CONICET | Buscador de Institutos y Recursos Humanos

The activation of α9α10 nicotinic receptors in cochlear hair cells can ameliorate acoustic trauma. Therefore, enhancing α9α10-mediated currents by means of a pharmacological potentiator may be useful in the prevention or treatment of noise-induced hearing loss. In the present work we aimed to characterized ascorbic acid (ASC) as a positive modulator in α9α10 injected X. laevis oocytes by two-electrode voltage-clamp recordings. Responses to 10μM acetylcholine (ACh) were potentiated by ASC in a concentration-dependent manner (0.1-30mM ASC). At 3mM ASC, a 64±6% (n=77) potentiation was observed. Potentiation was more pronounced at lower (250±50%, 3μM ACh, n=9) than at higher (140±40%, 1mM ACh, n=6) ACh concentrations. No significant changes in the half maximal concentration of ACh and Hill coefficients were observed in the pressence of 3mM ASC (EC50 = 18±1, nHill = 0.8±0.1, n=5-9). Oxidized ASC blocked acetylcholine (ACh)-evoked responses at a 3mM concentration (72±5% block at 1µM ACh, n=3), suggesting that reducing ASC is the active compound. Alternatively, the equally reducing stereoisomer D-iso-ASC also had a blocking effect (55±10% block at 1µM ACh, n=3), indicating that a stereospecific interaction is also necessary. To test if a redox mechanism is involved in ASC potentiation we mutated the extracellular residues CC192/193SS (Torpedo α1 numbering). This substitutions did not abolish ASC potentiation (90±10% potentiation at 1mM ACh, n=6), ruling out the participation of these residues in the mechanism. Altogether, our results indicate that ASC potentiates α9α10-mediated responses through an allosteric and/or redox mechanism which does not involve C192-C193 disulfide bond. Consequently, our results suggest that ASC has a potential therapeutic use in noise-induced hearing loss.