Effect of roscovitine pretreatment on in vitro maturation of oocytes and their subsequent developmental after chemical activation in dromedary camel (Camelus dromedarius).

Studies were conducted to evaluate an optimal concentration of roscovitine needed to maintain abattoir origin oocytes at germinal vesicle stage in experiment 1 and their subsequent maturation and developmental competence after chemical activation in experiments 2 and 3, respectively. The cumulus-oocyte complexes (COCs) aspirated from ovaries collected from a local slaughterhouse were cultured in TCM-199 based pre-maturation medium supplemented with 25, 50 or 75 μM roscovitine, depending on the experimental group. After 24 h, the COCs were denuded of cumulus, fixed and stained with aceto-orcein and examined for their nuclear status. They were classified as germinal vesicle, diakinesis, metaphase-I, metaphase-II and those with degenerated, fragmented, scattered, activated or without visible chromatin as others. In experiment 2, the COCs pre-matured in media supplemented with 50 μM roscovitine for 24 h were washed and kept for in vitro maturation along with another group of freshly collected COCs for 30 h. All the oocytes were fixed and stained to evaluate their nuclear status as described above. In experiment 3, all mature oocytes obtained from the COCs pre-matured in media supplemented with 50 μM roscovitine and those obtained from freshly collected group were activated by 5 mM ionomycin. Activated oocytes were cultured in embryo culture medium for a period of 7 days to evaluate their developmental potential. The proportion of oocytes at GV stage in the group pre-matured in media with 50 μM-was significantly (P < 0.01) higher when compared with the group having 25 μM of roscovitine. No difference was found in the proportion of GV stage oocytes in this group when compared with the freshly collected COCs. None of the oocytes reached to M-II stage in any of the three treatment groups. In experiment 2, no difference was observed in the proportion of oocytes reaching M-II stage between the groups after 30 h of in vitro culture; however, higher proportion of oocytes (P < 0.05) were classified as others in the pre-maturation group when compared with the group having freshly collected oocytes. In experiment 3, no difference was observed in the proportion of oocytes cleaving and those developing to the blastocyst stage between the pre-matured and freshly matured groups. In conclusion, the present study, for the first time, demonstrates the possible use of roscovitine as a meiotic inhibitor for camel oocytes. Keeping in view the ability of these oocytes to mature and develop to the blastocyst stage at par with the fresh oocytes, more flexible schedules for maturation and manipulation of such oocytes could be developed.