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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