Synaptic Circuitry of Auditory Neurons

National Institute On Deafness And Other Communication Disorders

Investigators

Linked publications & trials

Abstract

The focus of the Section on Neuronal Circuitry this year has been generating data for scientific presentations and publications. Projects fall into two major categories: 1) Intrinsic electrical properties and synaptic inputs of medical olivocochlear (MOC) neurons in the brainstem, and 2) Influence of synaptic outputs of MOC neurons in the cochlea. Synaptic inputs of olivocochlear neurons in the brainstem and synaptic outputs in the cochlea Medial olivocochlear (MOC) neurons have cell bodies in the brainstem, where they receive excitatory synaptic inputs conveying sound information from the cochlea via the cochlear nucleus. Very little is known about the neuronal circuitry driving activity of MOC neurons because their cell bodies are difficult to locate in un-stained brain preparations. We completed the initial goal of the lab to develop and characterize a reliable genetic technique to perform single cell patch-clamp recordings from the MOC neurons in brain slices from mice. Using this technique, we aim to understand the diversity of synaptic inputs of MOC neurons to fully understand how the neurons are activated and modulated during auditory perception. We also aim to determine how the MOC neuron baseline electrical properties may change to contribute to neuronal hyperactivity that has been shown to occur in pathological situations such as in tinnitus or hyperacusis. Experimental accomplishments include characterization of the ChAT-IRES-Cre x tdTomato mouse line as a reliable tool for identifying MOC neurons in brainstem slices for single cell patch-clamp electrophysiology experiments. Using this mouse line, we discovered a pathway of sound-driven inhibition of MOC neurons by neurons of the medial nucleus of the trapezoid body (MNTB), and are characterizing the effect of this inhibition on MOC neuron activity. The initial report was published in January 2020 (JNeurosci). A manuscript describing the short-term synaptic plasticity of MNTB-MOC synapses is in preparation. In addition, a project investigating the integration of excitatory and inhibitory synaptic inputs to MOC neurons using a novel brain slice preparation is in the final stages of experimentation. A methods manuscript describing the technique was published in 2021 (JoVE). To complement patch-clamp electrophysiology projects using the wedge slice, we are currently developing a computational model of MOC neurons, including addition of accurate post-synaptic current waveforms and intrinsic electrical conductances from experimental data collected in our own laboratory, which will investigate the integration of excitatory and inhibitory synaptic inputs to MOC neurons to determine how sound based inhibition alters MOC function and the MOC effects on the cochlea. These experiments will elucidate the mechanisms of synaptic activation and inhibition of MOC neurons, which in turn drives their inhibition of mechanical activity in the cochlea, thus shaping the cochlear response to sound. In a collaborative project with Dr. Lisa Goodrich at Harvard University we aimed to determine the single cell transcriptome of LOC and MOC neurons. For this project, members of the Section on Neuronal Circuitry performed patch-clamp recordings from identified LOC and MOC neurons, sucked out the cell contents, then analyzed the single-cell transcriptome of the neurons using the PatchSeq methodology. We were trained to do the cDNA library preparation by members of the Goodrich Lab. The samples were then transferred to Dr. Robert Morell in the Genomics and Computational Biology Core, who performed sequencing and initial analysis of the samples. Unfortunately, this project requires many individuals to be in the lab simultaneously, and has been put on hold indefinitely due to the COVID pandemic. We have collaborated with Dr. Hui Cheng in the NIDCD statistics core for analysis of our patch-clamp electrophysiology results. Synaptic outputs of olivocochlear neurons in the cochlea MOC neurons project to the cochlea, where they decrease the mechanical movement of the basilar membrane by inhibiting cochlear OHCs. OHC activity enhances signaling to cochlear IHCs and shapes cochlear tuning curves and gain. MOC synapses onto OHCs are implicated in inhibiting OHC via coupling of cholinergic channel coupling to an SK potassium conductance. This effect is implicated in improved hearing in background noise, and protection of the cochlea against sound trauma. MOC neurons are also thought to release other neurotransmitters in the cochlea with poorly described effects on cochlear function. Recent work has detailed GABAergic responses in type II spiral ganglion neurons, while immunohistochemistry suggests a GABAergic synapse between MOC efferent and type II afferent neurons. This work was presented at the ARO midwinter meeting. In addition, a review on neurotransmitters released in the cochlea was recently published in Hearing Research (Kitcher, Pederson and Weisz 2021). VGLUTs in OHCs, and the central innervation patterns of type II cochlear afferent neurons In a now-concluded collaborative project with Dr. Rebecca Seal at the University of Pittsburgh, the specific vesicular glutamate transporters (VGLUTs) employed by OHCs to load glutamate into presynaptic vesicles for release onto spiral ganglion afferents was determined. This led to generation of mutant mice lacking the VGLUTs from either inner hair cells (IHC) or OHC, or both. Use of these mutant mice allows isolation of afferent signaling by either type of hair cell, in order to determine the unique contribution that each pathway makes to perception of sound. We used these different mouse lines in an experiment in which the animals were exposed to noise, then their cochlear nuclei assessed for activation of the immediate early gene c-Fos. We determined that the OHC-type II afferent pathway can indeed respond to acoustic signals at non-pathological levels of sound, and evoke activation of neurons in the brainstem. This work was published in JNeurosci, with Dr. Weisz as co-contributing author (Weisz et al 2021).