New insights on trazodone effect in promoting neuronal differentiation of adult murine hippocampal neural progenitor cells

New insights on trazodone effect in promoting neuronal differentiation of adult murine hippocampal neural progenitor cells

S234 P.1.g. Basic and clinical neuroscience − Neuropharmacology We fit the uptake data to sigmoidal dose–response curves and determined IC50 values. ...

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S234

P.1.g. Basic and clinical neuroscience − Neuropharmacology

We fit the uptake data to sigmoidal dose–response curves and determined IC50 values. To assess monoamine release, we preloaded the NET, DAT, or SERT expressing HEK 293 cells with respective neurotransmitter before adding the test substances to initiate monoamine release. The monoamine release was compared with vehicle control using ANOVA followed by Dunnett’s tests. In addition, we assessed binding affinities to monoamine receptors using HEK 293 cell membranes expressing the respective human receptors. Results: All novel phenethylamine derivatives most potently inhibited the NET. 3,4-DMMC and 4-MA inhibited the SERT more potently than the DAT while 5-IT, 3-MMC, and N-methyl2-AI inhibited the DAT slightly more potently than the SERT. All drugs caused monoamine release. Additionally, 5-IT exhibited high affinity for the 5-HT2A receptor and N-methyl-2-AI exhibited strong binding to the a2A receptor. Conclusion: All substances strongly inhibited the NET, as it is known for classic amphetamines, suggesting marked sympathomimetic properties. 3,4-DMMC and 4-MA inhibited the SERT more strongly than the DAT, exhibiting pharmacological properties similar to 3,4-methylenedioxymethamphetamine (MDMA) and suggesting similar empathogenic effects. 5-IT is structurally closely related to the tryptamine hallucinogens and its strong stimulation of the 5-HT2A receptor likely results in hallucinogenlike properties as well. References [1] B¨ackberg, M., Beck, O., Hult´en, P., Rosengren-Holmberg, J., Helander, A., 2014. Intoxications of the new psychoactive substance 5-(2-aminopropyl)indole (5-IT): a case series from the Swedish STRIDA project. Clin Toxicol 52(6), 618–624. [2] Blanckaert, P., van Amsterdam, J., Brunt, T., van den Berg, J., Van Durme, F., Maudens, K., van Bussel, J., 2013. 4-Methyl-amphetamine: a health threat for recreational amphetamine users. J Psychopharmacol 27(9), 817–822.

P.1.g.037 New insights on trazodone effect in promoting neuronal differentiation of adult murine hippocampal neural progenitor cells F. Mancini1 ° , V. Bortolotto2 , G. Mangano1 , C. Milanese1 , L. Polenzani1 , F. Garofolo1 , S. Tongiani1 , M. Grilli2 1 Angelini S.p.A., R&D Angelini Research Center, Piazzale della Stazione 00071 S. Palomba-Pomezia Roma, Italy; 2 University of Piemonte Orientale, Dept Pharmaceutical Sciences, Via Bovio 6 - 28100 Novara, Italy Introduction: Adult hippocampal neurogenesis (ahNG) is a peculiar form of neuroplasticity occurring in mammalian brain. Studies in rodent models suggested that generation of new hippocampal neurons in adulthood plays a role in forms of memory, mood regulation and responsiveness to stress. AhNG is also deregulated in neuropsychiatric diseases, including major depressive disorder [1]. Indeed, several groups demonstrated that chronic treatment with antidepressants can result in increased hippocampal neurogenesis, suggesting that this property may contribute to the therapeutic activity of these drugs [2]. Trazodone (TZD), an antidepressant drug acting through 5-HT receptors and inhibiting the 5-HT transporter, has different pharmaco-therapeutic profiles according to the dose used. For instance antidepressant activity occurs following administration of high doses, while effects on sleep are evident at much lower doses [3]. Methods: The in vitro model of murine adult hippocampal neural progenitor cells (ahNPC) was used to evaluate the effect of

TZD on ahNG [4], the expression of serotoninergic and adrenergic receptor subtypes was investigated by RT-PCR. Then the effects of TZD (0.0003–100 mM) added to ahNPC cultures for 24 h were investigated. The proliferation rate of NPC in presence/ absence of TZD was assessed in comparison with fluoxetine and paroxetine. The effect of TZD and reference molecules on neuronal and non neuronal differentiation of adult NPC was evaluated. For these studies the following phenotypic markers were utilized in single/double immunofluorescence: nestin (for stem cells/undifferentiated progenitors), MAP-2 (for mature neurons), GFAP (for astrocyte) and NG2 (for oligodendrocyte precursors). Results: The results confirmed the expression of SERT, serotoninergic (5-HT1A, 5-HT2A, 5-HT2C) and adrenergic receptors (a1a, a1b, a2a, a2b) in ahNPC. Experiments assessing ahNPC differentiation in presence of TZD revealed that the drug is to selectively promote only neuronal differentiation. Indeed, compared to vehicle, TZD from 0.01 to 0.3 mM significantly increased the percentage of mature (MAP-2+ cells) and immature neurons (MAP-2+/nestin+ cells) (One-Way ANOVA, Tukey post hoc, p < 0.001) generated in vitro by ahNPC under differentiating conditions. Moreover, TZD had no effect on ahNPC differentiation toward non neuronal lineages as tested by evaluating percentages of GFAP and NG2 immunopositive cells (One-Way ANOVA, Tukey post hoc, p > 0.05). At all tested concentrations TZD, unlike paroxetine and fluoxetine, was devoid of negative effects on the viability of ahNPC and their progeny (One-Way ANOVA, Tukey post hoc, p > 0.05). Additionally, TZD had no effect on ahNPC proliferation. Conclusions: Together these data showed, for the first time, that TZD is a proneurogenic drug which specifically promotes neuronal differentiation of murine ahNPC. Additionally they revealed interesting differences between TZD and other antidepressant drugs, which are known to promote ahNG in vivo. Studies aimed at dissecting the relative contribution of distinct receptor subtypes and downstream intracellular pathways to TZD proneurogenic effects are currently ongoing in murine and also in human NPC cultures. References [1] Apple, D.M., Fonseca, R.S., Kokovay, E., 2016. The role of adult neurogenesis in psychiatric and cognitive disorders. Brain Res http:// dx.doi.org/10.1016/j.brainres.2016.01.023. [2] Eisch, A.J., Petrik, D., 2012. Depression and hippocampal neurogenesis: a road to remission? Science 338, 72−75. [3] Stahl SM., 2009. Mechanism of action of trazodone: a multifunctional drug. CNS Spectr; 14(10): 536−46. [4] Meneghini, V., Bortolotto, V., Francese M.T., Dellarole, A., Carraro, L., Terzieva, S., Grilli, M., 2013. High-mobility group box-1 protein and b-amyloid oligomers promote neuronal differentiation of adult hippocampal neural progenitors via receptor for advanced glycation end products/nuclear factor-úB axis: relevance for Alzheimer’s disease. J Neurosci 33, 6047–6059.

P.1.g.038 Antimuscarinic-induced convulsions in fasted animals after food intake produced no spontaneous seizures, behavioral changes, or neuronal damage N. Enginar1 ° , A. Nurten2 , A. Zengin T¨urkmen2 , G.I. G¨undo˘gan3 , Z.G. Oz¨unal1 1 Istanbul Medical School, Istanbul, Turkey; 2 Department of Physiology, Faculty of Medicine, Yeni Y¨ uzyıl University, Istanbul, Turkey; 3 Department of Histology and Embryology, Yeni Y¨uzyıl University, Istanbul, Turkey Background: Mice and rats treated with scopolamine, atropine or biperiden after fasting for two days or less develop convulsions