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Notes: Abstract. The aim of the present study was to characterize putative muscarine receptors on sympathetic nerve terminals in the rabbit trachea. Release of endogenous noradrenaline from in vitro incubated rabbit tracheae was evoked by electrical field stimulation (3 Hz, 540 pulses) and quantified by high performance liquid chromatography with electrochemical detection.   The muscarine receptor agonist oxotremorine inhibited the evoked release of noradrenaline completely at 1 μmol/l (EC50: 64 nmol/l). The concentration response curve was very steep (Hill coefficient of 2.3). Scopolamine shifted the concentration response curve of oxotremorine to the right (–log KB 8.48) demonstrating specific, inhibitory muscarine receptors. Several subtype-preferring muscarine receptor antagonists also shifted the concentration response curve of oxotremorine to the right. The rank order of potency was (–log KB or pA2*): scopolamine (8.48)〉AF-DX 384 (7.88*; slope of Schild plot 1.1)〉(R)-trihexyphenidyl (7.87)〉4-DAMP (7.85) 〉AQ-RA 741 (7.77) 〉 methoctramine 6.18) 〉 pirenzepine (6.0)〉p-fluoro-hexahydrosiladifenidol (p-FHHSiD, 5.68). When these affinity constants were plotted against reported –log Ki values determined in binding studies on human cloned muscarine receptor subtypes (m1–m5), the best correlation was obtained for m2. Indomethacin (3 μmol/l), which on its own increased the evoked noradrenaline release by about 45%, affected neither the inhibitory effect of oxotremorine nor the antagonistic potency of methoctramine or p-FHHSiD. After preincubation for 48 min with 300 μmol/l phenoxybenzamine, which has been shown to inactivate muscarine receptors irreversibly, the concentration response curve of oxotremorine was shifted 5.2fold to the right and the maximal inhibition was reduced by 50%, whereas the slope remained steep (Hill coefficient 2.6). These experiments indicated that a fraction of about 22% of the muscarine receptors has to be occupied by oxotremorine to produce half-maximum inhibition of noradrenaline release; the dissociation constant of oxotremorine at the prejunctional muscarine receptors was 0.33 μmol/l.   In conclusion, the sympathetic nerve terminals in the rabbit trachea are endowed with inhibitory M2-like muscarine receptors for which methoctramine displayed a low affinity. Since a large receptor reserve could be excluded, the steep concentration response curve of oxotremorine suggests that activation of muscarine receptors has to reach a threshold level before the onset of an inhibitory effect.

Notes: Abstract. Overflow of endogenous noradrenaline from rat isolated tracheae was evoked by electrical field stimulation (3 Hz, 540 pulses) in the presence of yohimbine, desipramine and tyrosine. Isoprenaline 100 nmol/l increased the evoked overflow of noradrenaline by about 65%. This effect was antagonized by propranolol (100 nmol/l) and the β 2-selective adrenoceptor antagonist ICI 118,551 (100 nmol/l), but not by the β 1-selective adrenoceptor antagonist CGP 20712 A (100 nmol/l). The β 2-selective adrenoceptor agonist formoterol (1–100 nmol/l) also facilitated the evoked overflow of noradrenaline, but maximally by only about 25% at 10 nmol/l, i.e. formoterol behaved as a partial agonist at these facilitatory β-adrenoceptor. This assumption is also supported by the observation that formoterol (10 nmol/l) acted as antagonist against isoprenaline (100 nmol/l). Mechanical removal of the mucosa resulted in a 30% decrease in tissue noradrenaline and a 55% reduction of the evoked overflow of noradrenaline. In mucosa-denuded preparations isoprenaline failed to facilitate noradrenaline overflow. In the presence of indomethacin (3 μmol/l) the evoked overflow of noradrenaline from mucosa containing preparations was increased by about 50%, but isoprenaline still further facilitated the evoked noradrenaline overflow by about 40%. In conclusion, the overflow of noradrenaline in the rat trachea is facilitated via β 2-adrenoceptors, an effect which requires an intact airway mucosa.