Catechol-O-methyltransferase activity in rat brain primary neuronal and glial cell cultures and its inhibition by novel drugs

To analyze the intracellular catechol-O-methyltransferase (COMT) activity i n cultured cells, a substrate (dihydroxybenzoic acid) was added to the tiss ue culture plates and the product (vanillic acid) was analyzed with reverse d-phase HPLC with coulometric detection. The cultures, prepared from variou s regions of the brain, were immunohistochemically characterized. Basal COM T activities were similar in all glial cultures while in some neuron-enrich ed cultures the enzyme activity was lower. Preincubation with nitrocatechol COMT inhibitors, entacapone or tolcapone (30-300 nM), decreased COMT activ ity in all culture types with slightly different potencies. An atypical inh ibitor of O-methylation, CGP 28104, had no effect on COMT activity in any o f these cultures. In conclusion: 1) this method enables straight-forward an alysis of a large number of COMT activity samples in cultured cells, 2) COM T activity is almost equally distributed throughout the brain, and 3) tolca pone was a more potent COMT inhibitor than entacapone in cultures containin g glial cells.