Assessment of double ovulation to halve Xenopus laevis use for eggs

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The African claw-toed frog, Xenopus laevis, is used in many laboratories because of its eggs. Unlike the eggs of many other species, frog eggs (spawn) are laid in large numbers and develop outside the other in ordinary pond water. This makes them ideal for observing embryonic development and doing experiments such as testing the consequences of a tissue graft or injecting small amounts of DNA (genetic material) to find out how genes work in a living organism. Xenopus frogs are particularly useful because they normally live in water and so can be kept easily in water-filled tanks all year round. To get them to lay eggs (ovulate) researchers inject the females with a hormone. If injected in the evening, the frogs lay eggs into the tank the following day. (This used to be used clinically as a pregnancy test since the hormone is elevated in a pregnant woman's urine.) Currently, labs using Xenopus ovulate frogs and then rest them for three months ("single ovulation"). However, although not routine, experiments have shown that a frog can be ovulated weekly over several weeks with only gradual decline in the number of eggs produced. This may reflect breeding in the wild where frogs have a limited breeding (rainy) season. We have conducted some pilot experiments to show that ovulating females two times, a week apart ("double ovulation") with three month's rest before this is repeated, yields similar numbers per ovulation that are not obviously less than single ovulation. In this project we will test this double ovulation regime to see if yield numbers are reliably maintained. Importantly, to ensure that the procedure does not cause additional stress to the animals, either immediately or over time, we will assess stress levels by tracking egg numbers and quality (which are both known to deteriorate for sick or otherwise stressed animals) and measure levels of a stress-hormone (corticosterone) that is released by the animals under stressful conditions. If these measures show that egg yields hold up with little or no increase in stress or harm to the animals, they will allow labs to use half as many animals as they do now. Worldwide, Xenopus users have a strong community and new methods are rapidly shared. There are no barriers to take-up of double ovulation, and we plan to disseminate our results using proven channels of the Xenbase website, national Stock Centres in the UK and USA and through training courses, as well as publication of our data and methods in journals and conferences.