Polyethylene glycol fusion repair of severed rat sciatic nerves reestablishes axonal continuity and reorganizes sensory terminal fields in the spinal cord

Peripheral nerve injuries result in the rapid degeneration of distal nerve segments and immediate loss of motor and sensory functions;behavioral recovery is typically poor.We used a plasmalemmal fusogen,polyethylene glycol(PEG),to immediately fuse closely apposed open ends of severed proximal and distal axons in rat sciatic nerves.We have previously reported that sciatic nerve axons repaired by PEG-fusion do not undergo Wallerian degeneration,and PEG-fused animals exhibit rapid(within 2–6 weeks)and extensive locomotor recovery.Furthermore,our previous report showed that PEG-fusion of severed sciatic motor axons was non-specific,i.e.,spinal motoneurons in PEG-fused animals were found to project to appropriate as well as inappropriate target muscles.In this study,we examined the consequences of PEG-fusion for sensory axons of the sciatic nerve.Young adult male and female rats(Sprague–Dawley)received either a unilateral single cut or ablation injury to the sciatic nerve and subsequent repair with or without(Negative Control)the application of PEG.Compound action potentials recorded immediately after PEG-fusion repair confirmed conduction across the injury site.The success of PEG-fusion was confirmed through Sciatic Functional Index testing with PEG-fused animals showing improvement in locomotor function beginning at 35 days postoperatively.At 2–42 days postoperatively,we anterogradely labeled sensory afferents from the dorsal aspect of the hindpaw following bilateral intradermal injection of wheat germ agglutinin conjugated horseradish peroxidase.PEG-fusion repair reestablished axonal continuity.Compared to unoperated animals,labeled sensory afferents ipsilateral to the injury in PEG-fused animals were found in the appropriate area of the dorsal horn,as well as inappropriate mediolateral and rostrocaudal areas.Unexpectedly,despite having intact peripheral nerves,similar reorganizations of labeled sensory afferents were also observed contralateral to the injury and repair.This central reorganization may contribute to the improved behavioral recovery seen after PEG-fusion repair,supporting the use of this novel repair methodology over currently available treatments.

Polyethylene glycol fusion repair of severed sciatic nerves accelerates recovery of nociceptive sensory perceptions in male and female rats of different strains

Successful polyethylene glycol fusion(PEG-fusion)of severed axons following peripheral nerve injuries for PEG-fused axons has been reported to:(1)rapidly restore electrophysiological continuity;(2)prevent distal Wallerian Degeneration and maintain their myelin sheaths;(3)promote primarily motor,voluntary behavioral recoveries as assessed by the Sciatic Functional Index;and,(4)rapidly produce correct and incorrect connections in many possible combinations that produce rapid and extensive recovery of functional peripheral nervous system/central nervous system connections and reflex(e.g.,toe twitch)or voluntary behaviors.The preceding companion paper describes sensory terminal field reo rganization following PEG-fusion repair of sciatic nerve transections or ablations;howeve r,sensory behavioral recovery has not been explicitly explored following PEG-fusion repair.In the current study,we confirmed the success of PEG-fusion surgeries according to criteria(1-3)above and more extensively investigated whether PEG-fusion enhanced mechanical nociceptive recovery following sciatic transection in male and female outbred Sprague-Dawley and inbred Lewis rats.Mechanical nociceptive responses were assessed by measuring withdrawal thresholds using von Frey filaments on the dorsal and midplantar regions of the hindpaws.Dorsal von Frey filament tests were a more reliable method than plantar von Frey filament tests to assess mechanical nociceptive sensitivity following sciatic nerve transections.Baseline withdrawal thresholds of the sciatic-mediated lateral dorsal region differed significantly across strain but not sex.Withdrawal thresholds did not change significantly from baseline in chronic Unoperated and Sham-operated rats.Following sciatic transection,all rats exhibited severe hyposensitivity to stimuli at the lateral dorsal region of the hindpaw ipsilateral to the injury.However,PEG-fused rats exhibited significantly earlier return to baseline withdrawal thresholds than Negative Control rats.Furthermore,PEG-fused rats with significantly improved Sciatic Functional Index scores at or after 4 weeks postoperatively exhibited yet-earlier von Frey filament recove ry compared with those without Sciatic Functional Index recovery,suggesting a correlation between successful PEG-fusion and both motor-dominant and sensory-dominant behavioral recoveries.This correlation was independent of the sex or strain of the rat.Furthermore,our data showed that the acceleration of von Frey filament sensory recovery to baseline was solely due to the PEG-fused sciatic nerve and not saphenous nerve collateral outgrowths.No chronic hypersensitivity developed in any rat up to 12 weeks.All these data suggest that PEG-fusion repair of transection peripheral nerve injuries co uld have important clinical benefits.

Conundrums and confusions regarding how polyethylene glycol-fusion produces excellent behavioral recovery after peripheral nerve injuries

Current Neuroscience dogma holds that transections or ablations of a segment of peripheral nerves produce： （1） Immediate loss of axonal continuity, sensory signaling, and motor control; （2） Wallerian rapid （1-3 days） degeneration of severed distal axons, muscle atrophy, and poor behavioral recovery after many months （if ever, after ablations） by slowly-regenerating （1 mm/d）, proximal-stump outgrowths that must specifically reinnervate denervated targets; （3） Poor acceptance of microsutured nerve allografts, even if tissue-matched and immune-suppressed. Repair of transections/ablations by neurorrhaphy and well-specified-sequences of PEG-fusion solutions （one containing polyethylene glycol, PEG） successfully address these problems. However, conundrums and confusions regarding unorthodox and dramatic results of PEG-fusion repair in animal model systems often lead to misunderstandings. For example, （1） Axonal continuity and signaling is re-established within minutes by non-specifically PEG-fusing （connecting） severed motor and sensory axons across each lesion site, but remarkable behavioral recovery to near-unoperated levels takes several weeks; （2） Many distal stumps of inappropriately-reconnected, PEG-fused axons do not ever （Wallerian） degenerate and continuously innervate muscle fibers that undergo much less atrophy than otherwise-denervated muscle fibers; （3） Host rats do not reject PEG-fused donor nerve allografts in a non-immuno-privileged environment with no tissue matching or immunosuppression; （4） PEG fuses apposed open axonal ends or seals each shut （thereby preventing PEG-fusion）, depending on the experimental protocol; （5） PEG-fusion protocols produce similar results in animal model systems and early human case studies. Hence, iconoclastic PEG-fusion data appropriately understood might provoke a re-thinking of some Neuroscience dogma and a paradigm shift in clinical treatment of peripheral nerve injuries.

Protective effects of gonadal hormones on spinal motoneurons following spinal cord injury

Spinal cord injury（SCI） results in lesions that destroy tissue and disrupt spinal tracts, producing deficits in locomotor and autonomic function. The majority of treatment strategies after SCI have concentrated on the damaged spinal cord, for example working to reduce lesion size or spread, or encouraging regrowth of severed descending axonal projections through the lesion, hoping to re-establish synaptic connectivity with caudal targets. In our work, we have focused on a novel target for treatment after SCI, surviving spinal motoneurons and their target musculature, with the hope of developing effective treatments to preserve or restore lost function following SCI. We previously demonstrated that motoneurons, and the muscles they innervate, show pronounced atrophy after SCI. Importantly, SCI-induced atrophy of motoneuron dendrites can be attenuated by treatment with gonadal hormones, testosterone and its active metabolites, estradiol and dihydrotestosterone. Similarly, SCI-induced reductions in muscle fiber cross-sectional areas can be prevented by treatment with androgens. Together, these findings suggest that regressive changes in motoneuron and muscle morphology seen after SCI can be ameliorated by treatment with gonadal hormones, further supporting a role for steroid hormones as neurotherapeutic agents in the injured nervous system.

Exercise promotes recovery after motoneuron injury via hormonal mechanisms

Injuries to spinal motoneurons manifest in a variety of forms,including damage to peripheral axons,neurodegenerative disease,or direct insult centrally.Such injuries produce a variety of negative structural and functional changes in both the directly affected and neighboring motoneurons.Exercise is a relatively simple behavioral intervention that has been demonstrated to protect against,and accelerate recovery from,these negative changes.In this article,we describe how exercise is neuroprotective for motoneurons,accelerating axon regeneration following axotomy and attenuating dendritic atrophy following the death of neighboring motoneurons.In both of these injury models,the positive effects of exercise have been found to be dependent on gonadal hormone action.Here we describe a model in which exercise,hormones,and brain-derived neurotrophic factor might all interact to produce neuroprotective effects on motoneuron structure following neural injury.