubated with or without AAPH in the presence of IBMX (100uM) or db-cAMP (2mM). Spontaneous GVBD of untreated DO incubated for 6 h was 73.8% (N=107), whereas GVBD of DO incubated with IBMX or db-cAMP was 6.8% (N=73) and 1.6% (N=62), respectively. Incubation of DO with AAPH (20mM) significantly (p<0.005) reversed the inhibition of GVBD produced by IBMX or db-cAMP (82.2%GVBD; N=45 and 76.7%GVBD; N=201, respectively). To assess the specificity of AAPH action, DO were incubated with either IBMX or db-cAMP and AAPH in the absence and presence of superoxide dismutase (SOD; 1700u/ml) or a mixture of SOD and catalase (2000u/ml). Only the combination of SOD and catalase significantly (p<0.005) inhibited GVBD induced by AAPH (IBMX+AAPH+SOD=76.9%GVBD; N=52) (IBMX+AAPH+SOD+catalase= 2.8%GVBD; N=72) (db-cAMP+AAPH+SOD=74.7% GVBD; N=87) (db-cAMP+AAPH+SOD+catalase= 19.7% GVBD; N=71). Analysis by high-pressure liquid chromatography did not reveal any change of either IBMX or db-cAMP following incubation with AAPH for 6h. Thus, hydrogen peroxide is the probable candidate that mediates the action of AAPH on induction of GVBD, which may implicate this reactive oxygen species in regulation of the resumption of meiosis. Supported by NIH-HD-35663.
This abstract is being presented on Monday, August 2 at 8:00 AM to 10:15 AM at CUB 2nd Floor Ballroom.