Regulation of the electrically evoked release of [3H]5-HT was examined in midbrain, frontal cortex and hippocampus preloaded slices obtained from wild-type and 5-HT1B knock-out mice. In the absence of any drug [3H]5-HT released was increased in midbrain and hippocampus but not in frontal cortex slices of [3H]5-HT1B knock out mice. The selective 5-HT1B agonist CP 93129 and the 5-HT1B/1D agonist sumatriptan, inhibited [3H]5-HT release in hippocampus and cortical slices obtained from control mice but had no effect in mutants. In the two projection areas studied, the non-selective 5-HT agonist 5-carboxyamidotryptamine (5-CT) inhibited [3H]5-HT release in both groups of mice, indicating that additional 5-HT receptors, other than 5-HT1B, might be involved in the regulation of [3H]5-HT release from 5-HT terminals. In slices containing midbrain raphe nuclei, CP 93129 had no effect in either group. In contrast, sumatriptan inhibited [3H]5-HT release in controls and mutants. The latter effect was blocked by the 5-HT1D antagonist GR 127935, but not the 5-HT1A antagonist (+)WAY 100135, thus suggesting that a 5-HT1D-like receptor, possibly 5-HT1D alpha, negatively regulates 5-HT release in mouse midbrain raphe nuclei.