144 Two-step in vitro maturation from small follicles versus IVM from large follicles and chemical oocyte activation in alpaca (Vicugna pacos)
D. Vargas-Donayre A , Z. Bravo A , H. Zeña A and M. Valdivia AA Laboratory of Reproductive Physiology, Biological Sciences Faculty, Universidad Nacional Mayor de San Marcos, Lima, Peru
Reproduction, Fertility and Development 35(2) 200-200 https://doi.org/10.1071/RDv35n2Ab144
Published: 5 December 2022
© 2023 The Author(s) (or their employer(s)). Published by CSIRO Publishing on behalf of the IETS
The main source of oocytes for the production of alpaca embryos in vitro is from large follicles, as the population features better developmental competence. Therefore, oocytes from smaller follicles are a potential source of achieving a higher number of embryos for use in reproductive biotechnologies. In the present report, we examined a two-step in vitro maturation (IVM) method, which included a prematuration culture period for enhancing the competence of oocytes from small follicles. Ovaries (n = 130) collected in the municipal slaughterhouse of Huancavelica were transported to Lima, and oocytes (n = 159) were harvested 20 to 24 h postmortem. Oocytes from large follicles ([LF] 3–7 mm) were collected by aspiration and cultured according to the standard one-step IVM method for 32 h in a humidified atmosphere containing 5% CO2 at 38.5°C. Oocytes retrieved from small follicles ([SF] < 3 mm) by slicing ovaries were cultured in medium supplemented with meiotic arrestors (C-type natriuretic peptide [CNP] and 3-isobutyl-1-methylxanthine [IBMX]) (Step 1) before culture in standard IVM medium (Step 2). Nuclear maturation was assessed by the presence of the first polar body, and parthenogenetic embryos were obtained by chemical activation with ionomycin and 6-dimethylaminopurine (6-DMAP). Presumptive embryos were in vitro cultured in Global Total medium (GlobalMed Group) in 5% CO2, 5% O2, and 90% N2 at 38.5°C, and the cleavage and blastocyst rates of in vitro culture were registered after 2 and 7 days. Mean nuclear maturation rate of oocytes from LF was similar to that of oocytes from SF undergoing IVM in the two-step method (P > 0.05) (56.9 ± 19.6% vs 55.8 ± 17.4%, respectively). There was no difference in mean cleavage rate and blastocyst formation frequency between the two IVM groups (P > 0.05) (30.0 ± 26.4%, 14.2 ± 15.1% vs 33.4 ± 15.7%, 2.1 ± 3.6%, in the one-step and two-step groups, respectively). In the present results, we demonstrated that two-step IVM allows oocytes from small follicles to acquire the in vitro developmental competence of oocytes from large follicles cultured in a standard one-step IVM method.
This research was supported by PROCIENCIA – CONCYTEC-Proyecto Investigación Básica, 2019-01-Convenio N° 428-2019-FONDECYT.
