Rapid, effective, and long-lasting behavioral recovery produced by microsutures, methylene blue, and polyethylene glycol after completely cutting rat sciatic nerves.
Behavioral function lost in mammals (including humans) after peripheral nerve severance is slowly (weeks to years) and often poorly restored by 1-2-mm/day, nonspecifically directed outgrowths from proximal axonal stumps. To survive, proximal stumps must quickly repair (seal) plasmalemmal damage. We report that, after complete cut- or crush-severance of rat sciatic nerves, morphological continuity, action potential conduction, and behavioral functions can be consistently (>98% of trials), rapidly (minutes to days), dramatically (70-85% recovery), and chronically restored and some Wallerian degeneration prevented. We assess axoplasmic and axolemmal continuity by intra-axonal dye diffusion and action potential conduction across the lesion site and amount of behavioral recovery by Sciatic Functional Index and Foot Fault tests. We apply well-specified sequences of solutions containing FDA-approved chemicals. First, severed axonal ends are opened and resealing is prevented by hypotonic Ca²⁺-free saline containing antioxidants (especially methylene blue) that inhibit plasmalemmal sealing in sciatic nerves in vivo, ex vivo, and in rat B104 hippocampal cells in vitro. Second, a hypotonic solution of polyethylene glycol (PEG) is applied to open closely apposed (by microsutures, if cut) axonal ends to induce their membranes to flow rapidly into each other (PEG-fusion), consistent with data showing that PEG rapidly seals (PEG-seals) transected neurites of B104 cells, independently of any known endogenous sealing mechanism. Third, Ca²⁺-containing isotonic saline is applied to induce sealing of any remaining plasmalemmal holes by Ca²⁺-induced accumulation and fusion of vesicles. These and other data suggest that PEG-sealing is neuroprotective, and our PEG-fusion protocols that repair cut- and crush-severed rat nerves might rapidly translate to clinical procedures.
Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, Texas; Section of Neurobiology, University of Texas at Austin, Austin, Texas, USA. bittner@mail.utexas.edu
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Bittner
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Keating
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Kane
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Britt
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Spaeth
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Fan
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Zuzek
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Wilcott
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Thayer
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Winograd
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Gonzalez-Lima
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Schallert
D000704:Analysis of Variance; D000818:Animals; D001522:Behavior, Animal / Q000187:drug effects*; D004195:Disease Models, Animal; D004576:Electromyography; D019054:Evoked Potentials, Motor / Q000187:drug effects; D005456:Fluorescent Dyes; D008751:Methylene Blue / Q000627:therapeutic use*; D008866:Microsurgery / Q000379:methods*; D009431:Neural Conduction / Q000187:drug effects / Q000502:physiology; D011092:Polyethylene Glycols / Q000627:therapeutic use*; D051381:Rats; D017207:Rats, Sprague-Dawley; D020127:Recovery of Function / Q000187:drug effects / Q000502:physiology*; D020426:Sciatic Neuropathy / Q000188:drug therapy / Q000503:physiopathology / Q000601:surgery; D013997:Time Factors; D014741:Video Recording
D005456:Fluorescent Dyes; D011092:Polyethylene Glycols; D008751:Methylene Blue