Regulation of hippocampal postnatal and adult neurogenesis by adenosine A2A receptor: Interaction with brain-derived neurotrophic factor.

Adenosine A2A receptor (A2A R) activation modulates several brain processes, ranging from neuronal maturation to synaptic plasticity. Most of these actions occur through the modulation of the actions of the neurotrophin brain-derived neurotrophic factor (BDNF). In this work, we studied the role of A2A Rs in regulating postnatal and adult neurogenesis in the rat hippocampal dentate gyrus (DG). Here, we show that A2A R activation with CGS 21680 promoted neural stem cell self-renewal, protected committed neuronal cells from cell death and contributed to a higher density of immature and mature neuronal cells, particularly glutamatergic neurons. Moreover, A2A R endogenous activation was found to be essential for BDNF-mediated increase in cell proliferation and neuronal differentiation. Our findings contribute to further understand the role of adenosinergic signaling in the brain and may have an impact in the development of strategies for brain repair under pathological conditions.

Can citation metrics predict the true impact of scientific papers?

Bibliometric quantification is frequently used as metrics for the evaluation of the scientific performance of researchers and institutions. The researchers' merit is usually assessed by the analysis of quantitative parameters such as the number of publications, the impact factor of journals, the total number of citations, or the h-index, although the limitations in translating these indicators into the impact of the outcome of scientific production are a matter of harsh criticism. To assess, based on factual evidences, the validity of traditional bibliometric analyses to conclude on the impact of papers to advance the state of the art, we carried out an innovative methodology on selected publications (test set). This methodology is based on identifying those citations of the test set papers that truly embed the methods, concepts, or hypotheses to build new knowledge and formulate conclusions. The results show that the percentage of citations that reflect the real impact of the papers of the test set has an average value of 12.4% of total citations and is not related to the impact factor of the journal where the test set papers were published. In conclusion, our analysis demonstrates factually, using experimental data, the total failure of using quantitative bulk citation analyses to conclude on the scientific impact of publications. Only a careful analysis of how the work described in papers was embedded on the subsequent work and/or conclusions of others can tell about the real contribution of a published work to the development of new knowledge and advancement of science.

Uptake and metabolism of arginine impact Plasmodium development in the liver.

Prior to infecting erythrocytes and causing malaria symptoms, Plasmodium parasites undergo an obligatory phase of invasion and extensive replication inside their mammalian host's liver cells that depends on the parasite's ability to obtain the nutrients it requires for its intra-hepatic growth and multiplication. Here, we show that L-arginine (Arg) uptake through the host cell's SLC7A2-encoded transporters is essential for the parasite's development and maturation in the liver. Our data suggest that the Arg that is taken up is primarily metabolized by the arginase pathway to produce the polyamines required for Plasmodium growth. Although the parasite may hijack the host's biosynthesis pathway, it relies mainly upon its own arginase-AdoMetDC/ODC pathway to acquire the polyamines it needs to develop. These results identify for the first time a pivotal role for Arg-dependent polyamine production during Plasmodium's hepatic development and pave the way to the exploitation of strategies to impact liver infection by the malaria parasite through the modulation of Arg uptake and polyamine synthesis.

BDNF modulates glycine uptake in hippocampal synaptosomes by decreasing membrane insertion of glycine transporter 2.

Glycine transporter 2 (GlyT2) is localized in the nerve terminals of glycinergic neurons, promoting glycine uptake and ensuring the refilling of glycinergic vesicles. Brain-derived neurotrophic factor (BDNF) activates its high affinity TrkB receptors, which occur in two isoforms, full length (TrkB-FL) and truncated (TrkB-T1/T2). After BDNF binding to TrkB receptor, several intracellular cascades are triggered, specifically PLC, Akt and MAPK signalling pathways. We herein show that BDNF decreases [(3)H]glycine uptake mediated by GlyT2 in isolated nerve endings (synaptosomes) obtained from rat hippocampus, by reducing the maximum velocity (Vmax) of transport while not influencing the transporter affinity constant (Km) for glycine. Western Blot analysis detected both TrkB receptor isoforms in the synaptosomes but the BDNF effect seems to be mediated by TrkB-FL since: 1) the tyrosine kinase inhibitor, k252a, prevented the effect of BDNF, and 2) the effect of BDNF was lost in the presence of specific inhibitors of TrkB signalling pathways, namely U73122, LY294002 and U0126 (inhibitors of PLC, Akt and MAPK pathways, respectively). Monensin, a transporter recycling inhibitor, prevented the BDNF action upon glycine uptake, suggesting that BDNF reduces GlyT2 insertion in the plasma membrane. It is concluded that BDNF effect upon glycine uptake into glycinergic nerve terminals requires the activation of the TrkB-FL receptor and its canonical signalling pathways and occurs by inhibiting GlyT2 membrane incorporation.

BDNF, via truncated TrkB receptor, modulates GlyT1 and GlyT2 in astrocytes.

Glycine transporters (GlyT), GlyT1 and GlyT2, are responsible for the termination of glycine-mediated synaptic activity through removal of neurotransmitter from synaptic cleft. Brain-derived neurotrophic factor (BDNF) activates its high affinity tropomyosin-related kinase (Trk) receptors, namely TrkB, which includes full length (TrkB-FL) and truncated (TrkB-T) isoforms. In this article we evaluated the influence of BDNF upon the activity of glycine transporters in astrocytes. We report that BDNF decreases GlyT1- and GlyT2- mediated [(3) H]glycine transport in primary cultures of astrocytes from rat cerebral cortex. BDNF decreased Vmax but not Km values of transport, which suggests that BDNF induces transporter internalization. Accordingly, dynasore, an inhibitor of dynamin/clathrin-dependent endocytosis, prevented the influence of BDNF upon GlyT-mediated transport. While quantifying mRNA and protein levels, we detected a predominance of truncated isoforms over the TrkB-FL receptor. The effect of BDNF was not abolished by specific inhibitors of PLCγ, PI3K and MAPK, indicating that it did not occur through TrkB-FL canonical pathways. However, BDNF action was lost in the presence of a Rho family-specific blocker (toxin B), a signaling pathway that has been associated to TrkB-T1. Furthermore, the effect of BDNF was abolished upon TrkB-T knockdown in astrocytes by RNA interference. Immunofluorescence assays confirmed an increased GlyT expression in endosomes upon BDNF incubation, which was prevented in the presence of either dynasore or toxin B. We conclude that BDNF, acting on TrkB-T1 receptors, inhibits glycine uptake in astrocytes by promoting GlyT internalization through a Rho-GTPase activity dependent mechanism.

GlyT1 and GlyT2 in brain astrocytes: expression, distribution and function.

GlyT1 and GlyT2 are the transporters responsible for glycine uptake from the synaptic cleft. The expression and function of these two glycine transporters in rat cortical cultured astrocytes over several maturation stages (10, 18 and 24 days in vitro) were herein investigated. Quantitative PCR and western blot showed that both GlyT1 and GlyT2 transcripts and protein were expressed in astrocytes in the examined maturation stages. Double detection of Glial fibrillary acidic protein (GFAP) and GlyT1/GlyT2 revealed that both transporters were detected in the cell body and in the processes of astrocytes. Furthermore, the double immunofluorescence analysis carried out in P21 rat brain slices corroborated the presence of both transporters in cortical and hippocampal astrocytes. The functional characterization of GlyT1 and GlyT2 in cultured astrocytes performed by [(3)H]glycine uptake experiments revealed that both transporters take up glycine in a concentration-dependent way, but with a very distinct affinity. Kinetic analysis revealed a K m of 51.15 ± 4.96 µM and a V max of 379.30 ± 10.31 pmol/min/mg for GlyT1 and a K m of 1,801 ± 148.9 µM and a V max of 5,730 ± 200.2 pmol/min/mg for GlyT2. It is concluded that astrocytes express functional GlyT2, which challenge previous findings that those cells would express only GlyT1, whereas GlyT2 was supposed to be restricted to the glycinergic nerve terminals. Therefore, the work herein reported provides new insights about glycinergic neurotransmission in the brain.

Age-related changes of glycine receptor at the rat hippocampus: from the embryo to the adult.

Glycinergic inhibitory transmission has been described in spinal cord, but rather disregarded in the brain. The spatial-temporal characterization of glycine receptors (GlyR) in the hippocampus over development is herein reported. GlyR expression increases from late embryonic stage (E18) to 7 days postnatal (P7) and decreases from P7 on. Quantitative real-time PCR showed that GlyR subunit expression changes over neuronal maturation with a preponderance of α2 and α3, over α1 and ß. In immature stages, GlyR delineate the cell body of neurons at the Dentate Gyrus and Cornus Ammonis 1 and 3 (CA1/CA3) and are composed of α2 and α3 subunits. At P7, synaptic GlyRα2ß can already be observed in the dendritic areas of Dentate Gyrus and of CA1/CA3. In the mature hippocampus, synaptic GlyR decrease and, although a few synaptic GlyRα1ß can still be detected in the dendritic layers, extrasynaptic α2/α3-containing GlyR and somatic localized GlyRα3 are the most abundant. Our results point towards an important function of a slow tonic activation of extrasynaptic GlyR, over a fast phasic activation of synaptic GlyRα1ß. We clearly show that GlyR are widely expressed in hippocampus and that their subcellular localization and subunit composition change over development.