Figure 1. The vasoactive agonists ATP and bradykinin induce translocation of NFATc1-GFP to the nucleus of CPAE cells.

A: NFATc1-GFP was localized to the cytoplasm of CPAE cells (upper panel). Application of extracellular ATP (5 μM, 5 min) caused nuclear translocation of NFAT in the same cells (lower panel, image recorded 20 min after ATP-application). B: Simultaneous measurements of [Ca²⁺]_i (rhod-2, grey trace) and nuclear NFAT translocation (black trace) in response to ATP stimulation ([Ca²⁺]_o=2 mM). NFAT translocation was quantified as the ratio NFAT_NUC/NFAT_CYT, normalized to the nuclear-cytosolic distribution of NFAT under basal conditions. C: Nuclear translocation was absent when ATP was applied in the absence of extracellular Ca²⁺ (ER Ca²⁺ release alone). D: Summarized data showing that nuclear translocation was dependent on [Ca²⁺]_o, sensitive to CsA (1 μM) and PKC inhibitor BIS VII (Bisindolylmaleimide VII). E: Nuclear translocation of NFAT after application of bradykinin (BK, 20 μM) under the same experimental conditions. F: Staining of the nucleus using SYTO 59 identified cytoplasmic localization of NFAT-GFP in non-stimulated cells (a through c) and nuclear localization of NFAT-GFP in stimulated (5 μM ATP, [Ca²⁺]_o = 2 mM) cells (d through f). Traces represent raw fluorescence intensity profiles of GFP and SYTO 59 fluorescence across the nucleus and adjacent cytoplasmic regions, normalized to maximum fluorescence. CCE: capacitative Ca²⁺ entry; CsA: cyclosporin A; ER: endoplasmic reticulum. *, statistically significant (P < 0.05) compared to control. Here and elsewhere: numbers in parentheses indicate number of individual cells tested.