Dopamine autoreceptor regulation of the kinetic state of striatal tyrosine hydroxylase.

Tyrosine hydroxylase isolated from striatal synaptosomes exhibits biphasic Lineweaver-Burk kinetics for its tetrahydrobiopterin cofactor, consistent with multiple Km forms of the enzyme. Incubation of striatal synaptosomes with forskolin (EC50 0.45 microM) or dibutyryl cyclic AMP (EC50 1.2 mM), results in activation of tyrosine hydroxylase, isolated from these synaptosomes via conversion of the enzyme to a single low Km form (Km 40 microM). The activation of synaptosomal tyrosine hydroxylase by forskolin or dibutyryl cyclic AMP is not additive and is similar to activation seen with cyclic AMP-dependent protein kinase phosphorylation of purified tyrosine hydroxylase. The addition of dopamine (IC50 1.0 microM) (with nomifensine and pargyline) or apomorphine (IC50 30 nM) to the synaptosomal incubation medium blocks the activation of tyrosine hydroxylase by forskolin. This effect of dopamine and apomorphine can in turn be blocked by preincubation of the synaptosomes with the dopamine receptor antagonist haloperidol (IC50 30 nM and 4.5 nM, respectively) or chlorpromazine (IC50 50 nM versus apomorphine). In contrast to the forskolin data above, dopamine failed to block the activation of tyrosine hydroxylase by dibutyryl cyclic AMP. Addition of dopamine to the tyrosine hydroxylase assay, in amounts equivalent to that carried over from the synaptosomal incubation with the tyrosine hydroxylase, had no effect on forskolin-activated enzyme. The observations that dopamine and apomorphine can block forskolin activation of tyrosine hydroxylase, that this blockade can in turn be prevented by preincubation with haloperidol or chlorpromazine, and that the amount of dopamine required for blockade of forskolin activation in synaptosomes has no effect on tyrosine hydroxylase when added to the enzyme assay constitute the first clear evidence of a presynaptic dopamine receptor (autoreceptor). This autoreceptor regulates the activity of tyrosine hydroxylase by preventing or reversing cyclic AMP-dependent activation of the enzyme, probably through a decrease in the phosphorylation state of tyrosine hydroxylase. Failure of dopamine to block dibutyryl cyclic AMP activation of tyrosine hydroxylase suggests that, if forskolin and dibutyryl cyclic AMP activate tyrosine hydroxylase through identical changes in phosphorylation state, then autoreceptor regulation of tyrosine hydroxylase must occur through a decrease in cyclic AMP levels.