Resealing of transected myelinated mammalian axon membranes in vivo: Possible mechanism underlying calcium dependence

Date of Award




Degree Name

Doctor of Philosophy (Ph.D.)

First Committee Member

John N. Barrett, Committee Chair


Resealing of transected rat dorsal root axons was investigated in vivo using exclusion of a membrane-impermeant dye to assay membrane resealing. Resealing was Ca$\sp{2+}$-dependent, requiring $\mu$M levels of extracellular. (Ca$\sp{2+}\rbrack$ to proceed, and was further accelerated in 1mM Ca$\sp{2+}.$ The percentage of resealed axons increased with time during the first 120 minutes. Two hours after transection, 84% of axons had resealed in saline containing 2 mM Ca$\sp{2+},$ 28%, resealed in 10 $\mu$M Ca$\sp{2+}$ and only 3% had resealed after 120 minutes in 3 mM BAPTA. Resealing was faster in smaller caliber axons $(<5\ \mu$m axoplasmic diameter) than in larger axons. The enhancing effect of Ca$\sp{2+}$ could be overcome by both non-specific cysteine protease inhibitors (e.g. leupeptin) and inhibitors specific for the calpain family of Ca$\sp{2+}$-activated proteases. Inhibition of phospholipase A$\sb{2}$ by 50-100 $\mu$M arachidonyl trifluoromethyl ketone did not inhibit resealing in the presence of 2 mM Ca$\sp{2+}.$ Resealing in low Ca$\sp{2+}$ was not enhanced by 50 $\mu$M or 1 mM colchicine and was slightly increased by 55 $\mu$g/ml (${\sim}180\ \mu$M) nocodazole, both of which are compounds that disrupt microtubules. Resealing in the absence of added extracellular Ca$\sp{2+}$ was enhanced by the presence of 0.5% dimethylsulfoxide (DMSO). Based on these results, we propose that activation of endogenous proteases by elevated intracellular Ca$\sp{2+}$ following injury enhances resealing of axon membranes in vivo. These findings are compatible with the hypothesis that resealing is facilitated by disruption of certain elements of the cytoskeleton and/or the interface between the cytoskeleton and the membrane.


Biology, Neuroscience; Biology, Cell; Biology, Animal Physiology

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