The contributions of aquaporin-4 to water exchange across the blood-brain barrier measured by filter-exchange imaging.

Purpose: Water exchange across the blood-brain barrier (WEX) is a promising biomarker for assessing the blood-brain barrier (BBB) integrity. However, the physiological mechanisms governing WEX remain unclear. This study was conducted to investigate the contribution of Na/K-ATPase (NKA) on the luminal side of endothelial cells and aquaporin-4 (AQP4) to WEX.

Methods: WEX was measured using filter-exchange imaging for BBB assessment (FEXI-BBB) on rats, and data were fitted using an adapted two-compartment crusher-compensated exchange rate (CCXR) model. Test-retest reliability of the vascular water efflux rate constant (k) was assessed. Ouabain and 2-(nicotinamide)-1,3,4-thiadiazole (TGN-020) were administered to inhibit NKA on the luminal side of endothelial cells and AQP4, respectively, to investigate their roles in WEX measured by FEXI-BBB.

Results: Fixing intravascular diffusivity in the two-compartment CCXR model significantly improved estimation accuracy and precision of k and other parameters. The test-retest experiment demonstrated that this method had good reproducibility in measuring k (intraclass correlation coefficient = 0.79). Administering TGN-020, which inhibits AQP4, significantly decreased k by 32% (k = 3.07 ± 0.81 s vs. 2.09 ± 1.10 s, p < 0.05). However, the ouabain-treated group showed no significant change in k compared with that of the control group (2.51 ± 0.58 s vs. 2.37 ± 1.02 s, p = 0.73) in the NKA inhibition experiment.

Conclusions: WEX decreased by 32% after administering TGN-020, but no downward trend was noted after administering ouabain. Our findings indicate that AQP4 expression/function, but not NKA activity on the luminal side of endothelial cells, plays a significant role in regulating WEX.