Nicotinic acetylcholine receptors (nAChRs) at zebrafish red and white muscle show different properties during development

Abstract

Nicotinic acetylcholine receptors (nAChRs) are highly expressed at the vertebrate neuromuscular junction (NMJ) where they are required for muscle activation. Understanding the factors that underlie NMJ development is critical for a full understanding of muscle function. In this study we performed whole cell and outside-out patch clamp recordings, and single-cell RT-qPCR from zebrafish red and white muscle to examine the properties of nAChRs during the first 5 days of development. In red fibers miniature endplate currents (mEPCs) exhibit single exponential time courses at 1.5 days postfertilization (dpf) and double exponential time courses from 2 dpf onwards. In white fibers, mEPCs decay relatively slowly, with a single exponential component at 1.5 dpf. By 2 and 3 dpf, mEPC kinetics speed up, and decay with a double exponential component, and by 4 dpf the exponential decay reverts back to a single component. Single channel recordings confirm the presence of two main conductance classes of nAChRs (∼45 pS and ∼65 pS) in red fibers with multiple time courses. Two main conductance classes are also present in white fibers (∼55 pS and ∼73 pS), but they exhibit shorter mean open times by 5 dpf compared with red muscle. RT-qPCR of mRNA for nicotinic receptor subunits supports a switch from γ to ε subunits in white fibers but not in red. Our findings provide a developmental profile of mEPC properties from red and white fibers in embryonic and larval zebrafish, and reveal previously unknown differences between the NMJs of these muscle fibers.© 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 916-936, 2016.

Adenosine Promotes Endplate nAChR Channel Activity in Adult Mouse Skeletal Muscle Fibers via Low Affinity P1 Receptors

“…The second evidence comes from the results obtained by patch-clamp recordings carried out at the endplate region of isolated mouse FDB fibres, where adenosine and the P1R agonist NECA significantly affected the NP o , the open frequency and the time constant of the adult nAChR channels. It is generally assumed that the kinetics of the MEPCs reflects the channel activity of the postsynaptic nAChRs (Katz and Miledi, 1973;Lu et al, 1993;Van der Kloot et al, 1994;Ahmed and Ali, 2016). In our control conditions, the rise and decay time of MEPCs were similar to those already reported elsewhere (Robertson and Wann, 1984;Tanzi and D Angelo, 1995;Petrov et al, 2009) and both values were significantly increased in the presence of adenosine.…”

Synaptic silencing of fast muscle is compensated by rewired innervation of slow muscle

“…Although the distinct compositions between fast and slow muscle AChR subunits were recently described (13), the mechanism remains unresolved. Ahmed and Ali reported that slow muscles expressed mRNA of  and  subunits (35). The absence of alternate subunit composition reported here also strongly supports the idea on July 31, 2020 http://advances.sciencemag.org/ Downloaded from that the subunit composition is regulated at the posttranslational level.…”

Motor neuron development in zebrafish is altered by brief (5-hr) exposures to THC (∆9-tetrahydrocannabinol) or CBD (cannabidiol) during gastrulation

“…We had previously found that the decay time course of mEPCs recorded from white fibers of 2 dpf embryos was bi-exponential in nature due to the presence of multiple classes of nAChRs 32 , 33 . Because changes in the kinetics often signify a change in the subunit composition of synaptic receptors, we examined the exponential decay of mEPCs but found that there was no significant difference amongst any of the treatments (Fig.…”