Spatial heterogeneity in the mechanisms contributing to acetylcholine-induced dilatation in the rabbit isolated ear

1. Using an X-ray microangiographic technique in rabbit isolated perfused ears preconstricted with 5-HT (300 nM) and histamine (300 nM), we investigated the combined actions of N^ω-nitro-L-arginine methyl ester (L-NAME) and indomethacin on acetylcholine-induced depressor responses.
2. Under control conditions, acetylcholine (10 nM–30 μM) induced a concentration-dependent reversal of the pressor response, reaching a maximum of 66.0±13.6% (n=6). In the presence of L-NAME (300 μM) and indomethacin (10 μM), this depressor action was reduced, reaching a maximum of 38.6±5.9% (n=6).
3. The control response was associated with substantial vasodilatation in the central ear artery (G₀), a smaller dilatory action on first generation branch arteries (G₁) and no effect on second generation branch arteries (G₂). In the presence of L-NAME and indomethacin, vasodilatation occurred in G₂ with no effect in G₀ or G₁.
4. Two calcium-activated K⁺ channels blockers, charybdotoxin (ChTX; 10 nM) and penitrem A (100 nM), further inhibited, but did not abolish, the L-NAME- and indomethacin-resistant response to acetylcholine (10 nM–300 μM). Both agents abolished the vasodilatory action of acetylcholine in G₂.
5. In conclusion, L-NAME and indomethacin induced a shift in acetylcholine-induced vasodilatation from G₀ and G₁ to G₂. This is consistent with the suggestion that nitric oxide dominates in larger vessels whilst other mechanisms dominate in smaller vessels. The L-NAME- and indomethacin-resistant component was inhibited by ChTX and penitrem A, suggesting it is mediated, at least in part, by activation of K_Ca channels and could therefore involve a hyperpolarising mediator such as endothelium-derived hyperpolarising factor.