Corticosterone Is Essential for Survival Through Frog Metamorphosis.

Thyroid hormone (TH) is required for frog metamorphosis, and corticosterone (CORT) increases TH signaling to accelerate metamorphic progression. However, a requirement for CORT in metamorphosis has been difficult to assess prior to the recent development of gene-editing technologies. We addressed this long-standing question using transcription activator-like effector nuclease (TALEN) gene disruption to knock out proopiomelanocortin (pomc) and disrupt CORT production in Xenopus tropicalis. As expected, mutant tadpoles had a reduced peak of plasma CORT at metamorphosis with correspondingly reduced expression of the CORT-response gene Usher syndrome type-1G (ush1g). Mutants had reduced rates of growth and development and exhibited lower expression levels of 2 TH response genes, Krüppel-like factor 9 (klf9) and TH receptor ß (thrb). In response to exogenous TH, mutants had reduced TH response gene induction and slower morphological change. Importantly, death invariably occurred during tail resorption, unless rescued by exogenous CORT and, remarkably, by exogenous TH. The ability of exogenous TH by itself to overcome death in pomc mutants indicates that the CORT-dependent increase in TH signaling may ensure functional organ transformation required for survival through metamorphosis and/or may shorten the nonfeeding metamorphic transition to avoid lethal inanition.

Xenopus Tadpole Tissue Harvest.

The procedures described here apply to Xenopus tadpoles from the beginning of feeding through the major changes of metamorphosis and are appropriate for downstream postoperative snap freezing for molecular analysis, fixation for histological analysis, and sterile organ culture. To the uninitiated, the most difficult aspects of tadpole tissue dissections are likely knowing the appearance and location of organs, and the difficulty manipulating and holding tadpoles in place to carry out the oftentimes fine and precise dissections. Therefore, images and stepwise instructions are given for the harvest of external organs (tail, head, eyes, tail skin, back skin, gills, thymus, hind limbs, forelimbs) and peritoneal organs (intestine, pancreas, liver, spleen, lungs, fat bodies, kidney/gonad complex), as well as brain, heart, and blood. Dissections are typically done under a dissection stereomicroscope, and two pairs of fine straight forceps, one pair of fine curved forceps, and one pair of microdissection scissors are sufficient for most tissue harvests.