Development of the rat conceptus in vitro and associated changed in components of culture medium

Rat conceptuses of pregnancy day 11 (embryonic age approximately 10-5 days) with embryos within the yolk and amniotic sacs and polar allantoic placental tissues were cultured by the 'New system' utilizing homologous serum in 60 ml roller bottles with 20% O2, 5% CO2 and 75% N2 gas environment (five conceptuses/10 ml serum). Embryonic growth and development was assessed by external morphology and somite counts, and by measurement of DNA, RNA and protein contents. The embryonic growth was considerably retarded beyond 24 h when culture was extended to 48 h. During continuous culture of conceptuses for 48 h, the partial pressure of O2 in the medium decreased gradually with simultaneous increase of pCO2 and decline of pH values. Glucose level was depleted considerably and small quantities of urea, uric acid and creatinine accumulated in the medium. The concentrations of electrolytes (Na+, K+, Cl-) and total Ca and P, and lactic dehydrogenase activity in the medium increased during the latter part of the 48 h culture period. The growth and differentiation of embryos were significantly improved by transferring conceptuses to fresh serum medium with 40% O2, 5% CO2 and 55% N2 gas phase after 22-24 h culture and continuing the culture for an additional 24 h. Embryos grown in vitro for 48 h developed 39 somites and showed a tenfold increase in DNA content and a five- to seven-fold increase in RNA and protein contents, over initial values.