Flavonoids are polyphenolic compounds that exist widely in the plant kingdom. Epidemiological reports have demonstrated an association between high dietary intake of flavonoids and lower incidence of heart diseases. Among the daily intake of flavonoids, about 60–75% is quercetin. Quercetin presents in fruits and vegetables mainly as glycosides such as rutin. The present study aimed to examine the effect of long-term oral administration of rutin on blood pressure in rats with and without hypertension. Male spontaneously hypertensive rats (SHR) and their normotensive controls, male Wistar Kyoto rats (WKY) were treated with rutin (10 mg/kg) or its vehicle (sodium carboxyl methyl cellulose, 1%/kg) for 14 days. They were then anaesthetized for the measurement of blood pressure with or without intravenous injections of phenylephrine (1.25, 2.5, 5 and 10 mg/kg) and acetylcholine (0.25, 0.5, 1 and 2 mg/kg). Rutin treatment did not change the blood pressure in both WKY and SHR. Phenylephrine concentration-dependently increased blood pressure and these increases were comparable in WKY and SHR, with and without rutin treatments. On the other hand, acetylcholine significantly decreased blood pressure in a concentration-dependent manner, and these hypotensive effects were greater in WKY than in SHR. Rutin treatment restored the hypotensive effects of acetylcholine in SHR. Therefore, the present findings suggest that long-term rutin treatment does not affect normal blood pressure regulation but enhances the vasodilatory effect of acetylcholine only in hypertensive rats.