Abstract

Serotonin 5-HT_2C receptors (5-HT_2CR) activate G_q proteins and are expressed in the central nervous system (CNS). 5-HT_2CR regulate emotion, feeding, reward, or cognition and may serve as promising drug targets to treat psychiatric disorders or obesity. Owing to technical difficulties in isolating cells from the CNS and the lack of suitable cell lines endogenously expressing 5-HT_2CR, our knowledge about this receptor subtype in native environments is rather limited. The hypothalamic mHypoA-2/10 cell line was recently established and resembles appetite-regulating hypothalamic neurons of the paraventricular nucleus (PVN), where 5-HT_2CR have been detected in vivo. Therefore, we tested mHypoA-2/10 cells for endogenous 5-HT_2CR expression. Serotonin or the 5-HT_2CR preferential agonist WAY-161,503 initiated cAMP response element (CRE)-dependent gene transcription with EC₅₀ values of 15.5 ± 9.8 and 1.1 ± 0.9 nM, respectively. Both responses were blocked by two unrelated 5-HT_2CR-selective antagonists (SB-242,084, RS-102,221) but not by a 5-HT_2AR (EMD-281,014) or 5-HT_2BR (RS-127,455) antagonists. By single-cell calcium imaging, we found that serotonin and WAY-161,503 induced robust calcium transients, which were also blunted by both 5-HT_2CR antagonists. Additionally we revealed, first, that 5-HT_2CR induced CRE activation via protein kinase C (PKC)-mediated engagement of extracellular-regulated kinases-1/2 and, second, that intrinsic activity of WAY-161,503 was in the range of 0.3–0.5 compared with serotonin, defining the frequently used 5-HT_2CR agonist as a partial agonist of endogenous 5-HT_2CR. In conclusion, we have shown that hypothalamic mHypoA-2/10 cells endogenously express 5-HT_2CR and thus are the first cell line in which to analyze 5-HT_2CR pharmacology, signaling, and regulation in its natural environment.