Protocol for assessing auditory function, cochlear synaptopathy, and afferent terminal morphology in laboratory mice

Here, we present a protocol for evaluating auditory function and synaptic molecular-structural integrity in mice, specifically female GluA3-knockout (GluA3^KO) mice raised in quiet or ambient sound levels. We describe steps for combining auditory brainstem response recordings, immunofluorescence, and electron microscopy. We detail procedures for assessing hearing function, the expression and localization of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-type glutamate receptor subunits, and the ultrastructure of cochlear-ribbon synapses. This protocol provides a reproducible framework for studying molecular-structural synaptopathy in mouse models of cochlear dysfunction. For complete details on the use and execution of this protocol, please refer to Pal et al.¹.