| Budget Amount *help |
¥2,800,000 (Direct Cost: ¥2,800,000)
Fiscal Year 2000: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1999: ¥1,600,000 (Direct Cost: ¥1,600,000)
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| Research Abstract |
The purposes of the study were to evaluate the role of NO, PGI_2 and cytochrome P450 metabolites on acetylcholine (ACh) induced vascular hyperpolarization, and to clarify the effect of advanced glycation end products (AGEs) on the regulation of the membrane potential of vascular smooth muscle cells in coronary arterial microvessels (CAMs). Methods : One end of an isoleted canine CAM (70-300μm) was cannulated and the other was ligated. CAMs were pressurized at 60 cm H_2O in an organ-chamber which was perfused with Krebs' solution containing 0.25μM of bis-(1,3-dibutylbarbituric acid)trimethionine oxonol (K_<BIS>), a non-toxic membrane potential-sensitive fluorescent dye. Fluorescent images of CAMs were observed through a microscope with an image intensifier, and recorded with a video system. The fluorescence intensity of CAMs was measured using NIH image software on a personal computer. In protocol 1, CAMs were divided into three groups (C, I, and OD). In group C (n=9), ACh (10^<-7> M to 10^<-5> M) was added to K_<BIS> without any inhibitors. In group I (n=10), CAMs were incubated in K_<BIS> with L-NNA (10^<-4> M) and indomethacin (IND, 10^<-5> M), and then ACh was added to K_<BIS>. In group OD (n=1), CAMs were incubated with L-NNA, IND, and 17-ODYA (5×10^<-6> M), and then ACh was added. In protocol 2, CAMs were incubated with AGEs (100 μg/ml) or with its vehicle (MEM sol with L-NNA and IND) for 90 minutes or 24 hrs, and then ACh was added. In protocoll, the ACh-induced hyperpolarization was similar between group C and I.However, 17-ODYA significantly attenuated the ACh-induced hyperpolarization compared with group I.In protocol 2, AGEs did not affect ACh induced hyperpolarization either in the 90 minutes or 24 hrs incubated groups. These data indicate that cytochrome P450 metabolites have a crucial role in ACh induced hyperpolarization in canine CAMs but NO and PGI_2 have not.. AGEs have no acute or subacute effects on the ACh-induced hyperpolarization.
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