Outflow and overflow of picogram levels of endogenous dopamine and DOPAC from rat striatal slices: improved methodology for studies of stimulus-evoked release and metabolism.

An improved method for the rapid and sensitive determination of endogenous dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) in superfusates of single rat striatal slices, using high-performance liquid chromatography (HPLC) coupled with 'coulometric' electrochemical detection (EC), is described. Superfusates are directly injected into an HPLC-EC system following addition of a small aliquot of solubilized ascorbate-oxidase. DA and DOPAC are both separated and quantitated in 3-5 min. Twelve to 15 samples can be analyzed each hour with a nominal detection limit of 1.0 pg per injection for each compound or 10-20 pg/ml of superfusate. The present method was used to study changes in DA and DOPAC outflow and overflow in superfusates of single striatal slices following electrical field stimulation, both in the absence and presence of the catecholamine uptake inhibitor nomifensine. Studies of 1 min superfusate collections clearly showed that electrical field stimulation produced a latent increase in DOPAC as compared to DA. The routine sensitivity and sample throughout of the method allows for studies of both outflow and overflow of DA and DOPAC, as well as studies involving time-dependent overflow of these compounds.