4.3. Cell Cultures

Primary rat IMCD cells were obtained and cultured as previously described [36,83,88,89]. MCD4 cells [34,36,90] were grown in DMEM/F-12 GlutaMAX^TM (Life Technologies, Darmstadt, Germany; #31331) supplemented with 5% (v/v) FCS and 5 µM of dexamethasone. MCD4 cells were used at 80–90% confluency. Their doubling time is ~26 h. The same batch of cells was used for the entire study. Cells were cultured up to passage 20–25. Changes in morphology and AQP2 expression were regularly monitored using an inverted microscope. Primary IMCD cells were seeded to be fully confluent (95–100%). Mycoplasma contamination of MCD4 cells was excluded prior to their freezing and storage in liquid nitrogen using commercially available kits.