Differential role of KCa in dilatation evoked by luminally-perfused purinoceptor agonists in rat small mesenteric arteries

Polly Winter & Kim A Dora, Department of Pharmacy & Pharmacology, University of Bath, Bath BA1 7AY.

It has been demonstrated that ATP can facilitate flow-induced dilatation in rat mesenteric arteries (Liu et al., 2004) . The aim of this investigation was to assess the EDHF component of the ATP-facilitated flow-induced dilatation response, and whether P2Y receptors played a role.

Male Wistar rats (200-250g) were killed by schedule 1 methods of the Animals Scientific Procedures Act 1986 ( UK). Segments of third order mesenteric vessels were cannulated, mounted in a pressure myograph (Danish Myotechnology), pressurized to 50mmHg and perfused with MOPS at 37 oC. Vessels were pre-contracted with phenylephrine to give an internal diameter of ~50-80 μm. A multi-channel syringe pump was used to perfuse agonists through the lumen at 90μl.min -1.

Luminal perfusion of vehicle alone had no effect on arterial diameter (n=5). Perfusion of ATP (10nM-3μM) stimulated concentration- and flow-dependent dilatation (pD2; 6.4±0.1; Rmax: 94.9±3.3%, n=11). The flow-dependent dilatation to ATP was unaffected by the NO synthase (NOS) inhibitor L-NAME (100μM). In the presence of L-NAME, selective inhibition of intermediate-conductance Ca2+-activated K+ channels (IKCa) with TRAM-34 (1μM) had no effect against the dilatation to 1μM ATP (TRAM-34; 72.5±17.1%, n=5 vs control; 78.7±7.1%), while selective inhibition of small-conductance KCa (SKCa) with apamin (50nM) greatly reduced the dilatation to 1μM ATP (27.1±13%, n=4) to a level not significantly different from that seen following combined inhibition of SKCa and IKCa (15.5±1.2%, n=4).

Perfusion of the non-hydrolyzable ADP analogue ADP b S (1 m M) stimulated flow-dependent dilatation (59.1±5.2%, n=8) that was fully blocked only after inhibition of NOS, SKCa and IKCa (3.3±1.5%, n=4). In contrast, luminal perfusion of UTP (3μM) stimulated flow-dependent dilatation (90.1±5.0%, n=5) that was not completely blocked by inhibition of NOS, SKCa and IKCa (31.4±11.4%, n=4). Raised extracellular K+ (35-40mM) had no further inhibitory effect on the UTP-facilitated, flow-dependent response (53.0±11.4%, n=4).

Endothelial cell KCa have been found to differentially contribute to the ATP-, ADPβS- and UTP-stimulated flow-dependent dilatation. This indicates a differential activation of purinergic receptor subtypes by the three agonists and suggests that a variation in the subsequent downstream effector pathways may also exist.

Liu, C et al. 2004. Am. J. Physiol. Heart Circ. Physiol. 286: H1688-H1695

This work was supported by the British Heart Foundation.