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The effects of RU24722 (14,15-dihydro-20,21-di-noreburnamine-14-ol) on tyrosine hydroxylase in central catecholaminergic neurons were studied in rats treated with different quantities of the molecule, and a time course was done for the minimal dose that gave the maximal effect. RU24722 induced increases in tyrosine hydroxylase activities and specific protein content in noradrenergic cells of the locus ceruleus and decreased all these parameters in dopaminergic neurons of the substantia nigra and ventral tegmental area. The results pointed out that the specific activity of newly synthesized tyrosine hydroxylase in the loci cerulei was potentially greater but was not expressed “in vivo” except 7 days after injection. The phenotypic specificity and the time course pattern of the action could be considered as a consequence of an induction mechanism. The comparison of long-term change in tyrosine hydroxylase values after pipe-roxane, RU24722, clonidine, and combined RU24722-clon-idine treatment demonstrated that an activation during a few hours did not induce tyrosine hydroxylase in central noradrenergic neurons. Clonidine antagonized the activating effect of RU24722 following its injection but did not affect its long-term induction properties.

An improved quantitative immunochemical determination of brain tyrosine hydroxylase (TH) concentrations was designed using direct transfer into nitrocellulose from 20-microns thick brain sections, followed by immunodetection and quantitative radioautography in three reference brain structures (locus ceruleus, substantia nigra, and ventral tegmental area). Results obtained by this methodology were similar to those obtained after extraction and Western blotting of the TH protein in control and reserpine-treated animals. Moreover, this methodology allows the combination of high sensitivity and high anatomical resolution in the study of the distribution of pharmacological effects. The locus ceruleus exhibited a significant posteroanterior distribution of TH protein concentration in control and reserpine-treated animals.

The subcellular distribution of the protein tyrosine hydroxylase (TH) after fractionation of rat brain tissue was studied by a sensitive technique of immunoblot quantification in the dopaminergic nigrostriatal and the dorsal noradrenergic pathways and in the ventrolateral medulla. This repartition indicates that in all catecholaminergic regions of the cell bodies studied, the contribution of the nerve endings to the total TH amount is very low (less than 7%), in contrast to that observed in the terminal fields. The correlative subcellular determination of the TH amount and activity in the same tissue could be a useful approach for studying experimentally induced mechanisms of catecholamine synthesis modulation in different brain catecholaminergic pathways.