| AIMTo investigate the mechanisms of the enhanced endothelium-dependent vasodilatation and the changes of large conductance Ca2+-activated potassium channels (BKca channels) in vascular smooth muscle cells in thoracic aorta of the early stage streptozotocin (STZ)-induced diabetic C57BL/6J mouse.METHODSVascular muscle tension and Phenylephrine(PE)-induced rhythmic activity in the isolated thoracic aorta of mice were compared. Radioimmunity was also used to detect the metabolite of prostaglandin I2 (PGI2), 6-Keto-prostaglandin F1a (6-Keto-PGF1a), in the blood serum. Meanwhile, single vascular smooth muscle cells were isolated by Collagenase and BKca channels current were recorded by patch clamp single channel recording technique in symmetric high potassium solution. Single channel current was amplified and filtered by EPC 9 patch clamp amplifer, then the data were analysised by TAG and TACFit 4.0.9 software.RESULTS1. We have investigated the cumulative concentration-respons curves to acetylcholine (ACh) and PE in 17 weeks, 22 weeks and 28 weeks diabetic and contrl mice aorta. Then 17 weeks diabetic and control mice were selected to be studied furtherly.2. PE could induce rhythmic activity in both 17 weeks diabetic and control mice aorta but the amplitude was markedly larger in diabetic mice than in controls [(4.9 1.7) % vs (12 5) %, P<0.01]. High K+ solution-induced contraction (619 114 mg, vs 393 138 mg, P<0.01), and ACh-induced relaxation [(81 8) %, vs (56 10) %, P<0.01] were notablely enhanced in diabetic mice than those in controls. Alone NG-nitro- L-arginine methyl ester (L-NAME) or 6-(Phenylamino)-5,8-quinolinedione (LY-83583) abolished the rhythmic activity and ACh-induced relaxation in controls but only partially inhibited them in diabetic mice. Indomethacin did not affect rhythmic activity but depressed ACh-induced relaxation observably. L-NAME plus indomethacin significantly depressed the rhythmic activity and ACh-induced relaxation than alone L-NAME did (P<0.01). Furthermore tetraethylammonium plus L-NAME abolished them in diabetic mice.3. 6-Keto-PGF1a in the serum was significantly higher in 17 weeks STZ-induced diabetic mice than age-matched controls [(1.8 1.0) g/L vs (0.5 0.3) g/L, P<0.01].4. 1 mmol/L TEA depressed ACh-induced endothelium dependent relaxation in 17 weeks diabetic mice significantly and pD2 of ACh-induced relaxation decreased notably (6.266 0.374 vs 6.918 0.453, P<0.01, n=10). But TEA did not affectpD2 of ACh-induced relaxation in control mice. Simultaneously, conductance of BLCa channels in vascular smooth muscle cells decreased but probability of open of BKCa channels increased (0.508 0.277 vs 0.108 0.058, n=6, P<0.01) by decreasing mean closed time in diabetic mice (15.3 14.8 vs 132.5 98, n=6,CONCLUSIONThe mechanism that enhanced endothelium-dependent vasodilatation in STZ-induced diabetic mice is possibly due to enhanced production of PGl2 and endothelium-derived hyperpolarizing factor (EDHF) and enhanced the activity of BKca channels in vascular smooth muscle cells by decreasing mean closed time. The phenomena maybe only take place in early stage of diabetic mice. |
