A double ovulation protocol for Xenopus laevis produces doubled fertilisation yield and moderately transiently elevated corticosterone levels without loss of egg quality

Abstract

AbstractThe African claw-toed frog,Xenopus laevis, is a well-established laboratory model for the biology of vertebrate fertilisation and development. For ovulation,X. laevisfemales are usually injected with chorionic gonadotropin, whereupon they lay typically hundreds to thousands of eggs in a day. Animals are re-used after being rested for a minimum of three months,. The literature suggests that adult females can lay much larger numbers of eggs in a short period. Here, we compared the standard “single ovulation” protocol with a “double ovulation” protocol, in which females were ovulated, then re-ovulated after seven days and then rested for three months before re-use. We quantified egg number, fertilisation rate (development to cleavage stage), and corticosterone secretion rate (a measure of stress response) over seven 3-month cycles. We found no differences in egg number-per-ovulation or fertilisation, and no long-term changes in any measures over the trial period. Corticosterone secretion was elevated by ovulation, similarly for the single ovulation as for the first ovulation in the double-ovulation protocol, but more highly for the second ovulation (to a level comparable to that seen following shipment). However, both groups exhibited the same baseline secretion rates by the time of the subsequent cycle. Double ovulation is thus transiently more stressful/demanding than single ovulation but within levels routinely experienced by laboratoryX. laevis. Noting that “stress hormone” corticosterone/cortisol secretion is linked to physiological processes, such as ovulation, that are not necessarily harmful to the individual, we suggest that the benefits of a doubling in egg yield-per-cycle per animal without loss of egg quality or signs of acute or long-term harm may outweigh the relatively modest and transient corticosterone elevation we observed. The double ovulation protocol therefore represents a potential new standard practice for promoting the “3Rs” (animal use reduction, refinement and replacement) mission for Xenopus research.