In vivo investigation of sensory hair cell function and development

National Institute On Deafness And Other Communication Disorders

Investigators

Linked publications & trials

Abstract

Summary and Background: Sensory hair cells are required to reliably transmit auditory and vestibular information to the brain. While the majority of hearing loss results from the loss of hair cells, there is accumulating evidence that in cases of noise-induced and age-related hearing loss, the pathology may be due to damage or loss of hair cell synapses rather than hair cells. In these latter cases, effective clinical treatment requires the restoration of synaptic connections. In order to restore these connections, it is critical to understand how they function and how they are assembled in vivo. Our studies combine genetic, molecular, and imaging-based approaches to identify the structural and functional processes underlying synapse formation and function in hair cells. For our studies we examine neuromast hair cells in the zebrafish posterior lateral-line system (pLL) in order to study hair-cell system development and function, in a live, transparent preparation. We have an extremely powerful collection of transgenic zebrafish that label synaptic structures to either assess synaptic morphology or function using genetically encoded fluorescent proteins. We are combining these microscopy-based approaches with CRISPR technology to create mutant zebrafish in order to identify genes required for synapse formation, function and regeneration. With this knowledge we aim to apply our understanding of these processes in order to understand how to properly reform hair cells and synaptic structures when they are loss or damaged after hearing loss. This report summarizes the 10th year for the Section on Sensory Cell Development and Function. Our main focus has been publishing high quality research projects and exploring ways to expand upon and initiate new research. We apply numerous advanced microscopy approaches to explore new avenues of live functional and developmental analyses. Projects in the lab: In vivo investigation of sensory system function Based on our published work in the neuromasts of the zebrafish pLL, the majority of hair cells and synapses are silent, but the silent cells can be recruited (unsilenced) after damage. This data indicates that silent hair cells may function as failsafe to ensure sensory information is transmitted even after damage or loss of active cells. In the pLL, multiple afferent neurons (4) innervate each neuromast. Currently it is unclear if all anatomically connected afferent neurons are activated when the majority of hair cells and synapses are silent. Due to the abundance of silent hair cells, we hypothesize that not all anatomically connected afferent neurons respond to neuromast stimulation. We are currently investigating the functional circuitry between neuromasts and afferent neurons using light sheet fluorescent microscopy. Constructing this circuitry is essential for our understanding of in vivo sensory circuit function. Determine what molecules are required for hair cell synapses formation Hair cell ribbon synapses are critical for hearing and balance and loss of these synapses is linked to age- and noise-related hearing loss. Currently what molecules are required to assembly these synapses are not fully understood. We are using mouse and zebrafish models investigate the role of presynaptic Neurexins in hair cell synapse assembly. For our work we acquire high resolution of synapses in neurexin mutants. In addition, we use functional imaging to explore how pre- and post-synaptic responses are reduced in neurexin mutants. Overall, this work will identify Neurexins that are critical to assemble ribbon synapses. Understanding what moleculaes are important for ribbon synapse formation is important to understand how to reform or repair these synapses after they are lost or damaged. Determine how hair cell synapses form in vivo Hair cell ribbon synapses are defined by a presynaptic density called a ribbon. How this unique and highly specialized synapse is formed is unclear. Using Airyscan confocal microscope in live zebrafish we are investigating ribbon formation. We are using tracking analyses to monitor ribbon transport in the cell. We are also exploring the role of microtubules networks in ribbon movement during development. Overall, this work will outline a dynamic series of events that underlies formation of a critical synapse required for sensory function. Determine how synaptic vesicle reach the hair cell synapses Ribbon synapses have high rates of spontaneous vesicle release and function without fatigue. To sustain this level of release, a continuous supply of synaptic vesicles must be trafficked to the presynapse. In neurons, the motor protein Kif1a has been shown to transport synaptic vesicles along microtubules to the presynapse. We are exploring whether Kinesin-mediated transport delivers synaptic vesicles to the hair cell presynapses. For this work we are creating Kinesin mutants, as well as using pharmacology to disrupt microtubules. In addition to Kinesin- mediated transport we will also explore what contribution endocytosis has to synaptic vesicle replenishment at ribbon synapses. Overall, this work will reveal how synaptic vesicles are transported to and maintained at the hair cell synapses and highlight the significance of synaptic vesicle pools for proper hearing and balance.