Studies of plasticity after neuronal injury
Neurons of the adult mammalian central nervous system (CNS) and peripheral nervous system (PNS) differ in their response to injury. PNS neurons have a robust growth response and are capable of regenerating their axons and reinnervating peripheral targets following nerve injury. However, despite this robust regenerative response, specificity of target reinnervation is often poor, leading to a suboptimal functional outcome. In contrast, injured CNS neurons have a poor growth response and completely fail to regenerate, but uninjured axons that survive the injury undergo some degree of spontaneous reorganisation that is accompanied by limited functional improvement. In theory, appropriate reorganisation of central connections could compensate for topographical errors in the periphery and enhancing the innate capacity for reorganisation after CNS injury could compensate for lost function. Thus, recovery from both types of injury may benefit from strategies to enhance CNS plasticity. This Thesis makes use of an experimental strategy to promote plastic changes, that is, the enzyme chondroitinase ABC (ChABC) which degrades chondroitin sulphate proteoglycans - a major class of inhibitory extracellular matrix molecules. Viral vector technology was used to optimise the delivery of ChABC by gene transfer to host spinal cord cells. A lentiviral vector expressing the ChABC gene (LV-ChABC) was delivered to the adult rat spinal cord to achieve long-term delivery of active ChABC in vivo. After peripheral nerve injury, where two forelimb nerves were cut and repaired with different levels of inaccuracy, electrophysiological studies demonstrated that LV-ChABC injection led to significant plasticity of central connections, such as reorganisation of high and low threshold polysynaptic reflexes. ChABC did not lead to any changes in reflex activity in uninjured animals. These results indicate that ChABC facilitates the amplification of compensatory changes in the spinal cord following injury to the periphery. ChABC treatment was also investigated in the context of CNS injury. Following contusion injury, a clinically relevant model which mimics the functional and pathological characteristics typical of a human spinal cord injury, LV-ChABC injection led to significant functional improvements, including enhanced conduction, reorganisation of spinal reflexes below the lesion and improved performance on a sensorimotor behavioural task. in vivo. as Finally, viral vector technology was used to develop a novel tool for the study of anatomical plasticity. By expressing the construct synaptopHluorin (SpH), lentiviral and adenoviral-associated viral vectors were used to label presynaptic terminals in vitro and in vivo. As the pHluorin component of the label is pH sensitive, this tool has the potential additional advantage of enabling the visualisation of synaptic function The SpH vector was used to label a major motor pathway, the corticospinal tract, well as sensory neurons, and to visualise their terminals as SpH-labelled puncta. Using this tool, it will be possible to study synaptogenesis in a quantifiable manner, with the potential to study the activity of these new synaptic terminals. Thus, this Thesis uses a gene delivery approach to deliver ChABC to the spinal cord after central or peripheral neuronal injury, leading to the promotion of plasticity and functional repair. The development of a novel tool for visualising synapses is also described, presenting a novel opportunity for the study of plasticity after neuronal injury.
Networked Digital Library of Theses and Dissertations