UC Irvine

For centuries regeneration was thought to be unattainable in the spinal cord, but recent tools and advancements imbue optimism. This dissertation serves to guide the field in using a genetically-labeled mouse for studying CST regeneration, and to assess wiring of CST regeneration that results from PTEN deletion.

Mice designed to have yellow-fluorescent protein (YFP) expressed by the CST (CST-YFP mice) were proposed to be useful for studying CST regeneration. We show by anterograde labeling, anterograde degeneration, and retrograde labeling that some YFP-labeled axons in the spinal cord are not of the CST. We further demonstrate that dense YFP-labeled arbors impede tracing individual YFP-labeled axons such as could be used to confirm CST origins. We also found that YFP fluorescence in these mice is faint for clearing-based 3D imaging as could be used for screening regenerative responses. These results overall limit the utility of CST-YFP mice for studying CST regeneration.

Deletion of PTEN has been demonstrated to enhance CST regeneration after spinal cord injury. For regeneration to be functionally meaningful, it is critical that the axons form synapses. We assessed whether CST axons that regenerated due to PTEN deletion form synapses at the level of electron microscopy. In PTEN-floxed mice that were injected with AAV-Cre near birth to delete PTEN and as adults given a complete thoracic crush lesion, we identified synaptic structures of regenerated CST axons caudal to the lesion. This result indicates that manipulation of the PTEN pathway can produce a regenerative response that may contribute to caudal circuitry.

Toward understanding regeneration that may contribute to circuitry of motor function, the laterality of CST regenerative growth that results from PTEN deletion was assessed. PTEN-floxed mice were injected with AAV-Cre near birth and as adults given a dorsal hemisection lesion in the low thoracic spinal cord. By anterograde tracing, CST laterality in control mice was found to be predominantly contralateral to the injected cortex. Laterality of regenerative growth caudal to the lesion was found to be variable and sometimes predominantly on the ipsilateral side. These results indicate that regenerative growth does not follow the contralateral rule of the non-injured CST.