The Ambystoma mexicanum, commonly known as the axolotl has the impressive ability to regenerate multiple body parts. However the molecular mechanisms underlying regeneration remain poorly understood at the molecular level. Here we describe a method creating a spinal cord injury in the axolotl and for manipulating microRNAs in vivo in the axolotl spinal cord. This technique can be adapted for altering gene expression using small moleucles like morpholinos or siRNAs into other tissues in the axolotl including limb and tail blastemas. The ability to spatially and temporally test gene function in vivo is an important technique that will help to elucidate the hierarchy of gene function during regeneration.