K⁺-induced vasodilatation contributes to active hyperaemia in cerebral vessels, where K⁺ released from active neurones relaxes the surrounding cerebral arteries, increasing blood flow to active areas of the brain. Indeed, vessels such as the rat middle cerebral artery exhibit vasodilator responses to small increases in extracellular K⁺; K⁺ can also cause metabolic dilatation of blood vessels in the periphery. K⁺ has also been implicated in endothelium-derived hyperpolarizing factor (EDHF)-mediated vasodilatation in peripheral vessels (Edwards et al., 1998). Therefore, we assessed the possibility that K⁺ could contribute to EDHF-type relaxations in the rat middle cerebral artery.

Male Wistar rats (200-300g) were killed by cervical dislocation and exsanguination. The brain was removed and placed immediately in ice-cold Krebs solution. Segments of the middle cerebral artery (length, ~2mm; diameter, ~150 µM) were mounted in a Mulveny-Halpern myograph containing Krebs solution at 37ºC, gassed with 95% O₂ and 5% CO₂. EDHF mediated relaxations were elicited by the protease-activated receptor 2 agonist, SLIGRL (20 nM–20 µM) in the presence of L-NAME (100 µM). The effects of range of K_Ca channel inhibitors: the SK_Ca inhibitor, apamin (50 nM); the B/IK Ca inhibitor, charybdotoxin (100 nM); the IK_Ca inhibitor, TRAM-34 (1 µM) and the BK_Ca inhibitor, iberiotoxin (100 nM) were assessed on K⁺ and SLIGRL-induced relaxations. The effects of the K_IR inhibitor, barium (30 µM) and the Na/K ATPase inhibitor, ouabain (1 µM), were also assessed on K⁺ and SLIGRL induced relaxation. Data are mean ± s.e.mean of 4 or more animals. Statistical comparisons were made using one way ANOVA with Bonferroni’s post test.

EDHF responses elicited by addition of SLIGRL (20 nM- 20 µM, R_max 78.1 ± 12.9, n=7) in the presence of L-NAME were unaffected by apamin or iberiotoxin, alone or in combination (Figure 1), but were almost completely inhibited by either charybdotoxin (100 nM, R_max -5.4 ± 5.2 %, n=3, Figure 2) or TRAM-34 (1 mM, R_max 10.0 ± 6.4% n=4 Figure 3), each alone. Furthermore, Ba²⁺ (30 µM, R_max 57.6 ±17.8, n=4) and ouabain (1 mM, R_max, 55.2 ± 2.8 n=5) inhibited EDHF relaxation and when combined relaxation was further attenuated (R_max 29.1 ± 5.4, n=5, Figure 4). Relaxations to raising extracellular [K⁺] (7.8-19.8 mM, R_max 91.6 ± 10.0 at 13.8 mM, n=4) while unaffected by apamin, iberotoxin, charybdotoxin and TRAM-34, were inhibited by either barium or ouabain (R_max 41.6 ± 8.5, and 41.8 ± 13.3, respectively, n=4) and in combination they almost abolished K⁺ relaxation (R_max 25.7 ± 3.7, n=4, Figure 4).

The Blockade of EDHF mediated relaxation by either charybdotoxin or TRAM-34, suggests that IK_Ca channel opening is a critical step in the EDHF response in the rat middle cerebral artery and is in agreement with a previous study using this vessel (Marrelli et al., 2003). Furthermore, the inhibition of relaxation induced by EDHF and exogenous K⁺ by barium and ouabain indicates that K⁺ may function as an EDHF in rat mid cerebral arteries.

Edwards et al. (1998) Nature, 396, 269-272
Marrelli et al. (2003) Am. J. Physiol. 285, H1590-H1599