Rat isolated mesenteric arteries do not reliably relax to an increase of extracellular concentration of potassium ([K⁺]_o). Mechanistic explanations for this differ between laboratories. The concentration of phenylephrine (PE) to which the arteries are exposed has been proposed to explain this discrepancy (Dora et al. 2002). Male Wistar rats (200-250 g) were killed humanely and sections of a third order branch of the superior mesenteric artery mounted for isometric tension recording in a Mulvany-type myograph containing bicarbonate-buffered physiological salt solution at 37°C. We found the concentration of PE or the resultant degree of tension development had no effect on the relaxation of arteries to an increase of [K⁺]_o. In addition, inhibition of BK_Ca by TEA (5 mM), charybdotoxin (100 nM) or iberiotoxin (100 nM) did not induce a relaxation when [K⁺]_o was increased from 5.9 to 13.8 mM. The hyperpolarization and relaxation to an increase of [K⁺]_o from 4.6 to 13.8 mM was enhanced by increasing the duration of exposure to PE. The effect of duration of exposure to the vasoconstricting agonist was less pronounced when contraction was stimulated by U46619, a thromboxane A2 agonist (PE vs U46619, n=11 P <0.01; Mann-Whitney U test). Moreover, the time dependence of the effect of PE was significantly attenuated (n = 8, P < 0.05) in the presence of diphenylboric acid 2-aminoethyl ester (2-APB; 75 μM), an IP₃ receptor inhibitor, and abolished by SKF96365 (10 μM), a store-operated Ca²⁺ channel (SOC) inhibitor (n = 8, P < 0.01). In contrast, 50 nM ryanodine, a concentration previously shown to induce the Ca²⁺ release from intracellular stores (Meissner 1986), significantly enhanced the relaxation to raised [K⁺]_o (n = 10; P < 0.005). We have previously shown that relaxation of PE-induced force of rat isolated mesenteric artery by raised [K⁺]_o is dependent upon extracellular sodium, being depressed by a reduction in extracellular sodium and enhanced by monensin, in the presence but not absence of extracellular sodium (Brochet et al. 2002; Brochet & Langton, 2003). In this study we show that blockade of BK_Ca channels does not result in relaxation to elevated [K⁺]_o. Increasing time of exposure to PE increased the amplitude and duration of the relaxation to raised [K⁺]_o. Contractions induced by U46619, a vasoconstrictor that does not release SR calcium, were significantly less susceptible to relaxation to raised [K⁺]_o than contractions induced by PE. Finally, activation of SOCs by store emptying, using low concentrations of ryanodine, augmented the [K⁺]_o-induced relaxation of PE-induced contractions. We interpret these data to suggest that augmentation the accumulation of [Na⁺]_i following activation of SOCs by PE and the depression of Na,K-ATPase activity in low [K⁺]_o both serve to potentiate the upturn in the activity of the Na,K-ATPase when [K⁺]_o is subsequently increased. This results in hyperpolarization and relaxation of the arterial smooth muscle.