MuSK Regulates Neuromuscular Junction Nav1.4 Localization and Excitability

• DOI: 10.1523/jneurosci.1279-23.2025
• Publication Date: 2025-01-29T18:50:45Z
• Authors: Lauren A. Fish; Madison D. Ewing; Kelly A. Rich; Chengjie Xi; Isabella Chen; Diego Jaime; Laura A. Madigan; Xueyong Wang; Justin L. Shahtout; Rita E. Feder; Katsuhiko Funai; Jan L. Christian; Kristi A. Wharton; Mark M. Rich; William D. Arnold; Justin R. Fallon

The neuromuscular junction (NMJ) is the linchpin of nerve-evoked muscle contraction. Broadly, the NMJ transduces nerve action potentials into muscle fiber action potentials (MFAPs). Efficient neuromuscular transmission requires cholinergic signaling, which generates endplate potentials (EPPs), and excitation, the amplification of an EPP by postsynaptic voltage-gated sodium channels (Nav1.4) to generate the MFAP. Compared to the cholinergic component, the signaling pathways that organize Nav1.4 are poorly characterized. Muscle-specific kinase (MuSK), in addition to its Ig1 domain-dependent role as the main organizer of acetylcholine receptors (AChRs), also binds BMPs via its Ig3 domain and shapes BMP-induced signaling. Using mice lacking the MuSK Ig3 domain (“ΔIg3-MuSK”), we probed the role of this domain at the NMJ. NMJs formed in ΔIg3-MuSK animals with pre- and postsynaptic specializations aligned at all ages examined. However, the ΔIg3-MuSK postsynaptic apparatus was fragmented from an early age. Synaptic electrophysiology showed that spontaneous and nerve-evoked acetylcholine release, AChR density, and endplate currents were comparable at WT and ΔIg3-MuSK NMJs. However, single fiber electromyography revealed that nerve-evoked MFAPs in ΔIg3-MuSK muscle were abnormal, exhibiting jitter and blocking. Nerve-evoked compound muscle action potentials and muscle force were also diminished. Finally, Nav1.4 levels were reduced at ΔIg3-MuSK NMJs, but not extrasynaptically, indicating that the observed excitability defects result from impaired synaptic localization of this ion channel. We propose distinct, domain-specific roles for MuSK at the NMJ: the Ig1 domain mediates agrin-LRP4-mediated AChR localization, while the Ig3 domain maintains postsynaptic Nav1.4 density, conferring the muscle excitability required to amplify cholinergic signals and trigger action potentials.