Inhibition of extraneuronal uptake
Springer Online Journal Archives 1860-2000
Summary To simultaneously determine the kinetics of removal, O-methylation and accumulation of 3H-isoprenaline, isolated rat hearts were perfused for 4 min with various concentrations of 3H-isoprenaline. The apparent K m for the O-methylation of 3H-isoprenaline (3.3±0.5 μM) was more than one order of magnitude lower than the corresponding value for the accumulation of unchanged amine (71.3±7.1 μM). The apparent K m for removal was very similar to that for accumulation (63.2±5.9 μM). At perfusion concentrations higher than 25 μM, i.e. when O-methylation was saturated, removal virtually equalled accumulation. However, at low substrate concentrations removal of 3H-isoprenaline was overwhelmingly followed by O-methylation; this led to a marked difference between rates of removal and those of accumulation. When initial rates of uptake of 3H-isoprenaline were determined after 1.5 min of perfusion of the hearts by the method of Graefe et al. (1978), the uptake of 3H-isoprenaline consisted of two components: a nonsaturable and a saturable (after subtraction of the nonsaturable component from the total uptake). The kinetic constants of the saturable component of uptake were higher than those obtained after 4 min perfusion (see above) (K m : 110±19 μM; V max: 80±4 nmoles·g−1·min−1). Corticosterone competitively inhibited the saturable component of uptake of 3H-isoprenaline (K m : 1.2 μM). During wash out of accumulated 3H-isoprenaline, O-methylation took place predominantly in one of the two extraneuronal compartments. The efflux of 3-O-methyl-3H-isoprenaline (3H-OMI), the O-methylated metabolite of 3H-isoprenaline, was characterized by a half time of about 1.2 min. O-methylation accelerated the loss of radioactivity from the tissue during wash out. The extraneuronal uptake of 3H-isoprenaline was characterized as a “pump and leak” system by means of steady-state kinetics of accumulation of 3H-isoprenaline. Half saturation of the steady-state accumulation was observed at a concentration of 104.5 ±18.5 μM 3H-isoprenaline; the leak component was characterized by a rate constant of 0.0359 min−1.
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