Prehearing efferent-inner hair cell synaptic strength and afferent synapse maturation are not altered in mice lacking the BK channel

GRACIELA KEARNEY; LUCAS VATTINO; DANIËL O. J. REIJNTJES; CAROLINA WEDEMEYER; ANDREA MEREDITH; SONJA PYOTT; ANA BELÉN ELGOYHEN; ELEONORA KATZ

Baltimore

Congreso; 40th Annual MidWinter Meeting; 2017

Association for Research in Otolaryngology (ARO)

Before the onset of hearing, inner hair cells (IHCs) receive efferent input from cholinergic medial olivocochlear (MOC) neurons. This input inhibits spontaneous activity of the IHCs and is thought to refine maturation of the afferent auditory system. Efferent inhibition before the onset of hearing has been hypothesized to drive pruning of supernumerary afferent (ribbon) synapses contacting the IHCs. However, the evidence to date has been limited and contradictory. Recent work combining pharmacology, electrophysiology, and immunofluorescence shows that L-type VGCCs suppress ACh release by activating BK channels expressed at MOC terminals. BK channel activation likely suppresses release by curtailing the duration of the terminal action potential. These observations suggest that mice lacking the BK channel (BKα-/- mice) should have enhanced efferent inhibition of the IHCs and, therefore, provide an in vivo model in which to test the hypothesis that efferent inhibition of IHCs contributes to the pruning of supernumerary afferent synapses. To probe the role of BK channels in efferent synaptic function, we isolated cochleas from BK channel knockout mice and their wild-type littermates (BKα−/−, BKα+/+) at P9-11 and measured evoked release during whole-cell voltage-clamp recordings from IHCs. We found no significant differences between genotypes in quantum content (BKα+/+ 0.69±0.17, BKα−/− 0.69±0.14, p=0.97; n=10) nor in the short term plasticity pattern upon high frequency stimulation. In addition, the number and size of IHC ribbons were tracked by immunofluorescence. In both genotypes, ribbons are smaller and greater in number at P6 compared to their mature morphology at P21. The number of ribbons/IHC decreased from P6 to P21 from 33 ± 1 (n=3) to 21 ± 1 (n = 4) in wildtype mice and from 34 ± 2 (n = 3) to 22 ± 0.7 (n = 5) in knockout mice. The size of individual ribbon synapses increased from P6 to P21 from 0.24 ± 0.03 (n = 4) to 0.33 ± 0.03 (n = 4) µm2 in wildtype mice and from 0.23 ± 0.03 (n = 3) to 0.33 ± 0.02 (n = 4) µm2 in knockout mice. These results suggest that the lack of BK channels at efferent terminals might be compensated for by other regulatory mechanisms that prevent synaptic strength and pruning of supernumerary afferent synapses from being altered by this genetic modification. Support:UBA&ANPCyT: EK,ABE; UMCG:SJP