Determination of release of endogenous dopamine from superfused substantia nigra slices

Abstract

Dopamine (DA) is synthesized and released not only from the terminals of the nigrostriatal dopaminergic neuronal pathway but also from cell bodies and dendrites in the substantia nigra (SN). In most of the in vitro studies on DA release in the SN, release of exogenously applied 3H-DA has been determined either from slices or from synaptosomes. However 3H-DA has some disadvantages; 3H-DA is taken up and released not only from dopaminergic cells but also from other neuronal elements and radiolabelled DA may not be evenly distributed with the releasable endogenous pool of the amine. Therefore we have developed a method for determination of the release of endogenous DA from superfused guinea pig SN slices. We have used a superfusion system in which 6 slices are placed into a microchamber. Samples of superfusate were collected every 10 min for up to 3 h and biochemical analyses were performed by electrochemical detection preceded by adsorbtion of DA on alumina and chromatography on a cation exchange column. Expected increases of the release of endogenous DA were obtained following d-amphetamine and potassium administration. The data indicate that it is possible to measure endogenous release of DA from guinea pig SN slices with standard HPLC technique and follow the release for a relatively long time.