Selective localization of annexin V within depolarized skeletal muscle

Objectives The uptake of annexin V in the periphery of ischemic lesions in the heart and brain cannot be fully explained by apoptosis; these sites have little to no cell death. We postulated that the altered transmembrane potential of ischemic cardio-myocytes and neurons may contribute to the increased uptake of annexin V in these regions. We wished to use active and inactive forms of fluorescent annexin V to study the effect of local i.m. injection of succinyl-choline, a depolarizing agent, in the thighs of mice.

Methods Adult Swiss Webster mice received an injection of 0.08 mg/kg of succinyl-choline into the medial aspect of the left calf muscle. After 1 min mice received 50 µg of annexin V-128-Alexa-680 or annexin V-137-Alexa-680 (an inactive mutant as a protein control) via penile vein injection. 2 hours later mice were sacrificed with removal of the overlying skin of the thigh and calves. The legs of mice were imaged with the eXplore Optix system (ART Advanced Research Technologies Inc, Montreal (Quebec), Canada) Acquired images were analyzed with OptiView software.

Results There was a 7.44 ± 3.43 fold increase of annexin V-128 uptake in the left (treated n=6) calf muscle as compared to the right (untreated) calf, but only a 1.59 ± 0.76 fold increase seen with annexin V-137 (protein control, n = 3 mice).

Conclusions Annexin V localizes to the site of depolarized muscle cells and the effect appears to be specific and not due to pooling of blood or increased capillary permeability.