NMDA expression on brain microvascular ECs. (A) Flow cytometry and (B) Western blotting independently confirm expression of NMDAr subunit 1. Antibody control in panel A is fluorescent secondary antibody only. (C) In response to chronic Hcy treatment (3 days, 20μM), NMDAr subunit 1 expression was significantly induced on the luminal surface of EC membranes. (D) Chronic treatment with Hcy (3 days, 20μM) and NMDA (3 days, 25μM) decreased expression of claudin-5 using whole-cell ELISA. Cotreatment of Hcy plus MK-801 (3 days, 20μM + 25μM) rescued the Hcy-mediated decrease in claudin-5 expression, which was not different from Ctrl or MK-801 alone.