Elsevier

Vascular Pharmacology

Volume 46, Issue 4, April 2007, Pages 271-277
Vascular Pharmacology

Relaxant effects of danshen aqueous extract and its constituent danshensu on rat coronary artery are mediated by inhibition of calcium channels

https://doi.org/10.1016/j.vph.2006.10.011Get rights and content

Abstract

In this study, we have investigated the actions of danshensu, an active, water-extractable component of the medicinal herb danshen (Salvia miltiorrhiza), on rat isolated coronary artery rings precontracted with 1 μM 5-hydroxytryptamine (5-HT) and its action compared to the water-extractable fraction of the herb. Extraction of the water-soluble fraction from danshen (S. miltiorrhiza) provided yield of 17.5% (35 g/200 g). The amount of danshensu determined in the crude danshen herb and in its aqueous fraction was 0.45 mg/g (0.045%) and 3.28 mg/g (0.33%). The danshen aqueous extract was 13 times less potent than danshensu in relaxing 5-HT-precontracted coronary artery rings; IC50 values were 930.3 ± 133.5 μg/ml and 71.5 ± 11.0 μg/ml. Removal of the endothelium did not significantly affect their vasodilator potencies; IC50 values were 842.1 ± 123.8 μg/ml and 84.8 ± 8.8 μg/ml. On the other hand, a potassium channel inhibitor tetraethylammonium (TEA, 10 mM) shifted their concentration–response curves by 1.7 and 2.2 folds. The possible involvement of Ca2+ channels was investigated in artery rings incubated with Ca2+-free buffer and primed with 1 μM 5-HT or 60 mM KCl for 5 min prior to addition of CaCl2 to elicit contraction. In 5-HT-primed preparations, the CaCl2-induced vasoconstriction was abolished by 2 mg/ml danshen aqueous extract and 200 μg/ml danshensu, whereas, in KCl-primed preparations, 10 mg/ml danshen aqueous extract and 600 μg/ml danshensu were required to abrogate the vasoconstriction. These findings suggest the vasorelaxant actions of danshen aqueous extract and danshensu were produced by inhibition of Ca2+ influx in the vascular smooth muscle cells. The opening of K+ channels had a minor contribution to the response, but endothelium-dependent mechanisms were not involved.

Introduction

Danshen is the dried root of Salvia miltiorrhiza that belongs to the family of Labiatae. It is a traditional Chinese medicine commonly used for the treatment of cardiovascular diseases such as angina pectoris, myocardial infarction and stroke (Ji et al., 2000). The cardio-protective efficacy of danshen has been studied in ischaemia–reperfusion experiments in animals (Fung et al., 1993, Ji et al., 2000, Kuang et al., 1996, Wu et al., 1993, Zhou and Ruigrok, 1990). The mechanism may involve the ability of danshen to enhance antioxidant defense enzyme activities to decrease or abolish the production of free radicals (Ji et al., 2003). Danshen has also been shown to attenuate the increase in intracellular calcium induced by anoxia–reoxygenation in isolated ventricular myocytes, which would decrease the transformation of xanthine oxidase from xanthine dehydrogenase to reduce the production of oxygen-free radicals (Cao et al., 2003). In addition, danshen lowered the viscosity of whole blood, accelerated electrophoresis of red blood cells, and improved peripheral circulation (Chen, 1981). The vasodilator and hypotensive actions of danshen probably contributed to these effects (Kamata et al., 1993, Lei and Chiou, 1986b, Li et al., 1990).

The content of danshen can be separated into lipid-soluble and water-soluble fractions. Its lipid-soluble fraction contains more than 30 diterpenoid tanshinones; the major active constituents include tanshinone I, IIA, B, cryptotanshinone, dihydrotanshinone I, methylenetanshinone, and isotanshinone IIA (Chang et al., 1990). The putative active components of its aqueous extract are danshensu, salvianolic acid B, lithospermic acids, protocatechuic acid, and rosemarinic acid (Kohda et al., 1989, Liu et al., 1992, Kamata et al., 1993, Chan et al., 2004). However, since the water decoction is the commonly used preparation/method in preparing Chinese folk medicine for human consumption, current interest in studying traditional Chinese medicine is on the aqueous fraction and ingredients.

In animal studies, danshensu has been shown to possess many of the actions of the danshen herb. Hence, it was reported to dilate coronary arteries (Dong and Jiang, 1982), inhibit platelet aggregation (Li et al., 1983), improve microcirculation (Cheng et al., 1987), and protect the myocardium from reperfusion injury of the ischaemic heart (Tang et al., 1989). The mechanism for some of its observe activity may be related to inhibition of Ca2+ aggregation in cardiac cells and prevention of Ca2+ overload (Cao et al., 2003). In addition, danshensu has been found to be able to scavenge oxygen-free radicals (Su et al., 1992, Zhao et al., 1996), inhibit myocardial cell apoptosis (Liu et al., 2001), and protect the endothelial cells against homocysteinemia (Chan et al., 2004). In the present study, we have extracted the aqueous fraction from a danshen herb and quantified contents of danshensu in this extract and in the crude danshen herb. We have also compared the actions of danshen aqueous extract and danshensu on coronary arteries isolated from rats.

Section snippets

Materials and methods

Experiments were performed on male Sprague–Dawley rats (250–300 g) bred and kept by the Laboratory Animal Services Centre of the Chinese University of Hong Kong. All experiments were performed under licence from the Government of the Hong Kong SAR and endorsed by the Animal Experimentation Ethics Committee of the Chinese University of Hong Kong.

Effects on 5-HT-precontracted tone

The precontracted tone produced by 1 μM 5-HT was 0.32 ± 0.05 g, which was sustained over the course of the experiment. Cumulative dosing of danshen aqueous extract and danshensu produced a concentration-dependent relaxation of the 5-HT-precontracted tone with IC50 values of 930.3 ± 133.5 μg/ml and 71.5 ± 11.0 μg/ml (P < 0.001, Fig. 1). Removal of the endothelium did not significantly affect their vasodilator potencies; IC50 values were 842.1 ± 123.8 μg/ml and 84.8 ± 8.8 μg/ml. On the other hand,

Discussion

The present study has demonstrated that the water-soluble fraction of danshen dried herb and one of its constituents, danshensu, have dilatory action on rat coronary arteries. Their vasorelaxant action was produced primarily by inhibition of Ca2+ influx in the vascular smooth muscle cells and a small component was mediated by the opening of K+ channels. The present findings do not support involvement of endothelium-dependent mechanisms in their action.

Previously, we have demonstrated a biphasic

Acknowledgements

The authors thank Ms. Ethel Ng for her technical assistance. Provision of financial support for the present study by The Chinese University of Hong Kong is acknowledged.

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