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Muscarinic acetylcholine receptors (mAChRs) are widely expressed in the CNS and peripheral nervous system and play an important role in modulating the cell activity and function. We have shown that the cholinergic agonist carbachol reduces the pigeon’s inwardly rectifying potassium channel (pKir2.1) ionic currents in native vestibular hair cells. We have cloned and sequenced pigeon mAChR subtypes M2–M5 and we have studied the expression of all five mAChR subtypes (M1–M5) in the pigeon vestibular end organs (semicircular canal ampullary cristae and utricular maculae), vestibular nerve fibers and the vestibular (Scarpa’s) ganglion using tissue immunohistochemistry (IH), dissociated single cell immunocytochemistry (IC) and Western blotting (WB). We found that vestibular hair cells, nerve fibers and ganglion cells each expressed all five (M1–M5) mAChR subtypes. Two of the three odd-numbered mAChRs (M1, M5) were present on the hair cell cilia, supporting cells and nerve terminals. And all three odd numbered mAChRs (M1, M3 and M5) were expressed on cuticular plates, myelin sheaths and Schwann cells. Even-numbered mAChRs were seen on the nerve terminals. M2 was also shown on the cuticular plates and supporting cells. Vestibular efferent fibers and terminals were not identified in our studies. Results from WB of the dissociated vestibular epithelia, nerve fibers and vestibular ganglia were consistent with the results from IH and IC. Our findings suggest that there is considerable co-expression of the subtypes on the neural elements of the labyrinth. Further electrophysiological and pharmacological studies should delineate the mechanisms of action of muscarinic acetylcholine receptors on structures in the labyrinth.

Acetylcholine (ACh) is the major neurotransmitter released from vestibular efferent terminals onto hair cells and afferents. Previous studies indicate that the two classes of acetylcholine receptors, nicotinic (nAChRs) and muscarinic receptors (mAChRs), are expressed by vestibular hair cells (VHCs). To identify if both classes of receptors are present in VHCs, whole cell, voltage-clamp- and current-clamp-patch recordings were performed on isolated pigeon vestibular type I and type II HCs during the application of the cholinergic agonists, acetylcholine and carbachol, and the cholinergic antagonists, d -tubocurarine and atropine. By applying in different combinations, these compounds were used to selectively activate either nAChRs or mAChRs. The effects of nAChR and mAChR activation on HC currents and zero electrode current potential ( V z ) were monitored. It was found that presumed mAChR activation decreased both inward and outward currents in both type I and type II HCs, resulting in either a depolarization or hyperpolarization. Conversely, nAChR activation mainly increased both inward and outward currents in type II HCs, resulting in a hyperpolarization of their V z . nAChR activation also increased outward currents in type I HCs resulting in either a depolarization or hyperpolarization of their V z . The decrease of inward and outward currents and the depolarization of the V z in type I pigeon HCs by activation of mAChRs represents a new finding. Ion channel candidates in pigeon vestibular HCs that might underlie the modulation of the macroscopic ionic currents and V z by different AChR activation are discussed.

The rodent vestibular system is immature at birth. During the first postnatal week, vestibular type I and type II hair cells start to acquire their characteristic morphology and afferent innervation. We have studied postnatal changes in the membrane properties of type I hair cells acutely isolated from the semicircular canals (SCC) of gerbils and rats using whole cell patch clamp and report for the first time developmental changes in ionic conductances in these cells. At postnatal day (P) 5 immature hair cells expressed a delayed rectifier K+ conductance ( GDR) which activated at potentials above approximately −50 mV in both species. Hair cells also expressed a transient Na+ conductance ( GNa) with a mean half-inactivation of approximately −90 mV. At P6 in rat and P7 in gerbil, a low-voltage activated K+ conductance ( GK,L) was first observed and conferred a low-input resistance, typical of adult type I hair cells, on SCC type I hair cells. GK,L expression in hair cells increased markedly during the second postnatal week and was present in all rat type I hair cells by P14. In gerbil hair cells, GK,L appeared later and was present in all type I hair cells by P19. During the third postnatal week, GNa expression declined and was absent by the fourth postnatal week in rat and the sixth postnatal week in gerbils. Understanding the ionic changes associated with hair cell maturation could help elucidate development and regeneration mechanisms in the inner ear.