In the gastrointestinal (GI) system, like in other organ systems, the histological structure is a key determinant of physiological function. Tissues form multiple layers in the GI tract to perform their specialized functions in secretion, absorption, and motility. Even at the single layer, the heterogeneous cell population performs a diverse range of digestive or regulatory functions. Although many details of such functions at the histological and cell biological levels were revealed by traditional methods such as cell sorting, isolation, and culture, as well as histological methods such as immunostaining and RNA in situ hybridization, recent advances in spatial single-cell technologies could further contribute to our understanding of the molecular makeup of GI histological structures by providing a genome-wide overview of how different genes are expressed across individual cells and tissue layers. The current minireview summarizes recent advances in the spatial transcriptomics field and discusses how such technologies can promote our understanding of GI physiology. © 2023 American Physiological Society. Compr Physiol 13:4709-4718, 2023.