Differential effects of 4‐aminopyridine on nerve injury severity‐dependent functional recovery and muscle atrophy in mice

Abstract

Peripheral nerve injury (PNI) represents a major public health problem that often leads to profound neurologic impairment, disuse muscle atrophy, and permanent disability. While the functional recovery following nerve transection and repair is disappointing, the impact of irreparable nerve gap remains poorly characterized. Recently we demonstrated a repurposing beneficial effect of 4-aminopyridine (4-AP), a potassium channel blocker, on the functional recovery and muscle atrophy after sciatic nerve crush injury in rodents. However, this effect of 4-AP is unknown in nerve transection and gap models. This study was designed to evaluate and compare the functional recovery and muscle atrophy between nerve transection and nerve gap models in the absence (saline) and presence of 4-AP treatment. Under deep anesthesia and aseptic conditions, complete nerve transection or nerve gap injury was performed in male C57BL/6 mice (n = 10/group). In novel Stepwise Nerve Transection and Glue (STG) group, right sciatic nerve was sequentially transected with fibrin gluing to prevent gap formation. In nerve gap model, a large gap was created by dissecting out 7 mm (Gap-7) of right sciatic nerve section where the proximal stump of the nerve was buried underneath the muscle with fibrin glue. Following surgery, 4-AP (40 µg, ip) or saline was given daily and sciatic function index (SFI) was determined weekly for 12 weeks to evaluate functional recovery. After 12 weeks, tibialis anterior (TA) muscles were harvested for wet weight and histomorphological analysis. To asses muscle atrophy, right TA (RTA) muscle mass is presented as a percentage of contralateral healthy left TA (LTA) muscle which was assigned 100%. We found that SFI in STG group significantly dropped from baseline -9.9±2.83 to -66±2.24 (P<0.001) on one week followed by a small but significant recovery after 2 weeks (-51±4.42; P<0.05 vs. week-1) and then a steady state level between 9-12 weeks (-63±4.33). In Gap-7 group, SFI also significantly dropped from -8.5±3.18 to -72.4±2.50 in one week (P<0.001) and continued to deteriorate until it reached a steady state level after 10 weeks (-89.7±2.51). 4-AP had no effect on the SFI recovery in either group. Muscle atrophy was more pronounced in Gap-7 group (28±2% of LTA, P<0.001) compared to STG group (89±1.5% of LTA, P<0.001). Similar to muscle mass, RTA muscle cross sectional area (CSA) and minimum ferret diameter (MFD) were also significantly smaller in Gap-7 group compared to STG group. Interestingly, 4-AP treatment caused a significantly better recovery in RTA muscle mass, CSA, and MFD in Gap-7 group compared to STG group. These preliminary findings demonstrate that post-injury functional recovery and muscle atrophy after PNI are directly related to the intervening nerve gap, and 4-AP exerts differential effects on functional recovery and muscle atrophy.