Changes in mRNA Expression of Nicotinic Acetylcholine Receptor Subunits during Embryonic Development of Mouse Masseter Muscle

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Abstract
Nicotinic acetylcholine receptors (nAChRs) switch from the embryonic-type (alpha 2 beta gamma delta subunits) to the adult-type (alpha 2 beta epsilon delta subunits), and disappear besides the neuromuscular junctions with the development of trunk and limb skeletal muscles. However, little is known about this process during the embryonic development of masseter muscle. To identify the time course of the nAChR transition from embryonic day (E) 11 to the newborn stage in mouse masseter muscle, we analyzed the expression level of delta, epsilon, and gamma subunit mRNAs by competitive polymerase chain reaction in combination with reverse transcription as well as distribution of delta subunit protein by immunohistochemistry. The nAChR delta subunit mRNA was initially detected at E11, showed an approximately 25-fold increase (p < 0.0001) between E11 and E17, and plateaued thereafter until the newborn stage. Immunostaining for delta subunit was observed in the whole portions of masseter myofibers at E17 and birth, suggesting that the nAChR elimination does not begin even at the newborn stage. The epsilon subunit mRNA initially appeared at E17, and increased in quantity by 144% (p < 0.0001) up to the newborn stage. The quantity of gamma subunit mRNA increased by approximately 240% (p < 0.0001) between E11 and E17, and then decreased by 22% (p < 0.05) from E17 value at the newborn stage. The beginning of the expression of the epsilon subunit mRNA was coincident with the beginning of the decrease in the quantity of the gamma subunit mRNA, suggesting that the nAChR subunit switch begins at E17.