Serotonin stimulation of alveolar macrophages from wild type and 5-HT2C deficient animals

Gene regulation upon serotonin stimulation was compared in wild-type and 5HT2C receptor kock-out mice. Serotonin (5-HT), known as neuromodulator, regulates immune responses and inflammatory cascades. The expression and function of 5-HT receptors on alveolar macrophages (AM), which are the major fraction of pulmonary immune cells, remain elusive. Therefore we determined the expression of 5-HT type 2 receptors and investigated the effects evoked by stimulation with 5-HT in AM compared to alveolar epithelial cells (AEC). Quantitative PCR (qPCR) analysis revealed expression of the receptors 5-HT2A and 5-HT2B in AEC and of 5-HT2C in AM. In AM, 5-HT (10-5 M) induced a rise in intracellular calcium concentration ([Ca2+]i) that was initiated by release of Ca2+ from intracellular stores and depended on extracellular Ca2+ in a sustained phase. This 5-HT-induced increase in [Ca2+]i was not observed in AM treated with the 5-HT2C-selective inhibitor RS102221 and in AM derived from 5-HT2C-deficient mice. AM stimulated with 5-HT (10-5 M) showed increased expression of CCL2 (MCP-1) mRNA as assayed by qPCR at 4 h and augmented production of CCL2 protein as determined by dot-blot assay and ELISA at 24 h. Notably, in 5HT2C-deficient AM, CCL2 production was not affected by 5-HT treatment. Moreover, transcriptional responses to 5-HT exposure assayed by microarray experiments were only observed in AM from wild type animals but not in AM derived from 5-HT2C-deficient mice. Taken together these data demonstrate the presence of functional 5-HT2C receptors on AM and suggest a role of 5-HT as novel modulator of AM function. These effects are exclusively driven by the 5-HT2C receptor, thereby providing the potential for selective intervention. primary isolated murine alveolar macrophages were stimulated with serotonin