The outward transport of catecholamines mediated by uptake₂ of the rat heart

Summary

The efflux of³H-catecholamines from the extraneuronal tissue of the rat heart was analysed (after inhibition of vesicular and neuronal uptake, monoamine oxidase and catechol-O-methyl transferase). In most experiments, hearts were first loaded with a tracer concentration of a³H-catecholamine and then washed out.

• 1. For all four catecholamines [³H-(±)-isoprenaline,³H-(±)-adrenaline,³H-(−)-noradrenaline, and³H-dopamine] the loading period resulted in virtually the same distribution pattern: most of the radioactivity distributed into “compartment III”. However, the rate constants for efflux from compartment III increased in the order³H-(−)-noradrenaline <³H-dopamine<³H-(±)-isoprenaline=³H-(±)-adrenaline.

• 2. O-methyl-isoprenaline (OMI, a potent inhibitor of uptake₂) caused a concentration-dependent and partial inhibition of the efflux of all³H-catecholamines; its IC50 (half-maximal inhibition of OMI-sensitive efflux) was very close to that for half-maximal inhibition of inward transport by uptake₂. It is concluded that there is not only (OMI-resistant) diffusional efflux of³H-catecholamines, but also (OMI-sensitive) outward transport of³H-catecholamines. The contribution by each of these processes to total efflux differed considerably from one³H-catecholamine to the next.

• 3. U-0521 (the COMT inhibitor used in this study) inhibited the OMI-sensitive efflux of³H-noradrenaline with an IC50 of about 100 μmol/l. However, no inhibitory effect was found for 10 μmol/l U-0521.

• 4. During the wash-out period (see above) various unlabelled substrates of uptake₂ were added to the perfusion fluid at a concentration equalling 2×K _m. Dopamine, which has a very highV _max for uptake₂, caused a small acceleration of the efflux of³H-isoprenaline (facilitated exchange diffusion); 5-hydroxytryptamine, which has an intermediateV _max for uptake₂, had no effect on the efflux of³H-isoprenaline; clonidine, which has a lowV _max for uptake₂, inhibited the efflux of³H-isoprenaline.

• 5. For a total of nine unlabelled substrates (added to the perfusion fluid at 2×K _m) significant correlations were obtained between the degree of inhibition of the efflux of³H-(±)-isoprenaline, on the one hand, and the logarithm of either theV _max or theK _m for uptake₂, on the other hand. Such results might be explained by either the dissociation of the substrate from the carrier on the inside or the return of the empty carrier to the outside being the rate-limiting step in the transport cycle. This would account for the differences betweenV _max-values and for the inhibitory effect of substrates with lowV _max for uptake₂ on the efflux of³H-isoprenaline.