Quantitative image mean squared displacement (iMSD) analysis of the dynamics of Aquaporin 2 within the membrane of live cells

Nanodomains are a biological membrane phenomenon which have a large impact on various cellular processes. They are often analysed by looking at the lateral dynamics of membrane lipids or proteins. The localization of the plasma membrane protein aquaporin-2 in nanodomains has so far been unknown. In this study, we use total internal reflection fluorescence microscopy to image Madin-Darby Canine Kidney (MDCK) cells expressing aquaporin-2 tagged with mEos 3.2. Then, image mean squared displacement (iMSD) approach was used to analyse the diffusion of aquaporin-2, revealing that aquaporin-2 is confined within membrane nanodomains. Using iMSD analysis, we found that the addition of the drug forskolin increases the diffusion of aquaporin-2 within the confined domains, which is in line with previous studies. Finally, we observed an increase in the size of the membrane domains and the extent of trapping of aquaporin-2 after stimulation with forskolin. •Aquaporin-2 diffusion analysis in the plasma membrane in MDCK cells by using Photoactivated Localization Microscopy.•Image mean squared displacement (iMSD) approach revealed that Aquaporin-2 is confined within membrane nanodomains.•Increase in the size of the membrane domains and the extent of trapping of Aquaporin-2 after stimulation with forskolin was observed•iMSD analysis shows Plasma Membrane Diffusion of Aquaporin-2 increases with Forskolin treatment.