Several studies have suggested that 5-HT7 receptors are involved in nociceptive processing but the exact contribution of peripheral versus central 5-HT7 receptors still needs to be elucidated. In the present study, the respective roles of peripheral and spinal 5-HT7 receptors in the modulation of mechanical hypersensitivity were investigated under two different experimental pain conditions. In a first set of experiments, the selective 5-HT7 receptor agonist, E-57431, was systemically, intrathecally or peripherally (intraplantarly) administered to rats sensitized by intraplantar injection of capsaicin. Oral administration of E-57431 (1.25–10 mg/kg) was found to exert a clear-cut dose-dependent reduction of capsaicin-induced mechanical hypersensitivity. Interestingly, intrathecal administration of E-57431 (100 μg) also inhibited mechanical hypersensitivity secondary to capsaicin injection. In contrast, a dose-dependent enhancement of capsaicin-induced mechanical hypersensitivity was observed after local intraplantar injection of E-57431 (0.01–1 μg). In a second set of experiments, E-57431 was systemically or intrathecally administered to rats submitted to neuropathic pain (spared nerve injury model). Significant inhibition of nerve injury-induced mechanical hypersensitivity was found after intraperitoneal (10 mg/kg) as well as intrathecal (100 μg) administration of E-57431 in this chronic pain model. These studies provide evidence that, under sensitizing neurogenic/neuropathic conditions, activation of 5-HT7 receptors exerts antinociceptive effects at the level of the spinal cord and pronociceptive effects at the periphery. The antinociceptive effect mediated by central 5-HT7 receptors seems to predominate over the pronociceptive effect at the periphery when a selective 5-HT7 receptor agonist is systemically administered.