Xiaomin Hou

Enhancement of voltage-gated K+ channels and depression of voltage-gated Ca2+ channels are involved in quercetin-induced vasorelaxation in rat coronary artery.

Quercetin is one of the most common flavonoids in the human daily diet. Its affects the coronary artery, especially L-type voltage-gated Ca2+ channels and voltage-gated K+ channels in the arterial smooth muscle cells, which are poorly understood. The present experiments were designed to study the myogenic effect of quercetin and its possible underlying mechanisms in the rat coronary artery. A wire myograph was used to observe the myogenic effects. Arterial smooth muscle cells were freshly isolated from the rat coronary artery and the intracellular free Ca2+ concentration was measured with molecular probe fluo-4-AM. The effects of quercetin on L-type voltage-gated Ca2+ channels and voltage-gated K+ channels were studied using a whole-cell patch clamp. Quercetin (3-30 µM) produced a depression and relaxation on the contraction induced by KCl or the thromboxane A2 analog 9,11-Dideoxy-9α,11α-methanoepoxy prostaglandin F 2α . The vasorelaxation was attenuated by 4-aminopyridine, a specific voltage-gated K+ channel inhibitor, but was not affected by the NG-nitro-L-arginine methylester ester (a nitric oxide synthesis inhibitor), glibenclamide (a ATP-activated K+ channel inhibitor), iberiotoxin (a Ca2+-activated K+ channel inhibitor), BaCl2 (an inward rectifier K+ channel inhibitor), or by endothelium denudation. At the same concentrations, quercetin reduced the KCl-induced elevation of the intracellular free Ca2+ concentration, inhibited the inward Ca2+ currents through L-type voltage-gated Ca2+ channels, and increased the outward K+ currents through voltage-gated K+ channels in the rat coronary artery smooth muscle cells. Collectively, our results demonstrate that quercetin possesses vasospasmolytic effects in RCA and suggest that depression of the Ca2+ influx through L-type voltage-gated Ca2+ channels and augmentation of voltage-gated K+ channel activity in the myocytes may underlie coronary relaxation.

Hesperetin inhibits rat coronary constriction by inhibiting Ca2+ influx and enhancing voltage-gated K+ channel currents of the myocytes

Hesperetin (HSP, one of the most common flavonoids in Citrus) has been reported to possess many benificial effects and is indicated for many diseases both as a therapeutic drug and as a supplement. Although its vascular effects have been extensively studied, little is known about its effects and the underlying mechanisms on coronary artery. In the present study, the myogenic effects of HSP were studied with a wire myograph in isolated rat coronary artery (RCA). Molecular probe and the patch clamp technique were used to study effects of HSP on intracellular free Ca(2+) concentration, inward Ca(2+) currents through L-type voltage-gated Ca(2+) channels (LVGC) and outward K(+) currents through voltage-gated K(+) channels (KV). HSP (0.01-0.1mM) concentration-dependently depressed concentration-contraction curves of both KCl and thromboxane receptor agonist 9,11-Dideoxy-9α,11α-methanoepoxy prostaglandin F2α (U46619), and relaxed RCA precontracted by the both vasoconstrictors. The vasospasmolytic effect was more potent in KCl- than in U46619-induced contraction. The vasorelaxation was attenuated by 4-aminopyridine, a specific KV inhibitor, but not affected by NG-nitro-L-arginine methylester ester, indomethacin, glibenclamide, iberiotoxin, BaCl2 or endothelium denudation. At the same concentrations, HSP inhibited extracellular Ca(2+) influx-induced contraction, reduced intracellular free Ca(2+) concentration, inhibited inward Ca(2+) currents through LVGC and increased outward K(+) currents through KV in the vascular smooth muscle cells (VSMCs) freshly isolated from RCA. Collectively, our results show that HSP is vasospasmolytic in RCA and suggest that the vasospasmolysis is mediated by inhibition of LVGC and enhancement of KV currents in RCA VSMCs.

Relaxation of Rat Aorta by Farrerol Correlates with Potency to Reduce Intracellular Calcium of VSMCs

Farrerol, isolated from Rhododendron dauricum L., has been proven to be an important multifunctional physiologically active component, but its vasoactive mechanism is not clear. The present study was performed to observe the vasoactive effects of farrerol on rat aorta and to investigate the possible underlying mechanisms. Isolated aortic rings of rat were mounted in an organ bath system and the myogenic effects stimulated by farrerol were studied. Intracellular Ca2+ ([Ca2+]in) was measured by molecular probe fluo-4-AM and the activities of L-type voltage-gated Ca2+ channels (LVGC) were studied with whole-cell patch clamp in cultured vascular smooth muscle cells (VSMCs). The results showed that farrerol significantly induced dose-dependent relaxation on aortic rings, while this vasorelaxation was not affected by NG-nitro-l-arginine methylester ester or endothelium denudation. In endothelium-denuded aortas, farrerol also reduced Ca2+-induced contraction on the basis of the stable contraction induced by KCl or phenylephrine (PE) in Ca2+-free solution. Moreover, after incubation with verapamil, farrerol can induce relaxation in endothelium-denuded aortas precontracted by PE, and this effect can be enhanced by ruthenium red, but not by heparin. With laser scanning confocal microscopy method, the farrerol-induced decline of [Ca2+]in in cultured VSMCs was observed. Furthermore, we found that farrerol could suppress Ca2+ influx via LVGC by patch clamp technology. These findings suggested that farrerol can regulate the vascular tension and could be developed as a practicable vasorelaxation drug.

Farrerol can attenuate the aortic lesion in spontaneously hypertensive rats via the upregulation of eNOS and reduction of NAD(P)H oxidase activity.

Farrerol, a typical natural flavanone, is the major active component of Rhododendron dauricum L. The objective of this study was to evaluate the attenuation effect of farrerol against the aortic lesions in spontaneously hypertensive rats (SHR) for the first time. Twelve-week-old male SHR were orally administered with farrerol (50mg/kg/day), verapamil (50mg/kg/day, positive control), or vehicle for 8 weeks (n=10 in each group). Age-matched Wistar-Kyoto rats (WKY) served as normal controls (n=10). Our results revealed that farrerol significantly reduced the systolic blood pressure in SHR (from 177±4mmHg to 158±5mmHg) and also dramatically attenuated the aortic lesion, which is characterized by decreased media thickness, wall area, media-lumen ratio, nuclei size and an increased nuclei number (P<0.05). Moreover, the levels of O2(-) along with NAD(P)H oxidase activity were reduced (P<0.05), while the activity of endothelial nitric oxide synthase (eNOS) was elevated (P<0.05) in aortic homogenates after the intervention of farrerol. Furthermore, farrerol upregulated the expression of eNOS in both of mRNA and protein levels, accompanied by the downregulated mRNA and protein expression of p22(phox) (P<0.05), an essential subunit for NADPH oxidase activity. Our findings indicated that farrerol has a significant protective effect against the aortic lesion in SHR, which may be related to the enhanced eNOS activity and reduced NAD(P)H oxidase activity.

Extracellular acidosis contracts coronary but neither renal nor mesenteric artery via modulation of H+,K+‐ATPase, voltage‐gated K+ channels and L‐type Ca2+ channels

What is the central question of this study? We asked whether there are any differences in responses to extracellular acidosis among arteries of different origin and whether K+ channels, Ca2+ channels and H+,K+‐ATPase are involved in extracellular acidosis‐induced coronary myogenic responses. What is the main finding and its importance? This report is the first to demonstrate that extracellular acidosis alkalinizes the cytosol of arterial smooth muscle cells freshly isolated from rat coronary artery and contracts the artery exclusively and to suggest that modulation of voltage‐gated K+ channels, voltage‐gated Ca2+ channels and H+,K+‐ATPase are involved in the contraction.

Apigenin relaxes rat intrarenal arteries: involvement of Cl- channels and K+ channels

The vasodilator effect of apigenin (API) was demonstrated in a number of vascular beds. We aimed to characterize the vasospasmolytic and electrophysiological effects of apigenin (API) in intrarenal arteries (IRAs). The vascular tone of male rat isolated IRAs was recorded with a myograph. Transmembrane Cl− currents through Ca2+-activated Cl− channels (CaCCs), K+ currents through voltage-gated K+ (Kv) channels and inwardly rectifier K+ (Kir) channels were recorded with patch clamp in the freshly isolated arterial smooth muscle cells (ASMCs). Preincubation with API (10-100 μM) concentration-dependently depressed the contractions induced by KCl, 9,11-dideoxy-9α,11α-methanoepoxy prostaglandin F2α (U46619), phenylephrine and vasopressin without significant preference and the IC50 values were 13.27-26.26 μM. Acute application of API elicited instant relaxations in the IRAs precontracted with these vasoconstrictors and the RC50 values were 5.80-24.33 μM. API relaxation was attenuated by chloride deprivation, CaCC blockers, Kv blocker and nitric oxide synthase inhibitor, but not by Kir blocker and cyclooxygenase inhibitor. At 10-100 μM, API depressed CaCC currents and Kir currents while enhanced Kv currents of IRA ASMCs. The present results demonstrate that API counteracts various vasoconstrictors noncompetitively and nonspecifically and suggest that modulation of CaCCs, Kv and Kir channels of IRA ASMCs is involved in its vasospasmolytic effects.

Structure‑activity associations in novel farrerol derivatives with vasorelaxant properties

To detect the structure‑activity associations of farrerol derivatives, the relaxation activity of farrerol derivatives was observed in isolated aortic rings pre‑contracted using phenylephrine in Sprague‑Dawley rats. All compounds tested in the present study produced a relaxation effect, which was significantly affected by the molecular structure. Using a collagen gel contraction assay, the present study further evaluated the inhibitiory effect of farrerol derivatives in a decreased collagen gel area, induced by Angiotensin II. The results indicated that farrerol derivatives could inhibit collagen contraction, and that the inhibitory effect was associated with the molecular structure of the compounds. Furthermore, the inhibitory strength of the different compounds was consistent with the results of vascular tension detection. The activity of the farrerol derivatives was closely associated with the molecular structure. The analysis indicated that an electron‑withdrawing substituent in the ortho position of the phenyl group (ring B) was crucial in order to observe improved vasorelaxation activity, whereas a hydroxyl or methoxy group was unfavorable. A para electron‑donating group was oberved to increase compound activity. In addition, when the B ring was heterocycle rather than a phenyl ring, the vasorelaxation ability was weakened.

MAPKs-mediated modulation of the myocyte voltage-gated K+ channels is involved in ethanol-induced rat coronary arterial contraction

ABSTRACT Acute coronary arterial spasm is contributory to ethanol‐induced heart ischemic events. The present experiments were designed to study contractive effect of ethanol in isolated rat coronary arteries (RCAs) stimulated mildly with vasoconstrictors and involvement of voltage‐gated potassium (KV) channels and mitogen‐activated protein kinases (MAPKs) of rat coronary arterial smooth muscle cell (RCASMC) in the spasm. The vascular tension was recorded with a wire myograph. KV currents of single freshly isolated RCASMC were assessed with patch clamp. Phosphorylation of RCASMC MAPKs (p38 MAPK and p44/42 MAPK) was assayed by Western blots. Mild stimulation, which was defined as 5–20% of 60mM KCl‐induced contraction, with thromboxane A2 mimetic U46619, endothelin‐1 or KCl tremendously increased contractive response of RCAs to ethanol (0.8–8.0mg/ml). Ethanol (8 and 25mg/ml) reduced the maximal KV currents by 53.6% and 56.6% respectively without concentration‐dependence. Both ethanol (8.0mg/ml) and U46619 (0.3&mgr;M) enhanced phosphorylation of p38 MAPK and p44/42 MAPK and there was a pronounced synergism between them. MAPK pathway inhibition with p38 MAPK inhibitor SB239063 and p44/42 MAPK inhibitor PD98059 depressed ethanol‐induced contraction by 22–55% in the stimulated RCAs and almost abrogated ethanol‐induced reduction in KV currents. The present results for the first time demonstrated that ethanol induces potent vasoconstriction in stimulated RCAs and depresses KV currents in RCASMCs. It is suggested that MAPKs pathways play an important role in the etiology of ethanol‐induced coronary artery diseases.