Role of media, protein and energy supplements on maintenance of morphology and DNA-synthesis of small preantral domestic cat follicles during short-term culture.

Small preantral follicles (40 to 90 microns in diameter) from domestic cats were cultured for 10 d using different media (M199 and Dulbecco's MEM) and protein (FCS and BSA) supplements. Culture efficacy was determined by Hoechst 33258 staining and estimation of Brom-desoxyuridine (BrdU)-incorporation into oocytes and granulosa cells. Culture in M199 + FCS and in DMEM + FCS resulted in 21.6% and 38.1%, respectively, of morphologically intact preantral follicles. Adding BSA increased the rate of normal follicles to 51.7% in M199 and to 58.6% in DMEM. Oocytes were found in 40% of the follicles, when DMEM and/or BSA supplementation was used, while M199 with FCS induced acute loss of oocytes in 85% of the follicles. About 10% of the oocytes contained degenerating chromatin. Measurement of BrdU-incorporation during culture allows for quick and effective assessment of follicle viability in vitro. Comparison of M199 and Dulbecco's MEM, both with FCS or BSA and DMEM with or without pyruvate/lactate, indicated that Dulbecco's MEM + BSA without pyruvate and lactate is the best medium for culture of cat follicles. However, further research of suitable medium supplements is needed.