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. 2018 Oct 1;9:4015. doi: 10.1038/s41467-018-06307-w

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 7 — IHC exocytosis is unaffected in adult Myo7a^fl/flMyo15-cre^+/- mice but efferent input is altered. a−d Calcium current (I_Ca) and corresponding changes in membrane capacitance (ΔC_m) recorded from IHCs of P16-P17 (a, b) and P48-P49 (c, d) control Myo7a^fl/fl (black) and knockout Myo7a^fl/flMyo15-cre^+/- mice (red). Recordings were obtained in response to 50 ms voltage steps (10 mV increments) from −81 mV and using 1.3 mM extracellular Ca²⁺ and at body temperature. For clarity, only maximal responses at −11 mV are shown. Panels b and d shows average peak I_Ca (bottom) and ΔC_m (top) curve from control Myo7a^fl/fl and Myo7a^fl/flMyo15-cre^+/- IHCs. e, f Membrane currents recorded from adult IHCs of control Myo7a^fl/fl (e) and Myo7a^fl/flMyo15-cre^+/- (f) mice before (left panels) and during superfusion of ACh (right panels). ACh elicited an instantaneous current only in the Myo7a^fl/flMyo15-cre^+/- IHC. g Steady-state slope conductance of the ACh-sensitive current (g_{ACh-sensitive}) at different age ranges in control Myo7a^fl/fl (black symbols) and Myo7a^fl/fl Myo15-cre^+/- (red symbols) IHCs. The isolated g_{ACh-sensitive} was obtained by subtracting the control current (left panels in e, f) from that in the presence of 100 μM ACh (right panels)³⁵. Number of IHCs is shown near the symbols (2 Myo7a^fl/fl mice at P22 and P33-P49; 3 Myo7a^fl/flMyo15-cre^+/- mice at P22; 5 Myo7a^fl/flMyo15-cre^+/-mice at P34-P48). h–k Voltage-clamp recordings obtained from adult IHCs of control Myo7a^fl/fl (h, P36) and Myo7a^fl/flMyo15-cre^+/- (i, j, P36; k P39) mice during the superfusion of 40 mM extracellular K⁺. Synaptic currents were only evoked in Myo7a^fl/flMyo15-cre^+/- IHCs. Panel j shows an expanded time scale of the blue area shown in panel (i). Panel k shows the effect of 1 μM strychnine on the ACh-induced synaptic currents. l, m TEM showing the IHC synaptic region from P37 control Myo7a^fl/fl (l) and Myo7a^fl/flMyo15-cre^+/- (m) mice. IHCs from Myo7a^fl/fl mice showed the characteristic synaptic organization of mature cells with efferent terminals forming axo-dendritic contact with the afferent fibers, as seen by the presence of active zones (l, black arrows). Scale bars, 1 μm. In Myo7a^fl/flMyo15-cre^+/- mice efferent fibers make direct axo-somatic contact with the IHC (m), which is characteristic of pre-hearing IHCs. Average data is shown as mean ± SEM