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Purinergic signaling has been implicated in many biological functions, including development. In this study, we investigate the functions of extracellular adenosine and adenosine receptors using a mouse embryonic stem cell (ESC) line and morula stages isolated from mouse embryos. Feeder-free mouse ESC was investigated in the absence and presence of the leukemia inhibitory factor (LIF), configuring undifferentiated cells and cells undergoing spontaneous differentiation. High alkaline phosphatase (ALPL) and low CD73 levels resulting in low adenosine (eADO) levels were characteristic for pluripotent cells in the presence of the LIF, while LIF deprivation resulted in augmented adenosine levels and reduced pluripotency marker expression, which indicated differentiation. Tracing ESC proliferation by BrdU labeling revealed that the inhibition of ALPL by levamisole resulted in a decrease in proliferation due to less eADO accumulation. Furthermore, caffeine and levamisole treatment, inhibiting adenosine receptor and eADO accumulation, respectively, reduced ESC migration, similar to that observed in the absence of the LIF. Pharmacological approaches of selective adenosine receptor subtype inhibition triggered specific adenosine receptor activities, thus triggering calcium or MAP kinase pathways leading to differentiation. In line with the in vitro data, mouse embryos at the morula stage were sensitive to treatments with A1 and A3 receptor antagonists, leading to the conclusion that A1 receptor and A3 receptor inhibition impairs proliferation and self-renewal and triggers inappropriate differentiation, respectively. The findings herein define the functions of eADO signaling in early development with implications for developmental disorders, in which adenosine receptors or ectonucleotidase dysfunctions are involved, and which could lead to malformations and miscarriages, due to exposure to caffeine.

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Following behavior is a well-documented foraging specialization in Neotropical birds, which consists of individuals of solitary or mixed-flocking species following other moving animals to capture fleeing prey. Here, we report two observations of the cryptic forest-falcon, Micrastur mintoni following troops of primates during an ornithological inventory in the eastern Brazilian Amazon. During both observations, the falcon emitted a typical vocalization and captured fleeing insects dispersed by the primates moving through the forest while foraging. This is the first report of an apparently commensal association between a Micrastur forest falcon and two species of primates.(AU)

O comportamento de following é uma especialização de forrageamento bem documentada em aves neotropicais, que consiste em indivíduos de espécies solitárias ou em bandos mistos seguindo outros animais em movimento para capturar presas afugentadas. Aqui reportamos duas observações do falcão-críptico, Micrastur mintoni seguindo grupos de primatas, durante um inventário ornitológico na Amazônia oriental brasileira. Durante as duas observações, o falcão emitiu uma vocalização típica e capturou insetos afugentados pelos primatas em movimento pela floresta enquanto forrageavam. Esse é o primeiro relato de uma associação aparentemente comensal entre um falcão florestal do gênero Micrastur e duas espécies de primatas.(AU)

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The environment can impose constraints on signal transmission properties such that signals should evolve in predictable directions (Sensory Drive Hypothesis). However, behavioral and ecological factors can limit investment in more than one sensory modality leading to a trade-off in use of different signals (Transfer Hypothesis). In birds, there is mixed evidence for both sensory drive and transfer hypothesis. Few studies have tested sensory drive while also evaluating the transfer hypothesis, limiting understanding of the relative roles of these processes in signal evolution. Here, we assessed both hypotheses using acoustic and visual signals in male and female antwrens (Thamnophilidae), a species-rich group that inhabits diverse environments and exhibits behaviors, such as mixed-species flocking, that could limit investment in different signal modalities. We uncovered significant effects of habitat (sensory drive) and mixed-species flocking behavior on both sensory modalities, and we revealed evolutionary trade-offs between song and plumage complexity, consistent with the transfer hypothesis. We also showed sex- and trait-specific responses in visual signals that suggest both natural and social selection play an important role in the evolution of sexual dimorphism. Altogether, these results support the idea that environmental (sensory drive) and behavioral pressures (social selection) shape signal evolution in antwrens.

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Calcium, the most versatile second messenger, regulates essential biology including crucial cellular events in embryogenesis. We investigated impacts of calcium channels and purinoceptors on neuronal differentiation of normal mouse embryonic stem cells (ESCs), with outcomes being compared to those of in vitro models of Huntington's disease (HD). Intracellular calcium oscillations tracked via real-time fluorescence and luminescence microscopy revealed a significant correlation between calcium transient activity and rhythmic proneuronal transcription factor expression in ESCs stably expressing ASCL-1 or neurogenin-2 promoters fused to luciferase reporter genes. We uncovered that pharmacological manipulation of L-type voltage-gated calcium channels (VGCCs) and purinoceptors induced a two-step process of neuronal differentiation. Specifically, L-type calcium channel-mediated augmentation of spike-like calcium oscillations first promoted stable expression of ASCL-1 in differentiating ESCs, which following P2Y2 purinoceptor activation matured into GABAergic neurons. By contrast, there was neither spike-like calcium oscillations nor responsive P2Y2 receptors in HD-modeling stem cells in vitro. The data shed new light on mechanisms underlying neurogenesis of inhibitory neurons. Moreover, our approach may be tailored to identify pathogenic triggers of other developmental neurological disorders for devising targeted therapies.

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BACKGROUND: Novel developmental functions have been attributed to the P2X7 receptor (P2X7R) including proliferation stimulation and neural differentiation. Mouse embryonic stem cells (ESC), induced with retinoic acid to neural differentiation, closely assemble processes occurring during neuroectodermal development of the early embryo. PRINCIPAL FINDINGS: P2X7R expression together with the pluripotency marker Oct-4 was highest in undifferentiated ESC. In undifferentiated cells, the P2X7R agonist Bz-ATP accelerated cell cycle entry, which was blocked by the specific P2X7R inhibitor KN-62. ESC induced to neural differentiation with retinoic acid, reduced Oct-4 and P2X7R expression. P2X7R receptor-promoted intracellular calcium fluxes were obtained at lower Bz-ATP ligand concentrations in undifferentiated and in neural-differentiated cells compared to other studies. The presence of KN-62 led to increased number of cells expressing SSEA-1, Dcx and ß3-tubulin, as well as the number of SSEA-1 and ß3-tubulin-double-positive cells confirming that onset of neuroectodermal differentiation and neuronal fate determination depends on suppression of P2X7R activity. Moreover, an increase in the number of Ki-67 positive cells in conditions of P2X7R inhibition indicates rescue of progenitors into the cell cycle, augmenting the number of neuroblasts and consequently neurogenesis. CONCLUSIONS: In embryonic cells, P2X7R expression and activity is upregulated, maintaining proliferation, while upon induction to neural differentiation P2X7 receptor expression and activity needs to be suppressed.

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