Luan Pereira Diniz

Astrocyte-induced Synaptogenesis Is Mediated by Transforming Growth Factor β Signaling through Modulation of d-Serine Levels in Cerebral Cortex Neurons

Background: Synapse formation and function is modulated by intrinsic and extrinsic non-autonomous factors. Results: Astrocytes induce synapse formation through TGF-β1 pathway. TGF-β1 synaptogenic property is dependent on d-serine signaling. Conclusion: TGF-β induces excitatory glutamatergic synapses in vertebrates. Significance: This is a novel molecular mechanism that might impact synaptic function and shed light on new potential therapeutic targets for synaptic deficit diseases. Assembly of synapses requires proper coordination between pre- and postsynaptic elements. Identification of cellular and molecular events in synapse formation and maintenance is a key step to understand human perception, learning, memory, and cognition. A key role for astrocytes in synapse formation and function has been proposed. Here, we show that transforming growth factor β (TGF-β) signaling is a novel synaptogenic pathway for cortical neurons induced by murine and human astrocytes. By combining gain and loss of function approaches, we show that TGF-β1 induces the formation of functional synapses in mice. Further, TGF-β1-induced synaptogenesis involves neuronal activity and secretion of the co-agonist of the NMDA receptor, d-serine. Manipulation of d-serine signaling, by either genetic or pharmacological inhibition, prevented the TGF-β1 synaptogenic effect. Our data show a novel molecular mechanism that might impact synaptic function and emphasize the evolutionary aspect of the synaptogenic property of astrocytes, thus shedding light on new potential therapeutic targets for synaptic deficit diseases.

Astrocyte transforming growth factor beta 1 promotes inhibitory synapse formation via CaM kinase II signaling.

The balance between excitatory and inhibitory synaptic inputs is critical for the control of brain function. Astrocytes play important role in the development and maintenance of neuronal circuitry. Whereas astrocytes‐derived molecules involved in excitatory synapses are recognized, molecules and molecular mechanisms underlying astrocyte‐induced inhibitory synapses remain unknown. Here, we identified transforming growth factor beta 1 (TGF‐β1), derived from human and murine astrocytes, as regulator of inhibitory synapse in vitro and in vivo. Conditioned media derived from human and murine astrocytes induce inhibitory synapse formation in cerebral cortex neurons, an event inhibited by pharmacologic and genetic manipulation of the TGF‐β pathway. TGF‐β1‐induction of inhibitory synapse depends on glutamatergic activity and activation of CaM kinase II, which thus induces localization and cluster formation of the synaptic adhesion protein, Neuroligin 2, in inhibitory postsynaptic terminals. Additionally, intraventricular injection of TGF‐β1 enhanced inhibitory synapse number in the cerebral cortex. Our results identify TGF‐β1/CaMKII pathway as a novel molecular mechanism underlying astrocyte control of inhibitory synapse formation. We propose here that the balance between excitatory and inhibitory inputs might be provided by astrocyte signals, at least partly achieved via TGF‐β1 downstream pathways. Our work contributes to the understanding of the GABAergic synapse formation and may be of relevance to further the current knowledge on the mechanisms underlying the development of various neurological disorders, which commonly involve impairment of inhibitory synapse transmission. GLIA 2014;62:1917–1931

Astrocytic control of neural circuit formation: highlights on TGF-beta signaling.

Brain function depends critically on the coordinated activity of presynaptic and postsynaptic signals derived from both neurons and non-neuronal elements such as glial cells. A key role for astrocytes in neuronal differentiation and circuitry formation has emerged within the last decade. Although the function of glial cells in synapse formation, elimination and efficacy has greatly increased, we are still very far from deeply understanding the molecular and cellular mechanism underlying these events. The present review discusses the mechanisms driving astrocytic control of excitatory and inhibitory synapse formation in the central nervous system, especially the mechanisms mediated by soluble molecules, particularly those from the TGF-β family. Further, we discuss whether and how human astrocytes might contribute to the acquisition of human cognition. We argue that understanding how astrocytic signals regulate synaptic development might offer new insights into human perception, learning, memory, and cognition and, ultimately, provide new targets for the treatment of neurological diseases.

Astrocyte Transforming Growth Factor Beta 1 Protects Synapses against Aβ Oligomers in Alzheimer's Disease Model

Alzheimers disease (AD) is characterized by progressive cognitive decline, increasingly attributed to neuronal dysfunction induced by amyloid-β oligomers (AβOs). Although the impact of AβOs on neurons has been extensively studied, only recently have the possible effects of AβOs on astrocytes begun to be investigated. Given the key roles of astrocytes in synapse formation, plasticity, and function, we sought to investigate the impact of AβOs on astrocytes, and to determine whether this impact is related to the deleterious actions of AβOs on synapses. We found that AβOs interact with astrocytes, cause astrocyte activation and trigger abnormal generation of reactive oxygen species, which is accompanied by impairment of astrocyte neuroprotective potential in vitro. We further show that both murine and human astrocyte conditioned media (CM) increase synapse density, reduce AβOs binding, and prevent AβO-induced synapse loss in cultured hippocampal neurons. Both a neutralizing anti-transforming growth factor-β1 (TGF-β1) antibody and siRNA-mediated knockdown of TGF-β1, previously identified as an important synaptogenic factor secreted by astrocytes, abrogated the protective action of astrocyte CM against AβO-induced synapse loss. Notably, TGF-β1 prevented hippocampal dendritic spine loss and memory impairment in mice that received an intracerebroventricular infusion of AβOs. Results suggest that astrocyte-derived TGF-β1 is part of an endogenous mechanism that protects synapses against AβOs. By demonstrating that AβOs decrease astrocyte ability to protect synapses, our results unravel a new mechanism underlying the synaptotoxic action of AβOs in AD. SIGNIFICANCE STATEMENT Alzheimers disease is characterized by progressive cognitive decline, mainly attributed to synaptotoxicity of the amyloid-β oligomers (AβOs). Here, we investigated the impact of AβOs in astrocytes, a less known subject. We show that astrocytes prevent synapse loss induced by AβOs, via production of transforming growth factor-β1 (TGF-β1). We found that AβOs trigger morphological and functional alterations in astrocytes, and impair their neuroprotective potential. Notably, TGF-β1 reduced hippocampal dendritic spine loss and memory impairment in mice that received intracerebroventricular infusions of AβOs. Our results describe a new mechanism underlying the toxicity of AβOs and indicate novel therapeutic targets for Alzheimers disease, mainly focused on TGF-β1 and astrocytes.

Interaction of amyloid-β (Aβ) oligomers with neurexin 2α and neuroligin 1 mediates synapse damage and memory loss in mice

Brain accumulation of the amyloid-β protein (Aβ) and synapse loss are neuropathological hallmarks of Alzheimer disease (AD). Aβ oligomers (AβOs) are synaptotoxins that build up in the brains of patients and are thought to contribute to memory impairment in AD. Thus, identification of novel synaptic components that are targeted by AβOs may contribute to the elucidation of disease-relevant mechanisms. Trans-synaptic interactions between neurexins (Nrxs) and neuroligins (NLs) are essential for synapse structure, stability, and function, and reduced NL levels have been associated recently with AD. Here we investigated whether the interaction of AβOs with Nrxs or NLs mediates synapse damage and cognitive impairment in AD models. We found that AβOs interact with different isoforms of Nrx and NL, including Nrx2α and NL1. Anti-Nrx2α and anti-NL1 antibodies reduced AβO binding to hippocampal neurons and prevented AβO-induced neuronal oxidative stress and synapse loss. Anti-Nrx2α and anti-NL1 antibodies further blocked memory impairment induced by AβOs in mice. The results indicate that Nrx2α and NL1 are targets of AβOs and that prevention of this interaction reduces the deleterious impact of AβOs on synapses and cognition. Identification of Nrx2α and NL1 as synaptic components that interact with AβOs may pave the way for development of novel approaches aimed at halting synapse failure and cognitive loss in AD.

Neuroprotective astrocyte-derived insulin/insulin-like growth factor 1 stimulates endocytic processing and extracellular release of neuron-bound Aβ oligomers

Synaptopathy underlying memory deficits in Alzheimers disease (AD) is increasingly thought to be instigated by toxic oligomers of the amyloid beta peptide (AβOs). Given the long latency and incomplete penetrance of AD dementia with respect to Aβ pathology, we hypothesized that factors present in the CNS may physiologically protect neurons from the deleterious impact of AβOs. Here we employed physically-separated neuron-astrocyte co-cultures to investigate potential non cell-autonomous neuroprotective factors influencing AβO toxicity. Neurons cultivated in the absence of an astrocyte feeder layer showed abundant AβO binding to dendritic processes and associated synapse deterioration. In contrast, neurons in the presence of astrocytes showed markedly reduced AβO binding and synaptopathy. Results identified the protective factors released by astrocytes as insulin and IGF1. The protective mechanism involved release of newly bound AβOs into the extracellular medium dependent upon trafficking that was sensitive to exosome pathway inhibitors. Delaying insulin treatment led to AβO binding that was no longer releasable. The neuroprotective potential of astrocytes itself was sensitive to chronic AβO exposure, which reduced insulin/IGF1 expression. Our findings support the idea that physiological protection against synaptotoxic AβOs can be mediated by astrocyte-derived insulin/IGF1, but that this protection itself is vulnerable to AβO buildup.Findings reveal a novel basis for protecting CNS neurons against Aβ oligomers (AβOs), neurotoxins believed to instigate neural damage leading to Alzheimer’s dementia. Results with spatially separated cocultures of astrocytes and hippocampal neurons show an exosome-like mechanism by which insulin/IGF1 from astrocytes clear bound AβOs from neuronal surfaces.

Flavonoid Hesperidin Induces Synapse Formation and Improves Memory Performance through the Astrocytic TGF-β1.

Synapse formation and function are critical events for the brain function and cognition. Astrocytes are active participants in the control of synapses during development and adulthood, but the mechanisms underlying astrocyte synaptogenic potential only began to be better understood recently. Currently, new drugs and molecules, including the flavonoids, have been studied as therapeutic alternatives for modulation of cognitive processes in physiological and pathological conditions. However, the cellular targets and mechanisms of actions of flavonoids remain poorly elucidated. In the present study, we investigated the effects of hesperidin on memory and its cellular and molecular targets in vivo and in vitro, by using a short-term protocol of treatment. The novel object recognition test (NOR) was used to evaluate memory performance of mice intraperitoneally treated with hesperidin 30 min before the training and again before the test phase. The direct effects of hesperidin on synapses and astrocytes were also investigated using in vitro approaches. Here, we described hesperidin as a new drug able to improve memory in healthy adult mice by two main mechanisms: directly, by inducing synapse formation and function between hippocampal and cortical neurons; and indirectly, by enhancing the synaptogenic ability of cortical astrocytes mainly due to increased secretion of transforming growth factor beta-1 (TGF-β1) by these cells. Our data reinforces the known neuroprotective effect of hesperidin and, by the first time, characterizes its synaptogenic action on the central nervous system (CNS), pointing astrocytes and TGF-β1 signaling as new cellular and molecular targets of hesperidin. Our work provides not only new data regarding flavonoid’s actions on the CNS but also shed light on possible new therapeutic alternative based on astrocyte biology.

Effects of Transforming Growth Factor Beta 1 in Cerebellar Development: Role in Synapse Formation

Granule cells (GC) are the most numerous glutamatergic neurons in the cerebellar cortex and represent almost half of the neurons of the central nervous system. Despite recent advances, the mechanisms of how the glutamatergic synapses are formed in the cerebellum remain unclear. Among the TGF-β family, TGF-beta 1 (TGF-β1) has been described as a synaptogenic molecule in invertebrates and in the vertebrate peripheral nervous system. A recent paper from our group demonstrated that TGF-β1 increases the excitatory synapse formation in cortical neurons. Here, we investigated the role of TGF-β1 in glutamatergic cerebellar neurons. We showed that the expression profile of TGF-β1 and its receptor, TβRII, in the cerebellum is consistent with a role in synapse formation in vitro and in vivo. It is low in the early postnatal days (P1–P9), increases after postnatal day 12 (P12), and remains high until adulthood (P30). We also found that granule neurons express the TGF-β receptor mRNA and protein, suggesting that they may be responsive to the synaptogenic effect of TGF-β1. Treatment of granular cell cultures with TGF-β1 increased the number of glutamatergic excitatory synapses by 100%, as shown by immunocytochemistry assays for presynaptic (synaptophysin) and post-synaptic (PSD-95) proteins. This effect was dependent on TβRI activation because addition of a pharmacological inhibitor of TGF-β, SB-431542, impaired the formation of synapses between granular neurons. Together, these findings suggest that TGF-β1 has a specific key function in the cerebellum through regulation of excitatory synapse formation between granule neurons.

Glia: dos velhos conceitos às novas funções de hoje e as que ainda virão

Descritas ha mais de 150 anos, as celulas gliais, constituintes do tecido nervoso juntamente com os neuronios, foram consideradas ate pouco tempo celulas de suporte do cerebro, passivas e a margem do seu funcionamento. Especialmente na ultima decada, as neurociencias foram palco de uma mudanca de paradigma relacionada a funcao e ao papel dessas celulas na fisiologia e patologia neurais. Neste artigo, discutimos como os avancos acerca do conhecimento sobre os astrocitos, o mais abundante tipo glial, contribuiram para o entendimento do funcionamento cerebral. Apresentamos evidencias da relacao entre disfuncoes gliais e doencas neurodegenerativas e desordens neurologicas, discutindo o potencial papel dessas celulas na elaboracao de abordagens terapeuticas para o sistema nervoso adulto.

Brain infusion of α-synuclein oligomers induces motor and non-motor Parkinson’s disease-like symptoms in mice

Abstract Parkinson’s disease (PD) is characterized by motor dysfunction, which is preceded by a number of non‐motor symptoms including olfactory deficits. Aggregation of &agr;‐synuclein (&agr;‐syn) gives rise to Lewy bodies in dopaminergic neurons and is thought to play a central role in PD pathology. However, whether amyloid fibrils or soluble oligomers of &agr;–syn are the main neurotoxic species in PD remains controversial. Here, we performed a single intracerebroventricular (i.c.v.) infusion of &agr;‐syn oligomers (&agr;‐SYOs) in mice and evaluated motor and non‐motor symptoms. Familiar bedding and vanillin essence discrimination tasks showed that &agr;‐SYOs impaired olfactory performance of mice, and decreased TH and dopamine levels in the olfactory bulb early after infusion. The olfactory deficit persisted until 45 days post‐infusion (dpi). &agr;‐ SYO‐infused mice behaved normally in the object recognition and forced swim tests, but showed increased anxiety‐like behavior in the open field and elevated plus maze tests 20 dpi. Finally, administration of &agr;‐SYOs induced late motor impairment in the pole test and rotarod paradigms, along with reduced TH and dopamine content in the caudate putamen, 45 dpi. Reduced number of TH‐positive cells was also seen in the substantia nigra of &agr;‐SYO‐injected mice compared to control. In conclusion, i.c.v. infusion of &agr;‐SYOs recapitulated some of PD‐associated non‐motor symptoms, such as increased anxiety and olfactory dysfunction, but failed to recapitulate memory impairment and depressive‐like behavior typical of the disease. Moreover, &agr;‐SYOs i.c.v. administration induced motor deficits and loss of TH and dopamine levels, key features of PD. Results point to &agr;‐syn oligomers as the proximal neurotoxins responsible for early non‐motor and motor deficits in PD and suggest that the i.c.v. infusion model characterized here may comprise a useful tool for identification of PD novel therapeutic targets and drug screening.