BackgroundThe primary pathological alterations of Pendred syndrome are endolymphatic pH acidification and luminal enlargement of the inner ear. However, the molecular contributions of specific cell types remain poorly characterized. Therefore, we aimed to identify pH regulators in pendrin-expressing cells that may contribute to the homeostasis of endolymph pH and define the cellular pathogenic mechanisms that contribute to the dysregulation of cochlear endolymph pH inSlc26a4−/−mice.MethodsWe used single-cell RNA sequencing to identify bothSlc26a4-expressing cells andKcnj10-expressing cells in wild-type (WT,Slc26a4+/+) andSlc26a4−/−mice. Bioinformatic analysis of expression data confirmed marker genes defining the different cell types of the stria vascularis. In addition, specific findings were confirmed at the protein level by immunofluorescence.ResultsWe found that spindle cells, which express pendrin, contain extrinsic cellular components, a factor that enables cell-to-cell communication. In addition, the gene expression profile informed the pH of the spindle cells. Compared to WT, the transcriptional profiles inSlc26a4−/−mice showed downregulation of extracellular exosome-related genes in spindle cells. Immunofluorescence studies in spindle cells ofSlc26a4−/−mice validated the increased expression of the exosome-related protein, annexin A1, and the clathrin-mediated endocytosis-related protein, adaptor protein 2.ConclusionOverall, cell isolation of stria vascularis from WT andSlc26a4−/−samples combined with cell type-specific transcriptomic analyses revealed pH-dependent alternations in spindle cells and intermediate cells, inspiring further studies into the dysfunctional role of stria vascularis cells inSLC26A4-related hearing loss.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12920-023-01549-0.