Fig. 1.

Depolarization of the endothelial cell (EC) membrane with ischemia in isolated lungs from wild-type (WT) and Akt-null mice. Lungs were preperfused for 30 min with the membrane potential-sensitive dye bis-oxonol (20 nM), and, where indicated, wortmannin (100 μM) or cromakalim (30 μM) were added. Images were acquired before (time 0) and at 0.5, 1, 2, 3, 4, and 5 min after the global cessation of flow (ischemia). Increased bis-oxonol fluorescence indicates cell membrane depolarization. Continuously perfused lungs (without ischemia) were used as a control. A: images were obtained at time 0 and at 0.5, 1, and 2 min of ischemia, as indicated at the top. Images are shown in pseudocolor with intensity as indicated by the sidebar. The imposed lines outline the margins of the microvessels. All images were acquired with the same exposure settings. Scale bar = 10 μm. B: fluorescence intensity quantified by MetaMorph software was plotted as a function of postischemic time from 0 to 5 min. The intensity for each time point was calculated as the mean value for six randomly selected fields. The dashed line indicates control perfusion with continuous flow (no ischemia). The plotted results represent means ± SE from 3 mice.