Spinal cord regeneration in Xenopus laevis

Abstract

Here we present a protocol for the husbandry of Xenopus laevis tadpoles and froglets, and procedures to study spinal cord regeneration. This includes methods to induce spinal cord injury (SCI); DNA and morpholino electroporation for genetic studies; in vivo imaging for cell analysis; a swimming test to measure functional recovery; and a convenient model for screening for new compounds that promote neural regeneration. These protocols establish X. laevis as a unique model organism for understanding spinal cord regeneration by comparing regenerative and nonregenerative stages. This protocol can be used to understand the molecular and cellular mechanisms involved in nervous system regeneration, including neural stem and progenitor cell (NSPC) proliferation and neurogenesis, extrinsic and intrinsic mechanisms involved in axon regeneration, glial response and scar formation, and trophic factors. For experienced personnel, husbandry takes 1-2 months; SCI can be achieved in 5-15 min; and swimming recovery takes 20-30 d.

Temporally distinct transcriptional regulation of myocyte dedifferentiation and Myofiber growth during muscle regeneration

“…In fact, the GO term “regulation of gene expression, epigenetic” (GO:0040029) was significantly enriched in profile F. All this suggested that at least a subset of these factors might be mechanistically involved in allowing post-mitotic myocytes to dedifferentiate into proliferative myoblasts and we thus decided to focus on transcription factors and epigenetic regulators. We further hypothesized that early-activated genes would play a particularly important role in the reprogramming and dedifferentiation of injured myocytes, and tested the hypothesis using gene knockdown experiments using antisense morpholino oligonucleotide (MO), a technique widely used to perform tissue-specific knockdown experiments in adult zebrafish [ 20 – 22 ] and other model organisms such as adult axolotls [ 23 ], xenopus larvae [ 24 ] or chick [ 25 ] and mouse [ 26 ] embryos. Briefly, lissamine-tagged MOs are injected and then electroporated into the LR muscle 3 h prior to myectomy injury (Figs.…”

The brain is required for normal muscle and nerve patterning during early Xenopus development

“…6a ) experiments were performed on stage-25 embryos. To prevent regeneration 66 , one segment of the spinal cord, sizing 50–100 μm in length, was completely removed by using two forceps with super-fine tips (Dumont #5ST, FST 11252-00). SC segments were removed at cervical level, immediately posterior to the hindbrain, and taking special care of minimizing the damage on the most rostral myotomes.…”

Three‐dimensional reconstruction of the cranial and anterior spinal nerves in early tadpoles of Xenopus laevis (Pipidae, Anura)

“…Since the mid‐20th century, the African clawed frog, Xenopus laevis , has become one of the most widely used model organisms in biology (Harland & Grainger, ; Porro & Richards, ). Especially in neurobiology, Xenopus embryos, tadpoles, and adults are used as models in pathological, developmental, physiological, and behavioral studies (Cannatella & De Sa, ; Cervino, Paz, & Frontera, ; Cline & Kelly, ; Dong et al, ; Edwards‐Faret et al, ; Frankenhaeuser & Huxley, ; Gouchie, Roberts, & Wassersug, ; Katz, Potel, & Wassersug, ; Lee‐Liu, Méndez‐Olivos, Muñoz, & Larraín, ; McKeown, Sharma, Sharipov, Shen, & Cline, ; Moreno, Tapia, & Larrain, ; Pieper, Eagleson, Wosniok, & Schlosser, ; Pratt & Khakhalin, ; Roberts, Walford, Soffe, & Yoshida, ; Schlosser & Northcutt, ; Simmons, Costa, & Gerstein, ; Wassersug & Hessler, ; Young & Poo, ). It is therefore surprising, that only one study (Paterson, ) on the anatomy of the cranial nerves of X. laevis tadpoles exists.…”