Catecholamine synthesis from [3H]tyrosine was studied in slices of striatum, cerebellum and substantia nigra of mice. If low concentrations of tyrosine (less than 5.5 microM) were added to the incubation medium, the slices released significant amounts of tyrosine into the medium during the incubation. Kinetic analysis of the same experiments indicated that medium tyrosine and not tissue tyrosine was the appropriate precursor for both dopamine (DA) synthesis and protein synthesis in striatal slice. Concentrations of medium tyrosine of 8.25 microM or greater were sufficient to prevent changes of medium tyrosine during incubation and thus maintained a constant specific activity of precursor. Increasing concentrations of medium K+ increased both the accumulation of [3H]DA and its release from striatal slices. However, accumulation was stimulated at a concentration of K+ (14 mM) that had no significant stimulatory effect on release, suggesting that the stimulatory effects of K+ on synthesis and release are mediated by separate processes. Release of 14CO2 from [1-14C]tyrosine closely paralleled the accumulation of [3H]DA from [3H]tyrosine. Release of preloaded [14C]DA closely paralleled that of [3H]DA synthesized from [3H]tyrosine, suggesting a common functional pool. The principal DA catabolite produced was dihydroxyphenylacetic acid (DOPAC). The appearance of labeled DOPAC in the media was greatly enhanced by K+ stimulation.