Alpha‐9 nicotinic acetylcholine receptor immunoreactivity in the rodent vestibular labyrinth

Abstract

Vestibular tissues (cristae ampullares, macular otolithic organs, and Scarpa's ganglia) in chinchilla, rat, and guinea pig were examined for immunoreactivity to the alpha9 nicotinic acetylcholine receptor (nAChR) subunit. The alpha9 antibody was generated against a conserved peptide present in the intracellular loop of the predicted protein sequence of the guinea pig alpha9 nAChR subunit. In the vestibular periphery, staining was observed in calyces around type I hair cells, at the synaptic pole of type II hair cells, and in varying levels in Scarpa's ganglion cells. Ganglion cells were also triply labeled to detect alpha9, calretinin, and peripherin. Calretinin labels calyx-only afferents. Peripherin labels bouton-only afferents. Dimorphic afferents, which have both calyx and bouton endings, are not labeled by calretinin or peripherin. In these experiments, alpha9 was expressed in both calyx and dimorphic afferents. A subpopulation of small ganglion cells did not contain the alpha9 nAChR but did stain for peripherin. We surmise that these are bouton-only afferents. Bouton (regularly discharging) afferents also show efferent responses, although they are qualitatively different from those in irregularly discharging (calyx and dimorphic) afferents, much slower and longer lasting. Thus, regular afferents are probably more affected via a muscarinic cholinergic or a peptidergic mechanism, with a much smaller superimposed fast nicotinic-type response. This latter response could be due to one of the other nicotinic receptors that have been described in studies from other laboratories.

Mechanisms of Efferent-Mediated Responses in the Turtle Posterior Crista

“…A possible postsynaptic localization of ␣9/␣10 nAChRs is currently controversial. Antibodies to ␣9 labeled calyces and most Scarpa's ganglion cells in rodents (Luebke et al, 2005). Consistent with this observation, calyx endings were labeled with ␣-BTX Wackym et al, 1995;Dailey et al, 2000).…”

“…There is evidence that ␣9/␣10 is present in mammalian type II hair cells (Elgoyhen et al, 2001;Cristobal et al, 2005;Luebke et al, 2005), yet inhibition is never seen in mammalian vestibular afferents (Goldberg and Fernandez, 1980;Marlinski et al, 2004). As our results with SK blockers show, an ␣9/␣10 receptor not linked to the activation of an SK channel can result in excitation.…”

The α10 nicotinic acetylcholine receptor subunit is required for normal synaptic function and integrity of the olivocochlear system

“…Acetylcholine (ACh), the principal neurotransmitter released by OC terminals (8), binds to postsynaptic nicotinic acetylcholine receptors (nAChRs) leading to calcium influx, activation of small-conductance calciumactivated potassium channels, and subsequent hair cell hyperpolarization (9 -17). As with electrical stimulation of the olivocochlear bundle (10), the result of OHC hyperpolarization is to reduce auditory afferent output via suppression of basilar membrane motion (18).Combined immunohistochemical (19,20), electrophysiological (9, 21-23), molecular biological (7, 24), and in situ hybridization studies (21,(25)(26)(27) suggest that the nAChR subtype present at efferent hair cell synapses is assembled from both ␣9 and ␣10 subunits. Although ␣10 subunits do not form ACh-gated ion channels, ␣9 subunits form functional homomeric nAChRs in Xenopus oocytes (25).…”

“…Combined immunohistochemical (19,20), electrophysiological (9, 21-23), molecular biological (7, 24), and in situ hybridization studies (21,(25)(26)(27) suggest that the nAChR subtype present at efferent hair cell synapses is assembled from both ␣9 and ␣10 subunits. Although ␣10 subunits do not form ACh-gated ion channels, ␣9 subunits form functional homomeric nAChRs in Xenopus oocytes (25).…”

Expression of nicotinic acetylcholine receptor subunit ?9 in type II vestibular hair cells of rats

“…ChAT and AChE activity were found in efferent terminals contacting the basal area of VHC II and the afferent chalices surrounding VHC I. Cholinergic receptors belong to 2 different classes: muscarine cholinergic receptors and nicotinic cholinergic receptors (nAChR). By using various nucleic acid and ligand-binding methods, nAChR were also localized in vestibular endorgans [14][15][16][17][18][19] . A total of 17 nAChR subunits (α1-α10, β1-β4, γ, δ, and ε) have been identified in vertebrate species which can coassemble to generate a wide variety of nAChR [20] .…”