Explants of rat adrenal medulla were cultured in defined medium for up to 22 hr. Addition of dopamine to the medium led to a diminution in the activity of tyrosine hydroxylase (tyrosine 3-monooxygenase; EC 1.14.16.2) in the tissue. The enzyme activity was inversely proportional to the concentration of dopamine in the culture medium. The extent of loss of tyrosine hydroxylase, as measured by immunochemical titration, corresponded to the degree of loss in enzyme activity under the same conditions. The decreased amount of enzyme protein was due to a decrease in the rate of synthesis of tyrosine hydroxylase. However, this effect was not specific in that the relative rate of tyrosine hydroxylase synthesis was not decreased. Metabolites of dopamine when added to the medium did not affect tyrosine hydroxylase activity. Two other adrenal medullary enzymes, monoamine oxidase (EC 1.4.3.4) and acid phosphatase (EC 3.1.3.2), were not affected by addition of dopamine to the medium. The results indicate that elevated cytoplasmic levels of dopamine decrease the concentration of tyrosine hydroxylase by inhibiting protein synthesis.