Constitutive Expression of the a10 Nicotinic Receptor Subunit Alone or in Combination with a9 Overexpression Fails to Maintain Cholinergic Responses in Inner Hair Cells After the Onset of Hearing

TARANDA J; BALLESTERO J; HIEL H; DE SOUZA F; GOMEZ CASATI ME; SAVINO J; LIPOVSEK M; RUBINSTEIN M; KATZ E; VETTER, D.E; ELGOYHEN AB

Baltimore, Maryland, USA

Congreso; 32nd Midwinter Meeting, Association for Research in Otolaryngology; 2009

Association for research in otolaryngology (ARO)

Efferent inhibition of cochlear hair cells is mediated by a9a10 nicotinic cholinergic receptors (nAChRs) functionally coupled to calcium-activated, small conductance (SK2) potassium channels. Before the onset of hearing (about day 12 in rats and mice) efferent fibers transiently make functional cholinergic synapses with inner hair cells (IHCs). The retraction of these fibers after the onset of hearing correlates with the cessation of transcription of the Chrna10 (but not the Chrna9) gene in IHCs. To further analyze this developmental change, we generated a strain of mice whose IHCs constitutively express a10 into adulthood by expressing the a10 cDNA under the control of the rat Pou4f3 gene promoter. In situ hybridization showed that the a10 mRNA is expressed in IHCs of 8-wk old transgenic mice, but not in wild type mice. Moreover, this mRNA is translated into a functional protein, since IHCs from P8-P10 (n=3) a10 transgenic mice backcrossed to a Chrna10-/- background (whose IHCs have no cholinergic function), displayed normal synaptic and acetylcholine (ACh)-evoked currents in patch-clamp recordings.  Thus, the a10 transgene restored nAChR function. However, in the a10 transgenic mice no synaptic or ACh-evoked currents were observed in P16-17 (n=4) IHCs, indicating developmental down-regulation of functional nAChRs after the onset of hearing, as normally observed in wild type mice. Moreover, the endogenous level of expression of the Chrna9 gene was not a limiting factor, since in double a10 and a9 transgenic mice (constitutively expressed under the Pou4f3 promoter) no synaptic or ACh-evoked currents were observed in P19-20 (n=5) IHCs. The lack of functional ACh currents correlated with the lack of SK2 currents. These results indicate that multiple features of the efferent postsynaptic complex to IHCs, in addition to the nAChR subunits, are down-regulated in synchrony after the onset of hearing, leading to lack of responses to ACh.