Copyright © 2004, European Society of Cardiology
AT1 receptors and L-type calcium channels: functional coupling in supersensitivity to angiotensin II in diabetic rats
Cardiovascular and Receptorology Laboratory, Department of Pharmacology and Toxicology, National Institute of Pharmaceutical Education and Research (NIPER), Sector-67, S.A.S. Nagar, Punjab-160062, India
* Corresponding author. Tel.: +91 172 2214697, +91 172 2214690; fax: +91 172 2214692. Email address: Kumar.Arun{at}pharma.med.uni-giessen.de ramaraop{at}yahoo.com
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 AT1 receptors and L-type calcium channels were performed using [3H] Ang II and [3H] 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 Emax (mg/mm2) and pD2 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. AT1 receptor saturation binding with [3H] Ang II revealed a significantly higher affinity (nM) and Bmax (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 [3H] PN200110 showed a higher affinity (nM) with no change in the Bmax (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 AT1 receptors and DHP-sensitive L-type calcium channels results in supersensitivity to Ang II in thoracic aorta isolated from diabetic rats.
KEYWORDS Angiotensin II; L-type calcium channel; Thoracic aorta; Diabetes; Streptozotocin
Abbreviations: Ang II, angiotensin II • AT1, angiotensin type I receptor • BAPTA-AM, 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis(acetoxymethyl ester) • Bmax, binding maxima • CCB, calcium channel blockers • CFK, calcium-free Krebs • CRC, concentration–response curve • DCC, dihydropyridine sensitive L-type calcium channels • DHP, dihydropyridine • DMEM, Dulbecco's modified essential medium • KCl, potassium chloride • Kd, dissociation constant • KHB, Krebs-Henseleit buffer • LOE-908, (R, S)-(3,4dihydro-6, 7-dimethoxy-isochinolin-1-yl)-2-phenyl-N, N-di [2-(2,3,4-trimethoxyphenyl) ethyl] acetamid mesylate • PE, Phenylephrine • STZ, streptozotocin • VSMC, vascular smooth muscle cell
1 Present address: Rudolf Buchheim Institut Für Pharmakologie, Universitätsklinikum Giessen, Frankfurter Strasse 107, D-35392, Giessen, Germany.
Time for primary review 29 days
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