SUBCLASSIFICATION OF PRESYNAPTIC ALPHA(2)-ADRENOCEPTORS - ALPHA(2D)-AUTORECEPTORS AND ALPHA(2D)-ADRENOCEPTORS MODULATING RELEASE OF ACETYLCHOLINE IN GUINEA-PIG ILEUM

The study was designed to classify in terms of alpha(2A), alpha(2B), a lpha(2C) and alpha(2D) the presynaptic alpha(2)-autoreceptors, as well as the alpha(2)-receptors modulating the release of acetylcholine, in the myenteric plexus-longitudinal muscle (MPLM) preparation of the gu inea-pig ileum. A set of antagonists was chosen that was able to discr iminate between the four subtypes. Small pieces of the MPLM preparatio n were preincubated with H-3-noradrenaline or H-3-choline and then sup erfused and stimulated electrically. The stimulation periods used (H-3 -noradrenaline: 3 trains of 20 pulses, 50 Hz, train interval 60 s; H-3 -choline: single trains of 30 pulses, 0.2 Hz) did not lead to alpha(2) -autoinhibition or inhibition of H-3-acetylcholine release by endogeno us noradrenaline. The alpha(2)-selective agonist 5-bromo-6-(2-imidazol in-2-ylamino)-quinoxaline (UK 14,304) reduced the evoked overflow of t ritium in both H-3-noradrenaline and H-3-choline experiments. Most (H- 3-noradrenaline) or all (H-3-choline) of the 10 antagonists shifted th e concentration-inhibition curves of UK 14,304 to the right. pK(d) val ues of the antagonists were calculated from the shifts. pK(d) values f rom H-3-noradrenaline experiments correlated with pK(d) values from H- 3-choline experiments (r = 0.981). It is concluded that alpha(2)-autor eceptors and alpha(2)-heteroreceptors modulating the release of acetyl choline in the MPLM preparation are of the same subtype. Comparison wi th antagonist affinities for prototypic native alpha(2) binding sites, binding sites in cells transfected with alpha(2) subtype genes, and p reviously classified presynaptic alpha(2)-adrenoceptors - all taken fr om the literature - indicates that both are alpha(2D). The results are consonant with the hypothesis that at least the majority of alpha(2)- autoreceptors belong to the alpha(2A/D) branch of the alpha(2)-adrenoc eptor tree, across mammalian or at least across rodent and lagomorph s pecies. The same may hold true for alpha(2)-adrenoceptors on non-norad renergic neurones.