Metabotropic glutamate receptor type 7 dimers at inner hair cell ribbon synapses of the mammalian cochlea: protection against hearing loss

• Applicant: Dr. Lisa Janina Klotz-Weigand, Ph.D.
• Subject Area: Molecular and Cellular Neurology and Neuropathology
• Term: since 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 517969465

Hearing impairment is often preceded by neurodegeneration at inner hair-cell (IHC) ribbon synapses in the cochlea. It was suggested that the primary cause of synapse loss during noise and aging in the cochlea is caused by glutamate overexposure. Pre-synaptically localized metabotropic glutamate receptors (mGluRs) are well suited to protect neurons from glutamate excitotoxicity since they can limit pre-synaptic glutamate release. Interestingly, mGluR7 is the only mGluR type that has been correlated with age-related and noise‐induced hearing deficits. Recently, we analysed expression and localization of the mGluR7 isoforms mGluR7a and mGluR7b in mouse cochlear wholemounts, using confocal microscopy and 3D reconstructions. We observed a pre-synaptic localization of both mGluR7 isoforms at the IHC ribbon synapse and a reduced amount of these receptors at synapses encoding higher frequencies and in older animals. Besides, we also showed a pre-synaptic localization of mGluR4 and mGluR8b there. MGluR7 represents the mGluR with lowest affinity to glutamate, therefore we hypothesize that activation of mGluR7 by excessive glutamate could protect the IHC ribbon synapses from noxious stimuli. For proper function mGluRs need to assemble into homo- or heterodimers. Various studies described the formation of heterodimers of mGluR types that revealed distinct characteristics. In this grant, we will therefore first describe the molecular composition of the pre-synaptic mGluR7 isoforms, using a set of complementary anatomical, biochemical and pharmacological techniques including immunohistochemical co-stainings, proximity ligation assays on cochlear wholemounts and co-immunoprecipitations (brain and cochlear tissue). For selected mGluR dimers the obtained data will be complemented by time-resolved FRET (TR-FRET) measurements. Our preliminary data showed a co-localization of mGluR7 isoforms with mGluR4 or mGluR8b, suggesting a heterodimerization. Furthermore, immunoprecipitations showed that all receptor combinations formed heterodimers when co-expressed in HEK-293 cells, while only some of them were detectable in native brain lysates. During a research stay in Montpellier, I performed initial TR-FRET measurements of the mGluR7b/8b heterodimer that indicated a functional receptor heterodimer. During a research stay in Göttingen, I learned the measurement of Ca2+-currents by whole-cell patch-clamp recordings in the cochlea. It has been described that mGluR activation can limit Ca2+ currents and thereby also pre-synaptic glutamate release. Therefore, we will pharmacologically manipulate the activity of mGluR7 dimers. To support our hypothesis that pre-synaptic mGluR7 receptors could protect the IHC ribbon synapses from noxious stimuli, we will characterize the hearing phenotype of mGluR7 knockout animals. The characterized mGluR7 dimers might represent a new molecular target suited for pharmacological concepts that protect the cochlea against noxious stimuli.

• DFG Programme: Research Grants