Early Alterations in Structural and Functional Properties in the Neuromuscular Junctions of Mutant FUS Mice

Amyotrophic lateral sclerosis (ALS) is manifested as skeletal muscle denervation, loss of motor neurons and finally severe respiratory failure. Mutations of RNA-binding protein FUS are one of the common genetic reasons of ALS accompanied by a 'dying back' type of degeneration. Using fluorescent approaches and microelectrode recordings, the early structural and functional alterations in diaphragm neuromuscular junctions (NMJs) were studied in mutant FUS mice at the pre-onset stage. Lipid peroxidation and decreased staining with a lipid raft marker were found in the mutant mice. Despite the preservation of the end-plate structure, immunolabeling revealed an increase in levels of presynaptic proteins, SNAP-25 and synapsin 1. The latter can restrain Ca2+-dependent synaptic vesicle mobilization. Indeed, neurotransmitter release upon intense nerve stimulation and its recovery after tetanus and compensatory synaptic vesicle endocytosis were markedly depressed in FUS mice. There was a trend to attenuation of axonal [Ca2+](in) increase upon nerve stimulation at 20 Hz. However, no changes in neurotransmitter release and the intraterminal Ca2+ transient in response to low frequency stimulation or in quantal content and the synchrony of neurotransmitter release at low levels of external Ca2+ were detected. At a later stage, shrinking and fragmentation of end plates together with a decrease in presynaptic protein expression and disturbance of the neurotransmitter release timing occurred. Overall, suppression of synaptic vesicle exo-endocytosis upon intense activity probably due to alterations in membrane properties, synapsin 1 levels and Ca2+ kinetics could be an early sign of nascent NMJ pathology, which leads to neuromuscular contact disorganization.

Automated summary

Amyotrophic lateral sclerosis (ALS) is a disease characterized by skeletal muscle denervation, motor neuron loss, and respiratory failure. Mutations in the FUS gene are a common genetic cause of ALS, resulting in degeneration. In mutant FUS mice at the pre-onset stage, early structural and functional alterations in diaphragm neuromuscular junctions (NMJs) were observed. Lipid peroxidation and decreased lipid raft staining were found in the mutant mice. Despite preserved end-plate structure, increased levels of presynaptic proteins and impaired neurotransmitter release were observed in FUS mice. These findings suggest that alterations in membrane properties, synapsin 1 levels, and calcium kinetics could be early indicators of nascent NMJ pathology in ALS.