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Ethnopharmacological relevance Gelsemium sempervirens L. is a traditional medicinal plant mainly distributed in the southeastern of the United States, employed in phytotheraphy and homeopathy as nervous system relaxant to treat various types of anxiety, pain, headache and other ailments. Although animal models showed its effectiveness, the mechanisms by which it might operate on the nervous system are largely unknown. This study investigated for the first time by a real-time PCR technique (RT-PCR Array) the gene expression of a panel of human neurotransmitter receptors and regulators, involved in neuronal excitatory signaling, on a neurocyte cell line. Materials and methods Human SH-SY5Y neuroblastoma cells were exposed for 24 h to Gelsemium sempervirens at 2c and 9c dilutions (i.e. 2 and 9-fold centesimal dilutions from mother tincture) and the gene expression profile compared to that of cells treated with control vehicle solutions. Results Exposure to the Gelsemium sempervirens 2c dilution, containing a nanomolar concentration of active principle gelsemine, induced a down-regulation of most genes of this array. In particular, the treated cells showed a statistically significant decrease of the prokineticin receptor 2, whose ligand is a neuropeptide involved in nociception, anxiety and depression-like behavior. Conclusions Overall, the results indicate a negative modulation trend in neuronal excitatory signaling, which can suggest new working hypotheses on the anxiolytic and analgesic action of this plant.