Phylogenetic differences in agonist efficacy of á9á10 nicotinic cholinergic receptors

MOGLIE M; LIPOVSEK M; MOSKOWITZ H; FUCHS PA; ELGOYHEN AB

Congreso; Nicotinic Acetylcholine Receptors 2014, Wellcome Trust Scientific Meetings; 2014

Wellcome Trust

The á9 and á10 nicotinic acetylcholine receptor (nAChR) subunits are expressed in cochlear hair cells. They form the receptor that mediates efferent inhibition of the activity of these cells, a mechanism thought to modulate the dynamic range of hearing. It is a unique receptor formed by two alpha subunits with a characteristic mixed nicotinic-muscarinic pharmacological profile. An evolutionary analysis of both subunits has shown that á10 exhibits signatures of positive selection only along the mammalian lineage, strongly suggesting the acquisition of a novel function (Franchini & Elgoyhen, 2006). These differences in their evolutionary history correlate with the divergent calcium permeability found between rat and chicken á9á10 nAChRs (Lipovsek et al., 2012). Here, we assayed the functional consequences of the acquisition of non-synonymous substitutions in mammalian á10 subunits on agonist efficacy. Recombinant á9 and á10 subunits cRNA were synthesized in vitro and micro-injected in Xenopus Laevis oocytes. Responses to drugs were measured using two microelectrode voltage clamp technique. Cholinergic agonists such as Choline (Ch) and DMPP showed higher efficacy on chicken á9á10 receptors (percentage of maximal response to acetylcholine (ACh): Ch:88±6%;n=6, DMPP:32±3%;n=5) compared to rat á9á10 nAChRs (Ch:37±2% ; n=10, DMPP:0,6±0,3%;n=6). Accordingly, responses to Ch were 72±3% (n=3) of the maximal response to ACh in chicken hair cells. Finally, responses to Ch in oocytes expressing a Ratá9-Chickená10 hybrid receptor, rendered a relative maximal response to ACh that was similar to that of chicken á9á10 nAChRs (87±3%; n=4). The evidence presented so far clearly supports the role of á10 subunits as main determinants of agonist efficacy. Thus, the non-synonymous substitutions accumulated on mammalian á10 might have resulted in differential synaptic dynamics at the efferent-hair cell synapse.