Aim The molecular interactions between transient receptor potential vanilloid subtype 4 channels (TRPV4) and cell junction formation were investigated in the human and mouse urogenital tract. Materials and Methods A qualitative study was performed to investigate TRPV4 channels, adherence junctions (AJs) and tight junctions (TJs) in kidney, ureter and bladder tissues from humans and wild‐type and transgenic TRPV4 knockout (−/−) mice with immunohistochemistry, Western blotting, immunoprecipitation and reverse trasnscription‐PCR. Cell junction formation in the wild‐type and TRPV4 knockout (−/−) mouse was evaluated with immunohistochemistry and transmission electron microscope (TEM) techniques. Results TRPV4 channels are predominantly located in membranes of epithelial cells of the bladder, ureter and the collecting ducts of the kidney. There is a molecular interaction between the TRPV4 channel and the AJ. TEM evaluation showed that AJ formation is disrupted in the TRPV4 −/− mouse resulting in deficient intercellular connections and integrity of the epithelium. Conclusions TRPV4 is believed to be a mechanoreceptor in the bladder. This study demonstrates that TRPV4 is also involved in intercellular connectivity and structural integrity of the epithelium.