Figure 2.

Direct activation of TRPC6 increases lung vascular permeability. (A) WT or Trpc6^−/− MLECs were loaded with Fura 2-AM for 25 min, after which they were exposed to buffer containing 2 mM Ca²⁺ or Ca²⁺-free buffer. Changes in intracellular Ca²⁺ were determined in response to OAG by converting 340:380 excitation ratio intensity, as described in Materials and methods. Each representative tracing is the mean calcium response of 20–25 cells in a given field. These experiments were repeated three times. (B) Isogravimetric WT or Trpc6^−/− mice lungs were perfused with 100 µM OAG for 20 min, after which intravascular pressure was raised by 10 cm of H₂O. Microvascular filtration coefficient (K_f,c) was determined from the slope of lung wet weight gain after normalization by lung dry weight, as described in Materials and methods. Plot shows mean ± SEM of K_f,c from four individual experiments. * indicates a significant increase in K_f,c over the values without OAG (0 µM; P < 0.05). (C) OAG (100 µM) was administered retroorbitally into the WT or Trpc6^−/− mice vasculature for 20 min, after which Evans blue–labeled albumin was injected and transendothelial leakage was determined at 30 min. Plot shows mean ± SEM of albumin leakage from four individual experiments. * indicates values that are significantly different from Trpc6^−/− lungs without OAG or after OAG administration in Trpc6^−/− mice (P < 0.05).