Influence of age of in vitro matured oocytes and electrical stimulations on the development of cattle nuclear transfer embryos

Theriogenology

39:292,

1993

INFLUENCE OF AGE OF IN VITRO MATURED OOCYTES AND ELECTRICAL STIMULATIONS ON THE DEVELOPMENT OF CATTLE NUCLEAR TRANSFER EMBRYOS V.H.RAO, P.CHESNE, J.P.RENARD and Y.HEYMAN Unite de Biologie du Developpement. INRA, Jouy-en-Josas, FRANCE. In our efforts to improve blastocyst formation in vitro by nuclear transfer embryos (NTE) produced from in vitro matured(IVM) oocytes we have been testing the influence of age of oocytes and different electrical stimulation pattems.Some of the interesting observations are presented here. Procedures for IVM of oocytes and nuclear transfer were described earlier (Heyman et al., 1992,12th ICAR,The Hague,vol2:688).Recipient oocytes were decoronized at 26h after the start of IVM and allowed to age at 39’C (Table 1). NTE were electrofused in 0.3M manitol by a d/c pulse of 2KV/cm.Multiple stimulations when used consisted of additional 54 or 3 pulses of O.SKV/cm administered at 1/2h intervals after the fusion stimulus.Between stimulations NTE were kept in B2+10%FCS at 390C in 5%CO2 in air.Subsequently fused NTE and IVF embryos from the same batch of oocytes were cultured on oviducal cell monolayers and evaluated for blastocyst formation on day 7. Table 1: k

--,--\--, 4/23( 17)a

l/24(4)‘-

0/12(0)b 0/25(0)b (25v

X2;Pday 90) from such blastocysts.Physiological changes between 32 and 36h of age apparently induce adequate reprogramming of donor nuclei and improve blastocyst formation at the same time as control IVF embryosDecline in cleavage rate but not blastocyst formation by 40h oocytes may indicate the onset of deleterious effects of ageing. We conclude that under the experimental conditions used there is an optimum window between 32 and 40h of age for better development of NTE produced from IVM oocytes. VHR is supported by APAU and DBT,Govt.of India.

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Copyright 0 1993 Butterworth-Heinemann