Goreisan (GRS) is a Kampo medicine that is widely prescribed for edema, migraine and diarrhea. Previous studies reported GRS to have diuretic effect and to mediate body‘s water balance. Aquaporin 2 (AQP2) is a water-selective channel that plays a crucial role in regulating water reabsorption in the kidneys, influencing the body‘s water content regulation. AQP2 on the apical membrane of the collecting duct facilitates water reabsorption via the cAMP–PKA signaling pathway. Thus, this study aimed to investigate the effects of GRS on AQP2 localization in a forskolin (FSK)-stimulated mouse inner medullary collecting duct (mIMCD-3) cell.
This study revealed that GRS dose-dependently inhibited FSK-induced cAMP production and increased intracellular Ca²⁺ concentration. A selective calcium-sensing receptor (CaSR: Gi, Gq protein-coupled GPCR) inhibitor, NPS-2143, partially counteracted the effect of GRS on cAMP production and intracellular Ca²⁺ concentration, indicating that GRS acts as an agonist for the CaSR. To evaluate AQP2 translocation, spheroids that recapitulate the luminal structure of the collecting ducts were established using a 3D cell culture. The colocalization area between the lumen (ZO-1⁺ area) and phosphorylated AQP2 (Ser 269), which is phosphorylated with translocation, was calculated to assess AQP2 trafficking. Pretreatment with GRS partially inhibited FSK-stimulated AQP2‘s trafficking into the lumen. These findings suggest that the diuretic effect of GRS is attributed to its ability to modulate AQP2 localization via the CaSR.