AT1 receptors and L-type calcium channels:: functional coupling in supersensitivity to angiotensin II in diabetic rats

Objective: The study was designed to investigate the role of calcium channels in enhanced angiotensin II (Ang II)-induced contraction in thoracic aortic rings from diabetic rats. Methods: Ang II-induced isometric tension was studied in thoracic aortic rings isolated from control or streptozotocin-induced (8 weeks) diabetic rats. Saturation binding studies at AT(1) receptors and L-type calcium channels were performed using [H-3] Ang II and [H-3] PN200110, respectively. Ang II-induced calcium influx was studied in fura-2-loaded single vascular smooth muscle cells isolated from thoracic aorta of control and diabetic rats. Results: Ang II did not induce contraction in calcium-free Krebs. In the presence of extracellular calcium, increased E-max (mg/mm(2)) and pD(2) to Ang II was observed in aortic rings from diabetic (795.54 +/- 38.19; 8.27 +/- 0.12) compared to control (230.09 +/- 25.45; 7.68 +/- 0.22) rats, respectively. Nimodipine but not verapamil or diltiazem dose-dependently blocked the Ang II-induced contractions in a noncompetitive manner and its -log IC50 was significantly lower in aortic rings from diabetic (8.81 +/- 0.10) compared to control (9.34 +/- 0.11) rats. The Ang II-induced increase in intracellular calcium levels was significantly enhanced (2.5-fold) in vascular smooth muscle cells from diabetic rats. AT(1) receptor saturation binding with [H-3] Ang II revealed a significantly higher affinity (nM) and B-max (pmol/mg protein) in aortic vascular membrane preparation from diabetic (0.31 +/- 0.04; 64.18 +/- 2.4) compared to control (0.52 +/- 0.02; 47.81 +/- 2.8) rats, respectively, while L-type calcium channel saturation binding with [H-3] PN200110 showed a higher affinity (nM) with no change in the B-max (fmol/mg protein) in diabetic (0.74 +/- 0.08; 4.52 +/- 0.40) compared to control (1.49 +/- 0.32; 5.43 +/- 0.60) aortic membranes, respectively. Conclusions: Our results suggest that Ang II-induced contraction is dependent on extracellular calcium, and enhanced functional coupling of AT, receptors and DHP-sensitivc L-type calcium channels results in supersensitivity to Ang II in thoracic aorta isolated from diabetic rats. (C) 2004 European Society of Cardiology. Published by Elsevier B.V. All rights reserved.