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Transcriptome analysis of cell types in the lateral superior olive sound localization network using PatchSeq

Dr. Bradley Winters (Anatomy and Neurobiology, Hearing Research Group, NEOMED)

The superior olivary complex in the brainstem of mammals compares information from the two ears (interaural) and extracts cues used for sound localization. These circuits are critical for selective attention and are altered in children with auditory processing disorder. Principal neurons (PNs) of the lateral superior olive nucleus (LSO) are a key component of this system that compare excitatory inputs driven by the ipsilateral ear with inhibitory inputs driven by the contralateral ear. The textbook view of the LSO is that it extracts interaural level differences (ILDs). Recent evidence challenges this thinking, suggesting a major function of the LSO is encoding interaural time differences (ITDs) for amplitude modulations and transient broadband sounds that are common in nature. ILD and ITD coding strategies place disparate demands on the cellular properties of LSO neurons. Furthermore, there is cellular diversity among LSO PNs that is not well understood including differences in transmitter systems, projection patterns, tonotopic distribution, and firing response type. Our overarching hypothesis is that there is functional segregation within this diversity for different binaural coding strategies.

A substantial portion of LSO PNs are inhibitory (I) and largely project ipsilaterally. The remainder are excitatory (E) and have variable projections. A major gap in our knowledge is that differences between these I/E cell types have not been examined. Our hypothesis is that their cellular properties are tuned for a form of ILD/ITD extraction, and that transmitter system and projection differences provide a means to segregate this information in upstream centers. To efficiently target I/E neurons and answer these questions, we will use knock-in reporter mice to target LSO cell types using fluorescent signals and characterize their electrophysiological and morphological properties. Preliminary studies in mice show that indeed I/E LSO neurons have large differences in excitability and dendritic morphology that would impact how they integrate sound information. Additionally, there are two action potential (AP) firing types observed within the transmitter types. To better understand what underlies these differences we propose to implement messenger RNA sequencing (RNAseq) in individual characterized LSO neurons using PatchSeq. Following patch-clamp recordings and imaging, the cell contents is drawn into the pipette and mRNAs present are quantified and sequenced. PatchSeq is an innovative and less biased approach to understanding what makes cell types different. These data will allow us to design more targeted electrophysiological studies of ion channel systems that are differentially expressed in LSO cell types. This method may also provide insights into morphological differences that cannot be addressed using electrophysiological methods.