Burn injury (BI) leads to neuromuscular dysfunction (NMD), and muscle weakness (MW). The relationship of motor nerve terminal and synaptic changes to MW after BI is unknown. Integrity of neuromuscular junction (NMJ) depends on tropic signals from nerve to muscle mediated via synaptic acetylcholine receptors (AChRs), and on retrograde cues from muscle including pivotal signaling via Akt (a.k.a., PKB) and glycogen synthase kinase-3 (GSK-3) to synapse. Terminal Schwann cells (TSC) function is also essential to NMJ integrity. We posit that inflammatory responses of BI, and mitochondrial dysfunction (MD) of NMJ tissues blunt tropic signaling, and TSC function leading to nerve and synaptic derangements, and these derangements are central to the NMD and MW. The major challenge is how to effectively reverse the inflammatory responses-mediated NMJ changes. TSCs constitutively express ?7AChRs isoform, and muscles up-regulate ?7AChRs after BI. Published, and our new data show that ?7AChRs stimulation with specific agonist, GTS-21, mitigates inflammatory responses, MD and enhances signaling via Akt/GSK-3. The hypothesis tested is that stimulation with GTS-21 of ?7AChRs, via its pluri-potent actions, will protect NMJ changes in BI mice.
Specific Aim 1 tests the hypothesis that synaptic and nerve aberrations of BI lead to NMD and MW. Using Thy1-YFP (yellow fluorescent protein) transgenic mice (depicting motor nerve) and fluorescent-bungarotoxin to label synaptic AChRs, time-dependent nerve and synaptic morphological changes after BI, and their relationship to muscle function (tension and fatigability) will be characterized. Use of Thy1-mito CFP (cyan fluorescent protein) transgenic mice depicting nerve MD function, and S100-GFP (green fluorescent protein) transgenic mice reflecting TSC status will elucidate the role of BI-induced nerve MD and TSC changes to MW.
Specific Aim 2 tests the hypothesis that ?7AChRs play a protective role in maintenance of NMJ integrity by modulating aberrant inflammatory responses, MD, and impaired Akt/GSK-3 signaling of BI. At fixed time points based on Specific Aim 1 results, morphological and functional changes at NMJ will be studied in wild type and ?7AChR knockout mice during early injury-phase and later recovery-phase of BI.
Specific Aim 3 tests the hypothesis that treatment with GTS-21, a specific agonist of ?7AChRs, will mitigate synaptic and nerve terminal changes of BI. The functional and morphological changes, the specificity and signaling pathways mediating beneficial effects of GTS-21 will also be elucidated. These studies will characterize the NMJ changes of BI, their relationship to MW, and pivotal protective role of ?7AChRs in NMJ integrity. Therapeutics with GTS-21 via a7AChRs stimulation is an untested novel approach to restore synaptic, and nerve terminal changes and MW of BI.
This project applies integrated morphologic, pharmacologic and genetic approaches to elucidate the relationship of synaptic & nerve terminal changes to muscle weakness of burn, and reverse these neuromuscular changes by ?7AChRs stimulation with GTS-21.
