Enhancing Motor and Sensory Axon Regeneration after Peripheral Nerve Injury Using Bioluminescent Optogenetics.

Introduction-Recovery from peripheral nerve injuries is poor even though injured peripheral axons can regenerate. Novel therapeutic approaches are needed. The most successful preclinical experimental treatments have relied on increasing the activity of the regenerating axons, but the approaches taken are not applicable to many nerve-injured patients. Bioluminescent optogenetics (BL-OG) is a novel method of increasing the excitation of neurons that might be similar to that found with activity-dependent experimental therapies. We investigated the use of BL-OG as an approach to promoting axon regeneration following peripheral nerve injury. Methods-BL-OG uses luminopsins, light-sensing ion channels (opsins) fused with a light-emitting luciferase. When exposed to a luciferase substrate, such as coelenterazine (CTZ), luminopsins expressed in neurons generate bioluminescence and produce excitation through their opsin component. Adeno-associated viral vectors encoding either an excitatory luminopsin (eLMO3) or a mutated form (R115A) that can generate bioluminescence but not excite neurons were injected into mouse sciatic nerves. After retrograde transport and viral transduction, nerves were cut and repaired by simple end-to-end anastomosis, and mice were treated with a single dose of CTZ. Results-Four weeks after nerve injury, compound muscle action potentials (M waves) recorded in response to sciatic nerve stimulation were more than fourfold larger in mice expressing the excitatory luminopsin than in controls expressing the mutant luminopsin. The number of motor and sensory neurons retrogradely labeled from reinnervated muscles in mice expressing eLMO3 was significantly greater than the number in mice expressing the R115A luminopsin and not significantly different from those in intact mice. When viral injection was delayed so that luminopsin expression was induced after nerve injury, a clinically relevant scenario, evoked M waves recorded from reinnervated muscles were significantly larger after injury in eLMO3-expressing mice. Conclusions-Treatment of peripheral nerve injuries using BL-OG has significant potential to enhance axon regeneration and promote functional recovery.

Effects of route of administration on neural exposure to platinum-based chemotherapy treatment: a pharmacokinetic study in rat.

The persisting need for effective clinical treatment of chemotherapy-induced neurotoxicity (CIN) motivates critical evaluation of preclinical models of CIN for their translational relevance. The present study aimed to provide the first quantitative evaluation of neural tissue exposed in vivo to a platinum-based anticancer compound, oxaliplatin (OX) during and after two commonly used dosing regimens: slow IV infusion used clinically and bolus IP injection used preclinically. Inductively-coupled plasma mass spectrometry analysis of dorsal root ganglia indicated that while differences in the temporal dynamics of platinum distribution exist, key drivers of neurotoxicity, e.g. peak concentrations and exposure, were not different across the two routes of administration. We conclude that the IP route of OX administration achieves clinically relevant pharmacokinetic exposure of neural tissues in a rodent model of CIN.

Bioluminescent Optogenetics: A Novel Experimental Therapy to Promote Axon Regeneration after Peripheral Nerve Injury.

Functional recovery after peripheral nerve injury (PNI) is poor, mainly due to the slow and incomplete regeneration of injured axons. Experimental therapies that increase the excitability of the injured axons have proven remarkably successful in promoting regeneration, but their clinical applicability has been limited. Bioluminescent optogenetics (BL-OG) uses luminopsins, fusion proteins of light-generating luciferase and light-sensing ion channels that could be used to increase neuronal excitability if exposed to a suitable substrate. Excitatory luminopsins were expressed in motoneurons of transgenic mice and in wildtype mice transduced with adeno-associated viral vectors. Intraperitoneal administration of coelenterazine (CTZ), a known luciferase substrate, generated intense bioluminescence in peripheral axons. This bioluminescence increased motoneuron excitability. A single administration of CTZ immediately after sciatic nerve transection and repair markedly enhanced motor axon regeneration. Compound muscle action potentials were 3-4 times larger than controls by 4 weeks after injury. The results observed with transgenic mice were comparable to those of mice in which the luminopsin was expressed using viral vectors. Significantly more motoneurons had successfully reinnervated muscle targets four weeks after nerve injury in BL-OG treated mice than in controls. Bioluminescent optogenetics is a promising therapeutic approach to enhancing axon regeneration after PNI.

Cancer Exacerbates Chemotherapy-Induced Sensory Neuropathy.

For the constellation of neurologic disorders known as chemotherapy-induced peripheral neuropathy, mechanistic understanding and treatment remain deficient. Here, we present the first evidence that chronic sensory neuropathy depends on nonlinear interactions between cancer and chemotherapy. Global transcriptional profiling of dorsal root ganglia revealed differential expression, notably in regulators of neuronal excitability, metabolism, and inflammatory responses, all of which were unpredictable from effects observed with either chemotherapy or cancer alone. Systemic interactions between cancer and chemotherapy also determined the extent of deficits in sensory encoding and ion channel protein expression by single mechanosensory neurons, with the potassium ion channel Kv3.3 emerging as one potential contributor to sensory neuron dysfunction. Validated measures of sensorimotor behavior in awake, behaving animals revealed dysfunction after chronic chemotherapy treatment was exacerbated by cancer. Notably, errors in precise forelimb placement emerged as a novel behavioral deficit unpredicted by our previous study of chemotherapy alone. These original findings identify novel contributors to peripheral neuropathy and emphasize the fundamental dependence of neuropathy on the systemic interaction between chemotherapy and cancer. SIGNIFICANCE: These findings highlight the need to account for pathobiological interactions between cancer and chemotherapy as a major contributor to neuropathy and will have significant and immediate impact on future investigations in this field.

Abnormal response of distal Schwann cells to denervation in a mouse model of motor neuron disease.

In several animal models of motor neuron disease, degeneration begins in the periphery. Clarifying the possible role of Schwann cells remains a priority. We recently showed that terminal Schwann cells (TSCs) exhibit abnormalities in postnatal mice that express mutations of the SOD1 enzyme found in inherited human motor neuron disease. TSC abnormalities appeared before disease-related denervation commenced and the extent of TSC abnormality at P30 correlated with the extent of subsequent denervation. Denervated neuromuscular junctions (NMJs) were also observed that lacked any labeling for TSCs. This suggested that SOD1 TSCs may respond differently than wildtype TSCs to denervation which remain at denervated NMJs for several months. In the present study, the response of SOD1 TSCs to experimental denervation was examined. At P30 and P60, SC-specific S100 labeling was quickly lost from SOD1 NMJs and from preterminal regions. Evidence indicates that this loss eventually becomes complete at most SOD1 NMJs before reinnervation occurs. The loss of labeling was not specific for S100 and did not depend on loss of activity. Although some post-denervation labeling loss occurred at wildtype NMJs, this loss was never complete. Soon after denervation, large cells appeared near SOD1 NMJ bands which colabeled for SC markers as well as for activated caspase-3 suggesting that distal SOD1 SCs may experience cell death following denervation. Denervated SOD1 NMJs viewed 7 days after denervation with the electron microscope confirmed the absence of TSCs overlying endplates. These observations demonstrate that SOD1 TSCs and distal SCs respond abnormally to denervation. This behavior can be expected to hinder reinnervation and raises further questions concerning the ability of SOD1 TSCs to support normal functioning of motor terminals.