(145) Fresh and Vitrified Oocytes from Young and Aged CD-1 Mice Produces Comparable Mouse Pup Survival Rates

India D. Napier¹; Elmer Umana²; Tony E Chavarria²; Gong Du²; Alexandre N Viana²; Susan E Erdman²

1. Department of Comparative Pathobiology, Cummings School of Veterinary Medicine, Grafton, MA, USA 

2. Division of Comparative Medicine, Massachusetts Institute of Technology, Cambridge, MA, USA

Abstract Text:

Vitrification is a rapid freezing technique that allows preservation of gametes, such as oocytes, for future use. As female mice age, their oocytes undergo biochemical and physiological changes that can impact their developmental competence and overall quality. These age-associated changes raise questions about whether aged mice are appropriate for developmental studies, and the effects of vitrification on aged oocyte viability for subsequent embryo development. In this study, we investigated the developmental outcomes of vitrified-thawed oocytes collected from young (7-11 weeks of age) and aged (17-21 weeks of age) super-ovulated CD-1 mice. Fresh (never cryopreserved) oocytes from young and aged mice served as control groups. Experiments were performed in duplicate using fresh or vitrified oocytes from both age groups. Young or aged murine oocytes were vitrified then stored at -195°C in liquid nitrogen. One hour later, vitrified oocytes were thawed, and the number of oocytes collected and identified as viable one hour after thaw were counted to determine recovery and survival rates, respectively. Then, fresh or thawed oocytes (collected from young or aged mice) were incubated overnight for in vitro fertilization using spermatozoa collected from 3-5-month-old, CD-1 proven breeder male mice. Sixteen to eighteen hours later, the number of 2-cell embryos were counted to calculate cleavage rates. Same-day surgical embryo transfers were performed using 2-4-month-old, pseudo-pregnant CD-1 recipients. Single-housed, pregnant dams were monitored until their pups were weaned between 21-23 days of age. The mean numbers of retrieved oocytes used for vitrification studies were similar (P > 0.05) between 7-11-week-old (294.5 ± 15.5) and 17-21-week-old (314 ± 71) super-ovulated mice. Similar (P > 0.05) numbers of fresh oocytes from young (336 ± 7) and aged (230 ± 40.5) mice were used for study. There were no significant differences (P > 0.05) in the average recovery rates of thawed oocytes for young (88.5% ± 1.72%) and aged (92.8% ± 2.11%) mice. The mean survival rates of vitrified-thawed oocytes were similar (P > 0.05) between young (80.7% ± 7.48%) and aged (73.1% ± 6.03%) mouse groups. There was no significant difference (P > 0.05) in the percentages of 2-cell embryos when using vitrified young (45.1% ± 5.70%) or aged (62.4% ± 4.45%) oocyte groups. Cleavage rates were also similar (P > 0.05) between fresh oocytes were used from 7-11-week-old (90.5% ± 0.50%) and 17-21-week-old (82.2% ± 1.10%) mice. The average numbers of weaned pups per dam were similar (P > 0.05) between the vitrified young (7.50 ± 1.22) and the aged (8.10 ± 0.94) mouse groups. Similar results (P > 0.05) were observed for fresh oocytes from 7-11-week-old (4.3 ± 1.0) and 17-21-week-old (7.3 ± 1.7) mice. All weaned pups aged to developmentally normal adult mice. Overall, female reproductive age had no impact on post-thaw survival rates of vitrified oocytes, 2-cell embryo cleavage rates, and weaned pup numbers in CD-1 mice. Thus, young and aged mice have comparable oocyte quality, indicating that older mice remain valuable models for reproductive and vitrification studies.