Mechanisms of Efferent Postsynaptic Assembly in Cochlear Hair Cells.
hair cells in the cochlea are innervated by efferent olivocochlear neurons that
originate in the superior olivary complex of the brainstem. These cholinergic synapses
regulate auditory sensitivity and frequency selectivity by suppressing the electromotile
activity of hair cells that amplifies sound signals transmitted to the brain.
Cholinergic signals in the postsynaptic hai... read morer cell are inhibitory, mediated by
Ca2+-permeable α9/10 nicotinic acetylcholine receptors (α9/10-nAChRs) that
are functionally coupled to small-conductance Ca2+-activated potassium (SK2) channels.
Ca2+ influx through α9/10-nAChRs rapidly activates nearby SK2 channels, resulting
in hair cell hyperpolarization and suppression of electromotile sound amplification.
Olivocochlear synapse activity in hair cells is critical for normal responses to sound
and provides protection against noise overexposure. The processes that direct the
assembly and function of nicotinic synapses in hair cells are not well understood. The
goal of the studies presented here is to identify novel proteins and mechanisms that
underlie the development of hair cell synapses and the proper localization and
functional coupling of α9/10-nAChRs and SK2 channels. We first characterize the
protein composition of postsynaptic sites in hair cells and show that this unusual
nicotinic synapse shares common components with nicotinic synapses elsewhere in the
nervous system. We also explore mechanisms and protein interactions that direct the
co-localization of α9/10-nAChRs and SK2 channels at the postsynaptic membrane.
Phenotypic comparison of mice lacking α9 or α10-nAChR subunits or SK2
channels has previously suggested that the SK2 channel is particularly important for the
functional synaptic localization of α9/10-nAChRs. In support of these prior
studies, we show that α9/10-nAChRs and SK2 channels interact with one another in a
heterologous cell expression system and demonstrate that the membrane expression of
α9/10-nAChRs is correlated with the presence and membrane expression of SK2
channels. We also identify α-actinin-1 as a binding partner of SK2 channels in
hair cells. We present evidence of two novel mechanisms by which hair cells may regulate
the nAChR-SK2 postsynaptic complex. We first demonstrate that alternative splicing of
the SK2 channel alters its protein interactions and surface expression. We further show
that Ca2+ differentially affects the interactions of SK2 splice variants with
α9/10-nAChRs and α-actinin-1, suggesting that postsynaptic Ca2+ signaling
represents another mechanism to modulate the function or localization of nAChRs and SK2
channels. Finally, we identify two additional binding partners of SK2 channels and
α9/10-nAChRs that may serve as adapter protein linkages to the postsynaptic
protein complex. Overall, our studies provide new insights into the molecular
organization of nicotinic synapses in cochlear hair cells and indicate that multiple
protein interactions and processes contribute to the coupling and regulation of
α9/10-nAChRs and SK2 channels.
Thesis (Ph.D.)--Tufts University, 2012.
Submitted to the Dept. of Neuroscience.
Advisor: Michele Jacob.
Committee: Eric Frank, Daniel Cox, and R. Keith Duncan.
Keyword: Neurosciences.read less