The extraneuronal O-methylation of 3H-(±)isoprenaline by guinea-pig tracheal rings in vitro

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Abstract
Summary Tracheal rings isolated from male guinea-pigs and incubated in Krebs' solution at 37° C O-methylated 3H-(±)isoprenaline by a saturable, high affinity mechanism. 1. With 3H-isoprenaline at 1 μmol·l−1, O-methyl 3H-(±)isoprenaline (3H-OMI) appeared in the tissue with a half-time for approach to steady state of approximately 10 min and was measured in the incubation medium after about 5 min, its concentration increasing linearly thereafter. With 3H-isoprenaline concentrations ranging from 1 to 200 μmol·l−1, the total formation of 3H-OMI (estimated from that contained in the tissue and the medium) was maintained at steady state rates for up to 60 min, after initial lag times of between 1 and 3 min. O-methylation obeyed Michaelis-Menten saturation kinetics; K m=6.14±0.13 μmol·l−1, V max=0.31±0.01 nmol·g−1·min−1. 2. The catechol-O-methyl transferase (COMT) inhibitor U-O521 and the extraneuronal uptake inhibitor corticosterone both reduced O-methylation of 3H-isoprenaline (0.1 μmol·l−1) by tracheal rings. However, U-O521 was fully inhibitory (IC50=2.6 μmol·l−1), but corticosterone inhibited by only 46% at concentrations up to 1 mmol·l−1. 3. The O-methylating activity of the “smooth muscle-rich” component of the trachea was approximately three-times greater than for complete tracheal rings. However, considerable activity was also associated with “cartilage-rich”, “smooth muscle-poor” sections. This activity did not seem to be associated with endothelial cells. 4. Histamine strongly inhibited O-methylation (IC50=30 μmol·l−1), but two other contractile agonists, 5-hydroxytryptamine and bethanechol, were weakly active and inactive, respectively.