Publication Date

2013-09-11

Availability

Open access

Embargo Period

2013-09-11

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PHD)

Department

Neuroscience (Medicine)

Date of Defense

2013-08-09

First Committee Member

Christine K. Thomas

Second Committee Member

Vance Lemmon

Third Committee Member

Kenneth J. Muller

Fourth Committee Member

Brian R. Noga

Fifth Committee Member

Ranu Jung

Abstract

Motoneuron death occurs with spinal cord injury, neuromuscular disease and age, resulting in muscle denervation. Although spared axons can reinnervate nearby muscle fibers, death of an entire motoneuron pool prevents muscle reinnervation by host motoneurons. The muscle will remain denervated completely. To reduce denervation-induced muscle atrophy, we have previously transplanted embryonic ventral spinal cord cells into a peripheral nerve near denervated muscles. This strategy shortens the distance axons have to regenerate to reinnervate muscles. Reinnervation reduced muscle atrophy but some muscle fibers remained denervated. Since neural activity is required for the survival of embryonic neurons and promotes axon growth, the aim of this study was to test whether brief electrical stimulation of embryonic neurons transplanted into peripheral nerve changes motoneuron survival, axon regeneration, motor unit formation and muscle properties 10 weeks after transplantation. The sciatic nerve was transected to induce muscle denervation, thereby mimicking the muscle consequences of motoneuron death. One week later, 200,000 embryonic day 14-15 ventral spinal cord cells, purified for motoneurons, were injected into the tibial nerve of adult Fischer rats close to gastrocnemii muscles. The transplanted neurons were the only source of neurons for muscle reinnervation because the proximal stump of the sciatic nerve had been tied to a hip muscle. Immediately after transplantation, the cells were stimulated for up to one hour using various protocols designed to determine the differential effects of pulse number, stimulation frequency, pattern and duration of stimulation on outcomes. Low frequency stimulation (1 Hz) for one hour resulted in more myelinated axons in the tibial nerve compared to intermittent stimulation, 3 minutes of stimulation or no stimulation. Continuous stimulation for one hour resulted in higher numbers of reinnervated motor units than 3 minutes of stimulation or no stimulation. Higher motor unit numbers resulted when the stimulation intervention included shorter rest times. Muscles with higher motor unit counts were stronger. Even without electrical stimulation, cell transplantation itself and reinnervation reduced muscle atrophy significantly. Muscles reinnervated by embryonic motoneurons could be excited by nerve stimulation whereas denervated muscles were unresponsive. Higher forces, relative to maximum, were attained in response to 15-50 Hz stimulation in reinnervated versus uninjured muscles. Reinnervated muscles were resistance to fatigue. The ability of embryonic neurons to respond to brief electrical stimulation, in vivo, by forming more motor units makes these reinnervated muscles more useful for production of functionally meaningful movements using patterned electrical stimulation.

Keywords

muscle denervation; cell transplantation; motoneuron survival; axon regeneration; muscle reinnervation; motor unit

Share

COinS