Impaired relaxation to acetylcholine in 2K-1C hypertensive rat aortas involves changes in membrane hyperpolarization instead of an abnormal contribution of endothelial factors

Abstract

The contribution of endothelial factors and mechanisms underlying decreased acetylcholine-induced relaxation and endothelial inhibitory action on phenylephrine-induced contraction were evaluated in aortas of two-kidney, one-clip hypertensive (2K-1C) and normotensive (2K) rats. Relaxation induced by acetylcholine in 2K-1C precontracted by phenylephrine was lower [Maximum Effect (ME): 71.33±3.36%; pD₂: 7.050±0.03] than in 2K (ME: 95.26±1.59%; pD₂: 7.31±0.07). This response was abolished by N^G-nitro-l-arginine (L-NNA) in 2K-1C, but was only reduced in 2K (ME: 29.21±9.28%). Indomethacin had no effect in 2K-1C, and slightly attenuated acetylcholine-induced relaxation in 2K. The combination of L-NNA and indomethacin almost abolished acetylcholine-induced relaxation in 2K-1C, while in 2K, the inhibition (ME: 56.61±8.95%) was lower than the effect of L-NNA alone. During the KCl-induced precontraction, 2K and 2K-1C aortas showed similar acetylcholine-induced relaxation (43.50±5.64% vs. 41.60±4.36%), which was abolished by L-NNA. The levels of cGMP produced in response to acetylcholine were not different between 2K and 2K-1C. The sensitivity to sodium nitroprusside was lower in phenylephrine-precontracted aortas from 2K-1C than 2K, as showed by the pD₂ values (7.72±0.20 vs. 8.59±0.17), and this difference was abolished in aortas precontracted by KCl. The membrane potential was less negative in 2K-1C than in 2K (−41.57±1.19 vs. −51.00±1.13 mV) and hyperpolarization induced by acetylcholine was lower in 2K-1C than in 2K aortas (6.00±0.66 vs. 13.27±1.61 mV). Phenylephrine-induced contraction in aortas with endothelium was similar in both groups, and increased by the endothelium removal. This increase was lower in 2K-1C (from 1.32±0.06 to 1.90±0.21 g) than 2K (from 1.49±0.07 to 2.83±0.18 g). L-NNA and the endothelium removal had similar effect in 2K-1C (1.85±0.18 g) and were lower in 2K (2.18±0.20 g). Indomethacin decreased phenylephrine-induced contraction only in 2K. In conclusion, our major finding was a selective defect in smooth muscle membrane hyperpolarization, which could explain the decreased relaxation to acetylcholine and the attenuated inhibitory effect of endothelium on the contractile function in 2K-1C aortas.