Dopamine turnover estimated by simultaneous LCEC assay of dopamine and dopamine metabolites

The estimation of changes in tissue levels of dopamine (DA) and its metabolites has been employed to approximate the turnover rate or release of DA. We have developed an assay using high pressure liquid chromatography with electrochemical detection to quantify DA, dihydroxyphenylacetic acid (DOPAC), 3-methoxytyramine (3-MT), and homovanillic acid (HVA) simultaneously in brain homogenates. The assay is simple, inexpensive, rapid and accurate; the sensitivity of the assay for DA and DOPAC was 10 pg, for HVA 50 pg, and 200 pg for 3-MT. Apomorphine treatment resulted in a reduction in tissue levels of DOPAC and HVA, while haloperidol administration increased tissue levels of DOPAC and HVA. Neither drug treatment caused changes in DA levels. Administration of d-amphetamine 2.5 mg/kg S.C. resulted 1 hour later in increases in the striatal concentration of DA (57%) and 3-MT (44%) and decreases in DOPAC (40%) and HVA (39%). These data are consistent with the agonistic effect of apomorphine and antagonistic effect of haloperidol on DA autoreceptors and post-synaptic receptors. In the case of amphetamine, the increases in the levels of DA and 3-MT probably reflect the ability of d-amphetamine to induce DA release and block re-uptake. The reduction of DOPAC and HVA levels are consistent with the ability of d-amphetamine to competitively inhibit monoamine oxidase activity as well as the neuronal reuptake of DA. The simultaneous estimation of DA, DOPAC, 3-MT, and HVA should facilitate the understanding of dopaminergic drugs with complicated mechanisms of action.