Biology subjects

Duarte, C. B.

Publications and source records attributed to Duarte, C. B..

5 recordsLinked to original sources

bioRxiv2026

Disease-associated variants alter a novel physiological role of hnRNP H2 in neuronal development

Heterogeneous nuclear ribonucleoprotein H2 (hnRNP H2) is a member of the hnRNP family of RNA-binding proteins that regulate neuronal function through multiple aspects of RNA metabolism. Mutations in the nuclear localisation signal (NLS) and quasi-RNA recognition motifs (qRRMs) of hnRNP H2 have recently been linked to a severe developmental syndrome, termed HNRNPH2-related neurodevelopmental disorder. Despite its emerging importance, the physiological roles of hnRNP H2 in neuronal development and synaptic maturation, and how disease-associated variants alter these processes to drive pathology, remain poorly understood. Here, we show that hnRNP H2 is expressed in hippocampal neurons and localises to the synaptic compartment. Knockdown of hnRNP H2 revealed essential roles of the protein in the regulation of neuronal morphology, dendritic spine maturation, synaptic transmission, and cortical neuronal migration in vivo. Disease-associated hnRNP H2 variants altered protein localisation and activity-dependent synaptic recruitment, resulting in defects in neuronal morphology, in vivo cortical neuronal migration and laminar orientation, dendritic spine maturation, and synaptic function. Together, these findings establish hnRNP H2 as a critical regulator of neurodevelopment and identify a molecular basis for HNRNPH2-related neurodevelopmental disorder.

neuroscience↗

bioRxiv2025

BDNF-TrkB signaling promotes synaptic GluN2A-NMDA receptor expression and network hyperexcitability in cultured hippocampal neurons and during status epilepticus

Brain-derived neurotrophic factor (BDNF) is a key modulator of synaptic function, acting through activation of TrkB receptors. This neurotrophic factor mediates synaptic plasticity and plays an important role in epileptogenesis, but the underlying molecular mechanisms have not been fully elucidated. In this work, we investigated the role of BDNF-TrkB signaling in the regulation of synaptic GluN2A-containing NMDA receptors (NMDAR), and the impact on network synchronization in cultured hippocampal neurons. Incubation with BDNF increased the synaptic surface expression of GluN2A-containing NMDAR in rat hippocampal synaptoneurosomes and in cultured hippocampal neurons. The effect in the latter preparation was time-dependent and required new protein synthesis. Mechanistically, we identified a signaling cascade involving hnRNPK, the non-receptor tyrosine kinase Pyk2, and protein kinase C (PKC) as essential for mediating BDNF-induced upregulation in the synaptic expression of GluN2A-containing NMDAR. Knockdown of hnRNPK or Pyk2, pharmacological inhibition of PKC, or expression of a phosphorylation -deficient Pyk2 mutant abolished BDNF-induced synaptic surface accumulation of Glu2A. Moreover, Pyk2 phosphorylation at Y402 was necessary for both basal and BDNF-induced synaptic GluN2A expression. Functional multielectrode array (MEA) recordings showed that endogenous BDNF and GluN2A-containing NMDAR contributed to the increase in network activity in cultured hippocampal neurons evoked by stimulation with a cocktail including bicuculline, 4-aminopyridine, and glycine. Importantly, BDNF-TrkB signaling also mediated the upregulation in the hippocampal synaptic surface expression of GluN2A-containing NMDAR, as determined in rats subjected to the pilocarpine model of temporal lobe epilepsy, where increased GluN2A synaptic expression was observed and shown to be TrkB-dependent. These findings unveil a crucial BDNF/TrkB-PKC-Pyk2-hnRNPK signaling axis that regulates synaptic GluN2A levels and network excitability, offering novel insights into the molecular basis of synaptic plasticity.

neuroscience↗

bioRxiv2024

Physioxia-modulated mesenchymal stem cells secretome has higher capacity to preserve neuronal network and translation processes in hypoxic-ischemic encephalopathy in vitro model

Hypoxic-ischemic encephalopathy (HIE) is one of the leading causes of child death worldwide. Most of the survivors develop various neurological diseases, such as cerebral palsy, seizures, and/or motor and behavioral problems. HIE is caused by an episode of perinatal asphyxia, which interrupts the blood supply to the brain. Due to its high energy demands, this interruption initiates glutamate excitotoxic pathways, leading to cell death. Umbilical cord mesenchymal stem cells (UC-MSCs) are gaining attention as a promising complement to the current clinical approach, based on therapeutic hypothermia, which has shown limited efficacy. Previous data have shown that priming MSCs under physiological culture conditions, namely soft platforms (3kPa) - mechanomodulated - or physiological oxygen levels (5% O2) - physioxia - leads to changes in the cellular proteome and their secretome. To evaluate how exposing MSCs to these culture conditions could impact their therapeutic potential, physiologically primed UC-MSCs or their secretome were added to an in vitro HIE model using cortical neurons primary cultures subjected to oxygen and glucose deprivation (OGD) insult. By comparing the neuronal proteome of sham, OGD insulted, and OGD-treated neurons, it was possible to identify proteins whose levels were restored in the presence of UC-MSCs or their secretome. Despite the different approaches that differentially altered UC-MSCs proteome and secretome, the effects converged on the re-establishment of the levels of proteins involved in translation mechanisms (such as the 40S and 60s ribosomal subunits), possibly stabilizing proteostasis, which is known to be essential for neuronal recovery. Interestingly, treatment with the secretome of UC-MSC modulated under physioxic conditions sustained part of the neuronal network integrity and modulated several mitochondrial proteins, including those proteins involved in ATP production. This suggests that the unique composition of the physioxia-modulated secretome may offer a therapeutical advantage in restoring essential cellular processes that help neurons maintain their function, compared to traditionally expanded UC-MSCs. These findings suggest that both the presence of UC-MSCs and their secretome alone can influence multiple targets and signaling pathways, collectively promoting neuronal survival following an OGD insult.

neuroscience↗

bioRxiv2024

Synaptic accumulation of GluN2B-containing NMDA receptors mediates the effects of BDNF-TrkB signalling on synaptic plasticity and in epileptogenesis

Brain-derived neurotrophic factor (BDNF) is a key mediator of synaptic plasticity and memory formation in the hippocampus. However, the BDNF-induced alterations in the glutamate receptors coupled to the plasticity of glutamatergic synapses in the hippocampus have not been elucidated. In this work we investigated the putative role of GluN2B-containing NMDA receptors in the plasticity of glutamatergic synapses induced by BDNF. Stimulation of hippocampal synaptoneurosomes with BDNF led to a significant time-dependent increase in the synaptic surface expression of GluN2B-containing NMDA receptors as determined by immunocytochemistry with colocalization with pre- (vesicular glutamate transporter) and post-synaptic markers (PSD95). Similarly, BDNF induced the synaptic accumulation of GluN2B-containing NMDA receptors at the synapse in cultured hippocampal neurons, by a mechanism sensitive to the PKC inhibitor G[O]6983. The effects of PKC may be mediated by phosphorylation of Pyk2, as suggested by western blot experiments analyzing the phosphorylation of the kinase on Tyrosine 402. GluN2B-containing NMDA receptors mediated the effects of BDNF in the facilitation of the early phase of long-term potentiation (LTP) of hippocampal CA1 synapses induced by {theta}-burst stimulation, since the effect of the neurotrophin was abrogated in the presence of the GluN2B inhibitor Co 101244. In the absence of BDNF, the GluN2B inhibitor did not effect LTP. Surface accumulation of GluN2B-containing NMDA receptors was also observed in hippocampal synaptoneurosomes isolated from rats subjected to the pilocarpine model of temporal lobe epilepsy, after reaching Status epilepticus, an effect that was inhibited by administration of the TrkB receptor inhibitor ANA-12. Together, these results show that the synaptic accumulation of GluN2B-containing NMDA receptors mediate the effects of BDNF in the plasticity of glutamatergic synapses in the hippocampus.

neuroscience↗

bioRxiv2022

Pilot MRI study of carbon monoxide (CO) against ischemic stroke in mice: blood brain barrier integrity and metabolic pattern

Although stroke is the main cause of brain damage worldwide, stroke therapies are based on blood reperfusion and do not target cerebral parenchyma. Ischemic stroke (representing 87% of all strokes) causes cerebral damage due to oxygen and tissue energy depletion, which lead to acidosis, inflammation, excitotoxicity and oxidative stress. Carbon monoxide (CO) is an endogenous gasotransmitter produced by heme oxygenase cleavage of the heme group. CO promotes cytoprotection by limiting inflammation and preventing cell death in several tissues including the brain. Previous studies have demonstrated the protective role of CO in the mouse ischemic stroke model, middle cerebral artery occlusion (MCAo) by histological analysis when CO is when applied before ischemia. Herein, there are two main novelties. First CO is administrated following stroke, which better mimics its potential future use as therapeutic drug. Secondly, imaging techniques were used to elucidate the effect of this gasotransmitter at the metabolic, vascular and anatomic levels. The putative neuroprotective effects of CO following MCAo were assessed by 3 i.p. injections of the CO-releasing molecule CORM-A1 (3 mg/kg), administered 6, 24 and 48h after reperfusion. Magnetic Resonance Imaging was performed 1 day and 7 days after reperfusion using T2-weighted, diffusion weighted images, proton spectroscopy (1H-MRS) and perfusion (dynamic contrast enhanced images). 1H-MRS also allowed the comparison between metabolite signatures at day 1 versus 7 day following MCAo. Furthermore, CORM-A1 limited the loss of blood-brain barrier (BBB) integrity as it reduced the edema formation. Furthermore, the CO donor minimized the metabolite load loss at an early stage after MCAo, both in striatum and cortex. In conclusion and based on MRI analysis, CO has a protective role in the recovery from stroke injury, mainly by acting on BBB integrity and brain metabolism.

neuroscience↗