Affinities of catecholamines for extraneuronal uptake in smooth muscle cells in trachealis muscle of guinea-pig

Summary

• 1. Tracheal segments from guinea-pigs pretreated with 6-hydroxydopamine were incubated with adrenaline, noradrenaline or isoprenaline at 37° C for various time intervals. Catechol-O-methyl transferase was inhibited by 100 μM U-0521. Tissues were prepared for fluorescence histochemistry and accumulated catecholamine in trachealis smooth muscle cells measured by fluorescence microphotometry.

• 2. The monoamine oxidase inhibitor, pargyline, markedly enhanced the accumulation of noradrenaline, but did not affect accumulation of isoprenaline. Pargyline (100 μM) was included in all further experiments with adrenaline or noradrenaline, but not in those with isoprenaline.

• 3. Net uptake into the smooth muscle cells approached steady state after incubation with amine for 60–120 min. The uptake of amine was linearly related to time for up to 10 min with 5 μM adrenaline, noradrenaline and isoprenaline and with 800 μM noradrenaline, but was linear only up to 5 min for 800 μM adrenaline and isoprenaline. Thus, an incubation time with amine of 5 min was selected for the initial rate kinetic study.

• 4. The uptake of the three catecholamines into the smooth muscle cells obeyed Michaelis-Menten saturation kinetics. The mean K _m values for adrenaline, noradrenaline and isoprenaline were 151 μM, 238 μM and 273 μM respectively. Hence, the affinities of the three catecholamines for extraneuronal uptake in the trachealis smooth muscle cells are of the same order of magnitude. Adrenaline may have a slightly higher affinity than the other two amines. This is contrary to the accepted generalization that isoprenaline has a higher affinity for extraneuronal uptake than adrenaline or noradrenaline.