Quantitative analysis of the ribbon synapse number of cochlear inner hair cells in C57BL/6J mice using the three-dimensional modeling method

Abstract

In mammals, the ribbon synapses of cochlear inner hair cells are a synaptic structure of the first sensory nerve in the pathway of acoustical signal transmission to the acoustic center, and it is directly involved in voice coding and neurotransmitter release. It is difficult to quantitatively analyze the ribbon synaptic number only using an electron microscope, because the ribbon synaptic number is relatively limited and their location is deep. In this study, the specific presynaptic structure-RIBEYE, and non-specific postsynaptic structure-GluR 2 & 3 in C57BL/6J mouse basilar membrane samples were treated by immunofluorescent labeling. Serial section was performed on the samples using a laser scanning confocal microscope, and then the serial sections were used to build three-dimensional models using 3DS MAX software. Each fluorescein color pair indicates one synapse, so the number of ribbon synapses of inner hair cells is obtained. The spatial distribution and the number of ribbon synapses of cochlear inner hair cells were clearly shown in this experiment, and the mean number of ribbon synapses per inner hair cell was 16.10+/-1.03. Our results have demonstrated the number of ribbon synapses is accurately calculated by double immunofluorescent labeling to presynaptic and postsynaptic structures, serial sections obtained using a laser scanning confocal microscope, and three-dimensional modeling obtained using 3DS MAX software. The method above is feasible and has important significance for further exploring the mechanism of sensorineural deafness.