An acetylcholine receptor lacking both γ and ε subunits mediates transmission in zebrafish slow muscle synapses

Abstract

Fast and slow skeletal muscle types in larval zebrafish can be distinguished by a fivefold difference in the time course of their synaptic decay. Single-channel recordings indicate that this difference is conferred through kinetically distinct nicotinic acetylcholine receptor (AChR) isoforms. The underlying basis for this distinction was explored by cloning zebrafish muscle AChR subunit cDNAs and expressing them in Xenopus laevis oocytes. Measurements of single-channel conductance and mean open burst duration assigned α2βδε to fast muscle synaptic current. Contrary to expectations, receptors composed of only αβδ subunits (presumed to be α2βδ2 receptors) recapitulated the kinetics and conductance of slow muscle single-channel currents. Additional evidence in support of γ/ε-less receptors as mediators of slow muscle synapses was reflected in the inward current rectification of heterologously expressed α2βδ2 receptors, a property normally associated with neuronal-type nicotinic receptors. Similar rectification was reflected in both single-channel and synaptic currents in slow muscle, distinguishing them from fast muscle. The final evidence for α2βδ2 receptors in slow muscle was provided by our ability to convert fast muscle synaptic currents to those of slow muscle by knocking down ε subunit expression in vivo. Thus, for the first time, muscle synaptic function can be ascribed to a receptor isoform that is composed of only three different subunits. The unique functional features offered by the α2βδ2 receptor likely play a central role in mediating the persistent contractions characteristic to this muscle type.

Acetylcholine Receptors Enable the Transport of Rapsyn from the Golgi Complex to the Plasma Membrane

“…In sofa potato , AChR subunits other than the δ subunit; α1, β1b, γ and ε, (Mongeon et al, 2011) are expected to remain at stages where their assembly is obstructed (Green and Millar, 1995). We examined the localization of AChR subunits using mAb35, which binds to the α1 subunit of the muscle nicotinic acetylcholine receptor.…”

Zebrafish model for congenital myasthenic syndrome reveals mechanisms causal to developmental recovery

“…Moreover, at all stages tested, the synaptic currents in wild-type muscle decayed along a single exponential time course (Table 1) owing to the fact that, unlike mammals and amphibians, the open burst durations for αβδε and αβδγ receptors are similar in zebrafish (3,13). By contrast, the synaptic currents in 48-hpf twi +/− fish, corresponding to the peak of the behavioral deficit, required the sum of three exponentials for fit (Table 1 and In any case, the open burst duration for α twi βδε differs from that of α twi βδγ receptors, pointing to a unique interaction with γ not seen with ε.…”

“…The methods used to record spontaneous and evoked synaptic currents under whole-cell voltage clamp were identical to those published previously (3,13,15). Synaptic currents were recorded using a HEKA Instruments EPC-10/2 amplifier (Instrutech) and 1 μM tetrodotoxin was added for recording spontaneous synaptic currents.…”

“…Thus, the continued expression of α twi into adulthood is tolerated because of the ε expression and associated recovery, raising the likelihood of unappreciated myasthenic cases that benefit from the γ−ε switch. neuromuscular | quinidine | nicotinic receptor T he developmental switch from the embryonic αβδγ to the adult αβδε muscle ACh receptor isoform occurs in every vertebrate species studied to date (1)(2)(3). Certain mutations in ε result in the lifelong myasthenic disorder slow-channel syndrome (SCS) (4)(5)(6), whereas mutations in the γ subunit are restricted to Escobar syndrome, characterized by improvement in neuromuscular performance during development (7,8).…”

Effects of Nitric Oxide on Neuromuscular Properties of Developing Zebrafish Embryos

“…Receptors expressing the γ subunit exhibit prolonged open channel times and generate prolonged macroscopic currents whereas those expressing the ε subunit exhibit short open channel times and generate shorter macroscopic currents [61]. Whilst a delay in the switch from γ to ε subunits may contribute to the effects of NO on EF fibres, it is unlikely to explain the effects observed within ES fibres as they do not exhibit a developmentally-related change in subunit composition [59]. Future molecular and biochemical studies are needed to identify whether one or more of these developmental phenomena are influenced by NO signalling.…”