Moreover, people working with animals have received appropriate education (FELASA course) as required by the Portuguese authorities. Embryonic Protosappanin B hypothalamic neurospheres culture Hypothalamic neuroprogenitor cells were isolated and cultured as floating neurospheres as previously described [8]. of undifferentiated cells (SOX-2+ cells). Additionally, fluoxetine-induced proliferation and maintenance of hypothalamic neuroprogenitor cells leads to changes in the mRNA levels of appetite regulator neuropeptides, including Neuropeptide Y (NPY) and Cocaine-and-Amphetamine-Regulated-Transcript (CART). This study provides the first evidence that SSRIs affect the development of hypothalamic neuroprogenitor cells with consequent alterations on appetite neuropeptides. Introduction Food intake and body weight Protosappanin B are centrally regulated by the hypothalamus, where arcuate nucleus (ARC) neurons sense and integrate peripheral signals of nutrition to downstream circuits [1], [2]. ARC neurons are divided in two populations acting together to regulate appetite: the orexigenic NPY/AgRP (Neuropeptide Y/Agouti-Related Protein) neurons and the anorexigenic POMC/CART (Pro-OpioMelanocortin/Cocaine-and-Amphetamine-Regulated-Transcript) neurons. Adult hypothalamic neurogenesis occurs at low rates in rodents but it is essential for body weight and feeding regulation [3], [4]. During the embryonic period, hypothalamic neuronal precursors are generated between days E10.5 and E14.5 [5], [6] and persist until adulthood Rabbit polyclonal to pdk1 [7]. In accordance, neuroprogenitor cells can be isolated from fetal and adult hypothalamus and these cells express neuropeptides important for the regulation of feeding [8]. Proliferation of hypothalamic neuroprogenitor cells during the perinatal period is influenced by maternal nutrition and hormones availability in mice [9], [10]. Notably, this will result in body weight and appetite defects in newborns that persist after weaning [11]. Moreover, these feeding alterations indicate a possible change of neuropeptides levels in hypothalamic cells. Based on these studies we can hypothesize that any molecule affecting hypothalamic cell proliferation during the neurodevelopment period can modify the programming of hypothalamic satiety pathways leading to persistent changes in newborns homeostasis. Selective serotonin reuptake inhibitors (SSRIs) are antidepressant drugs also known for their neurogenic effect in perinatal hippocampal and cerebellar neuroprogenitor cells [12], [13]. Additionally, SSRIs obtained from maternal lactation have proven to restore hippocampal neurogenesis in stressed rat offspring [14]. Nevertheless, the potential proliferative effect of SSRI in the perinatal hypothalamus is unknown and requires investigation since SSRIs are the drug of choice for treating depressed pregnant and postpartum women [15]. In fact, there is a 10C16% prevalence of depression during pregnancy and 25% of depressed women continue antidepressant use during pregnancy [16], [17]. As most SSRIs reach the fetus via the placenta and are detectable in breast milk [18], [19] a significant number of children are exposed to SSRIs during critical phases of hypothalamic neurodevelopment. Accordingly, it has been reported that maternal exposure to SSRIs results in low birth weight and modifications of the hypothalamic-pituitary-adrenal axis of human and rodent newborns [20], [21], [22], [23]. Therefore, in this study we investigated whether the SSRI fluoxetine alters the proliferation and differentiation of rat embryonic hypothalamic neuroprogenitor cells. Moreover, using an model previously described by our group [8], we evaluated the effect of fluoxetine in the expression levels of hypothalamic neuropeptides that regulate food intake, including orexigenic (NPY and AgRP) and anorexigenic (POMC and CART) neuropeptides. Materials and Methods Ethics statement All experimental procedures were performed in accordance with the European Union Directive 86/609/EEC for the care and use of laboratory animals. In addition, animals were housed in a licensed animal facility (international Animal Welfare Assurance number 520.000.000.2006) and the CNC animal experimentation board approved the utilization of animals for this project. Moreover, people working with animals have received appropriate education (FELASA course) as required by the Portuguese authorities. Embryonic hypothalamic neurospheres culture Hypothalamic neuroprogenitor cells were isolated and cultured Protosappanin B as floating neurospheres as previously described [8]. Briefly, hypothalamic tissue dissected from rat embryos (E18-19) were mechanically dissociated by means of a Pasteur pipette. Cells were suspended.
