Flow may modulate the ability of ascorbate to block EDHF-mediated vasodilatation in bovine long posterior ciliary artery Fiona J. Dowell1, Silvia Nelli2, Alison Stirrat2, William S. Wilson2& William Martin2. 1Institute of Comparative Medicine, University of Glasgow Vet School, G61 1QH & 2Institute of Biomedical and Life Sciences, University of Glasgow, G12 8QQ.

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Ascorbate blocks endothelium-derived hyperpolarizing factor (EDHF)-mediated vasodilatation in the bovine perfused ciliary circulation and rat perfused mesentery but not in bovine coronary artery rings in static tissue baths (McNeish et al., 2002, 2003). The aim of the present study was to compare the ability of ascorbate to block EDHF-mediated vasodilatation in a single vessel type when perfused and in a tissue bath.

A long posterior ciliary artery (LPCA) was dissected from bovine eyes obtained at a local abattoir and either cannulated and perfused at 2.5 ml min^-1 with Krebs solution at 37°C gassed with 95% O₂ / 5% CO₂ or cut into 2 mm rings and mounted in a wire myograph (set at transmural pressure equivalent to 70-100 mmHg). The thromboxane A₂-mimetic, U46619 (0.01-1 µM) was used to induce constriction and vasodilator responses were elicited using acetylcholine (ACh) or bradykinin (BK). The combination of 100 µM L-NAME and 10 µM indomethacin (L-N/Indo) was used to block nitric oxide synthase and cyclo-oxygenase, whereas the combination of apamin and charybdotoxin (ChTx/Ap; both 100 nM) was used to block EDHF. All blocking drugs were present for at least 20 min except ascorbate, which was included for 120 min. Data are expressed as mean ± s.e. mean, n5, with differences determined by ANOVA (Bonferroni post-test).

In the perfused LPCA, bolus doses of BK (0.1 pmol-10 nmol)induced dose-dependent vasodilatation (max. 69 ± 7%), which was reduced to 34 ± 4% (p<0.001) by L-N/Indo, to 27 ± 11% (p<0.001) by ChTx/Ap and to 7 ± 5% (p<0.001) by all 4 blockers. ACh (0.1-1000 nmol) also induced dose-dependent vasodilatation (max. 44 ± 8%), which was reversed to vasoconstriction of 55 ± 10% by L-N/Indo, of 34 ± 21% by ChTx/Ap and of 20 ± 14% (p<0.001 for each) by all 4 blockers. Ascorbate (50 µM) also blocked vasodilatation to BK (max. 35 ± 6%, p<0.001) and ACh (reversed to constriction of 50 ± 10%, p<0.001). In rings of LPCA, BK (1-1000 nM) induced concentration-dependent vasodilatation (max. 88 ± 5%). This was reduced to 72 ± 5% by L-N/Indo, to 51 ± 11% (p<0.01) by ChTx/Ap and to 14 ± 4% (p<0.001) by all 4 blockers. ACh (0.01-10 mM) also induced concentration-dependent vasodilatation (max. 86 ± 2%). This was reduced to 33 ± 9% (p<0.001) by L-N/Indo, to 21 ± 6% (p<0.001) by ChTx/Ap and to 4 ± 1% (p<0.001) by all 4 blockers. In stark contrast to our findings in perfused preparations, ascorbate (150 µM) had no effect on vasodilatation to either BK (92 ± 3%) or ACh (81 ± 2%) in rings of LPCA.

These results show that ascorbate blocks EDHF-mediated vasodilatation induced by BK and ACh in the bovine ciliary artery when it is perfused but not when it is suspended in a static wire myograph system. The possibility that flow is required to uncover the ability of ascorbate to block EDHF warrants further investigation.

McNeish et al. (2002) Br. J. Pharmacol. 135, 1801-1809.
McNeish et al. (2003) Br. J. Pharmacol. 138, 1172-1180.

Supported by the BHF and Wellcome Trust.