**2. Physiology of the NMJ**

This section introduces molecular basis of development and maintenance of the NMJ, and physiological features of nicotinic muscle AChR.

#### **2.1 NMJ synaptogenesis**

At the NMJ, MuSK is an indirect receptor for agrin (Valenzuela *et al.*, 1995; Dechiara *et al.*, 1996). Agrin released from the nerve terminal binds to LRP4 on the postsynaptic membrane (Kim *et al.*, 2008; Zhang *et al.*, 2008). Binding of LRP4 to agrin phosphorylates MuSK. Phosphorylated MuSK recruits the noncatalytic adaptor protein Dok-7 (Okada *et al.*, 2006). Once recruited, Dok-7 further facilitates phosphorylation of MuSK, and induces clustering of rapsyn and AChR by phosphorylating the subunit of AChR. Rapsyn self-associates and makes a homomeric cluster at the endplate, which serves as a scaffold for AChR. Rapsyn and AChR bind each other with a stoichiometry of 1:1. Rapsyn also binds to -dystroglycan and links the rapsyn scaffold to the subsynaptic cytoskeleton (Froehner *et al.*, 1990; Cartaud *et al.*, 1998; Ramarao & Cohen, 1998; Ramarao *et al.*, 2001). Except for LRP4, each of the above molecules is a CMS target.

#### **2.2 Physiology of the nicotinic muscle AChR**

176 Neuromuscular Disorders

above, AChR, rapsyn, MuSK, Dok-7, plectin, and NaV1.4 are associated with the postsynaptic membrane. Agrin, ColQ, and 2-laminin reside in the synaptic basal lamina. The only presynaptic disease protein identified to date is choline acetyltransferase (ChAT). A target molecule and its synaptic localization of glutamine-fructose-6-phosphate aminotransferase 1 (GFPT1) are still unresolved but the phenotypic consequence is the postsynaptic AChR deficiency. This chapter focuses on molecular bases of these three

This section introduces molecular basis of development and maintenance of the NMJ, and

At the NMJ, MuSK is an indirect receptor for agrin (Valenzuela *et al.*, 1995; Dechiara *et al.*, 1996). Agrin released from the nerve terminal binds to LRP4 on the postsynaptic membrane (Kim *et al.*, 2008; Zhang *et al.*, 2008). Binding of LRP4 to agrin phosphorylates MuSK. Phosphorylated MuSK recruits the noncatalytic adaptor protein Dok-7 (Okada *et al.*, 2006). Once recruited, Dok-7 further facilitates phosphorylation of MuSK, and induces clustering of rapsyn and AChR by phosphorylating the subunit of AChR. Rapsyn self-associates and makes a homomeric cluster at the endplate, which serves as a scaffold for AChR. Rapsyn and AChR bind each other with a stoichiometry of 1:1. Rapsyn also binds to -dystroglycan and links the rapsyn scaffold to the subsynaptic cytoskeleton (Froehner *et al.*, 1990; Cartaud *et al.*, 1998; Ramarao & Cohen, 1998; Ramarao *et al.*, 2001). Except for LRP4, each of the above

groups of CMS.

Fig. 1. Schematic of molecules expressed at the NMJ

physiological features of nicotinic muscle AChR.

**2. Physiology of the NMJ** 

**2.1 NMJ synaptogenesis** 

molecules is a CMS target.

Nicotinic AChRs are pentameric ligand-gated ion channels. The family of pentameric ligand-gated ion channels includes cationic AChRs, cationic serotonergic receptors (5HT3), anionic glycine receptors, and anionic GABAA and GABAC receptors (Keramidas *et al.*, 2004). Heteromeric neuronal nicotinic AChRs are comprised of various combinations of (2-7) and subunits (2-4), whereas homomeric AChRs are formed only by a single subunit (e.g., 7-9) (Mihailescu & Drucker-Colin, 2000). On the other hand, nicotinic muscle AChRs have only two forms: fetal AChR that carries the , , , and subunits encoded by *CHRNA1*, *CHRNB1, CHRND, CHRNG*, respectively, in the stoichiometry 2; and adult-type AChR that carries the subunit instead of the subunit in the stoichiometry 2 (Mishina *et al.*, 1986). The subunit is encoded by *CHRNE*. Nicotinic muscle AChR harbors two binding sites for ACh at the interfaces between the and / subunits (Lee *et al.*, 2009; Mukhtasimova *et al.*, 2009). Binding of a single ACh molecule opens the channel pore but for a short time. Binding of two ACh molecules stabilizes the open state of AChR, and AChR stays open for a longer time. Only cations pass through the channel pore of nicotinic AChRs. Unlike sodium, potassium, or calcium channels, AChRs, in general, have no selectivity for cations, but 7 AChRs have 10-20 times higher permeability for Ca2+ than for Na+.
