Errors introduced by a tritium label in position 8 of catecholamines

The neuronal and extraneuronal disposition of³H-7,8-and³H-7-labelled (-)-noradrenaline and dopamine was compared in in vitro studies. 1. In agreement with earlier studies, the present results show that the presence of a tritium label in position 8 (i.e., on the alpha-carbon) has two consequences: a) the rate of deamination declines and b) part of the deamination results in the formation of an unlabelled aldehyde plus tritium water; tritium water is recovered from the OMDA-fraction of the column chromatographic procedure of Graefe et al. (1973). 2. Whenever the deamination of a³H-catecholamine is reduced (by tritium in position 8), the intraneuronal³H-catecholamine concentration is increased. This increase, in turn, partly masks the decline in neuronal deamination (rat vas deferens). Irrespective of whether one dertermines the spontaneous efflux, the release of³H-noradrenaline by nerve stimulation or the release of³H-(-)-noradrenaline by the reserpine-like compound Ro 4-1284, the presence of tritium in position 8 distorts the results (experiments with rat vasa deferentia and/or rabbit aorta). 3. In the extraneuronal system of the rat heart, two intracellular enzymes inactivate³H-(-)-noradrenaline and³H-dopamine: catechol-O-methyl transferase (COMT) and monoamine oxidase (MAO). Any hindrance of deamination (by tritium in position 8, COMT intact) leads to a shift of the metabolism of the³H-catecholamines from the exclusively deaminated to the exclusively O-methylated metabolites. 4. No differences between³H-7,8-and³H-7-labelled catecholamines were found after inhibition of MAO and COMT (extraneuronal accumulation and rate constant for efflux from the extraneuronal compartment III of the rat heart). 5. These results indicate that the presence of tritium in position 8 of catecholamines introduces errors, if monoamine oxidase is active. This is important for the interpretation of earlier results, since virtually all samples of "³H-(-)-noradrenaline" and "³H-dopamine nominally labelled in position 7" were contaminated with varying amounts of tritium in position 8. 6. In some experiments, also³H-7-(-)-adrenaline was used. For adrenergic nerve endings, the rate of metabolism (deamination) declined in the order:³H-7-dopamine > ³H-7-(-)-noradrenaline > ³H-7-(-)-adrenaline. For the extraneuronal disposition the ranking order was:³H-7-(-)-adrenaline > ³H-7-(-)-noradrenaline=³H-7-dopamine. However, in the extraneuronal disposition of³H-(-)-adrenaline, O-methylation predominated over deamination.