MOTOR FUNCTION RESTORATION: THERAPEUTIC APPROACHES TO DENERVATION PATHOLOGIES

Abstract

Completely denervated muscles, as seen after motoneuron diseases such as amyotrophic lateral sclerosis, or after peripheral nerve and spinal cord injuries, do not respond to electrical stimulation and are thus not amenable for neural prostheses. Restoration of motor function requires either a timely return of innervation or an alternate approach to control contraction despite denervation. To do so, we first focused on transplantation of embryonic stem cell-derived motoneurons after prolonged denervation and followed by developing a model to optically control muscle contraction when reinnervation is not possible. For transplantation, embryonic stem cell-derived motoneurons were injected into the distal segment of the transected murine tibial nerve after prolonged denervation ranging from one to eight weeks. Success of reinnervation by transplanted motoneurons partially restored muscle force but decreased as the denervation period lengthened. Furthermore, teratocarcinomas developed from a residual multipotent cell population found despite neural differentiation. Tumours could be prevented by pre-transplant treatment with a DNA alkylating agent without compromising innervation from transplanted motoneurons. For optical control of contraction, we generated a transgenic mouse model expressing channelrhodopsin-2/H134R along the sarcolemma and T-tubules of skeletal muscle fibres. This model allowed optical control of contractions with forces comparable to those generated by nerve stimulation. Varying light pulse intensity, pulse duration and pulse frequency permitted to reproduce normal force generation patterns. Finally, we encountered a series of transplants that generated self-organized circuits resulting in spontaneous muscle contractions and evoked episodes of contraction bursts upon stimulation. This circuit was glutamate dependent and was modulated by inhibitory and muscarinic inputs. Despite differentiation, embryonic stem cell-derived motoneurons carry a risk of tumorigenicity. Pre-transplantation treatment with an anti-mitotic agent leads to survival and functional muscle reinnervation if performed before eight weeks of denervation. However, differentiation of stem cells towards a ventral spinal neuronal phenotype results in a mixed population enriched in motoneurons that, once transplanted in an environment disconnected from the spinal cord, forms variable degree of self-assemble microcircuits resulting in uncontrolled muscle contractions. An alternative for motor function restoration in permanently denervated muscles can be from incorporation of channelrhodopsin-2 in muscle fibers.