ABSTRACT
Extracellular adenosine, a key regulator of neuronal excitability, is metabolized by astrocyte-based enzyme adenosine kinase (ADK). We hypothesized that ADK might be an upstream regulator of adenosine-based homeostatic brain functions by simultaneously affecting several downstream pathways. We therefore studied the relationship between ADK expression, levels of extracellular adenosine, synaptic transmission, intrinsic excitability, and brain-derived neurotrophic factor (BDNF)-dependent synaptic actions in transgenic mice underexpressing or overexpressing ADK. We demonstrate that ADK: 1) Critically influences the basal tone of adenosine, evaluated by microelectrode adenosine biosensors, and its release following stimulation; 2) determines the degree of tonic adenosine-dependent synaptic inhibition, which correlates with differential plasticity at hippocampal synapses with low release probability; 3) modulates the age-dependent effects of BDNF on hippocampal synaptic transmission, an action dependent upon co-activation of adenosine A2A receptors; and 4) influences GABAA receptor-mediated currents in CA3 pyramidal neurons. We conclude that ADK provides important upstream regulation of adenosine-based homeostatic function of the brain and that this mechanism is necessary and permissive to synaptic actions of adenosine acting on multiple pathways. These mechanistic studies support previous therapeutic studies and implicate ADK as a promising therapeutic target for upstream control of multiple neuronal signaling pathways crucial for a variety of neurological disorders.
Subject(s)
Adenosine Kinase/physiology , Adenosine/physiology , Homeostasis/physiology , Synapses/physiology , Adenosine Kinase/genetics , Animals , Blotting, Western , Brain-Derived Neurotrophic Factor/physiology , CA1 Region, Hippocampal/physiology , CA3 Region, Hippocampal/physiology , Electrophysiological Phenomena/physiology , Excitatory Postsynaptic Potentials/physiology , Extracellular Space/metabolism , Immunohistochemistry , Mice , Mice, Inbred C57BL , Mice, Transgenic , Mossy Fibers, Hippocampal/physiology , Neuronal Plasticity/physiology , Patch-Clamp Techniques , Purines/metabolism , Receptor, Adenosine A2A/genetics , Receptor, Adenosine A2A/physiology , Receptors, GABA-A/physiology , Synaptic Transmission/physiologyABSTRACT
BACKGROUND: N-Methyl-D-aspartate receptors (NMDARs) play fundamental roles in basic brain functions such as excitatory neurotransmission and learning and memory processes. Their function is largely regulated by factors released by glial cells, including the coagonist d-serine. We investigated whether the activation of microglial CX3CR1 induces the release of factors that modulate NMDAR functions. METHODS: We recorded the NMDAR component of the field excitatory postsynaptic potentials (NMDA-fEPSPs) elicited in the CA1 stratum radiatum of mouse hippocampal slices by Shaffer collateral stimulation and evaluated D-serine content in the extracellular medium of glial primary cultures by mass spectrometry analysis. RESULTS: We demonstrated that CX3CL1 increases NMDA-fEPSPs by a mechanism involving the activity of the adenosine receptor type A2 (A2AR) and the release of the NMDAR coagonist D-serine. Specifically (1) the selective A2AR blocker 7-(2-phenylethyl)-5-amino-2-(2-furyl)-pyrazolo-[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine (SCH58261) and the genetic ablation of A2AR prevent CX3CL1 action while the A2AR agonist 5-(6-amino-2-(phenethylthio)-9H-purin-9-yl)-N-ethyl-3,4-dihydroxytetrahydrofuran-2-carboxamide (VT7) mimics CX3CL1 effect, and (2) the selective blocking of the NMDAR glycine (and D-serine) site by 5,7-dicholorokynurenic acid (DCKA), the enzymatic degradation of D-serine by D-amino acid oxidase (DAAO) and the saturation of the coagonist site by D-serine, all block the CX3CL1 effect. In addition, mass spectrometry analysis demonstrates that stimulation of microglia and astrocytes with CX3CL1 or VT7 increases D-serine release in the extracellular medium. CONCLUSIONS: CX3CL1 transiently potentiates NMDAR function though mechanisms involving A2AR activity and the release of D-serine.
Subject(s)
Chemokine CX3CL1/metabolism , Hippocampus/metabolism , Receptors, Adenosine A2/metabolism , Receptors, N-Methyl-D-Aspartate/metabolism , Serine/metabolism , Animals , Chromatography, Liquid , Excitatory Postsynaptic Potentials/physiology , Mass Spectrometry , Mice , Mice, Inbred C57BL , Neuroglia/metabolism , Organ Culture Techniques , Patch-Clamp TechniquesABSTRACT
In recent years several evidence demonstrated that some features of hippocampal biology, like neurogenesis, synaptic transmission, learning, and memory performances are deeply modulated by social, motor, and sensorial experiences. Fractalkine/CX(3)CL1 is a transmembrane chemokine abundantly expressed in the brain by neurons, where it modulates glutamatergic transmission and long-term plasticity processes regulating the intercellular communication between glia and neurons, being its specific receptor CX(3)CR1 expressed by microglia. In this paper we investigated the role of CX(3)CL1/CX(3)CR1 signaling on experience-dependent hippocampal plasticity processes. At this aim wt and CX(3)CR1(GFP/GFP) mice were exposed to long-lasting-enriched environment (EE) and the effects on hippocampal functions were studied by electrophysiological recordings of long-term potentiation of synaptic activity, behavioral tests of learning and memory in the Morris water maze paradigm and analysis of neurogenesis in the subgranular zone of the dentate gyrus (DG). We found that CX(3)CR1 deficiency increases hippocampal plasticity and spatial memory, blunting the potentiating effects of EE. In contrast, exposure to EE increased the number and migration of neural progenitors in the DG of both wt and CX(3)CR1(GFP/GFP) mice. These data indicate that CX(3)CL1/CX(3)CR1-mediated signaling is crucial for a normal experience-dependent modulation of hippocampal functions.
ABSTRACT
Microglia are highly motile phagocytic cells that infiltrate and take up residence in the developing brain, where they are thought to provide a surveillance and scavenging function. However, although microglia have been shown to engulf and clear damaged cellular debris after brain insult, it remains less clear what role microglia play in the uninjured brain. Here, we show that microglia actively engulf synaptic material and play a major role in synaptic pruning during postnatal development in mice. These findings link microglia surveillance to synaptic maturation and suggest that deficits in microglia function may contribute to synaptic abnormalities seen in some neurodevelopmental disorders.
Subject(s)
Brain/growth & development , Hippocampus/growth & development , Hippocampus/physiology , Microglia/physiology , Synapses/physiology , Animals , Brain/physiology , CX3C Chemokine Receptor 1 , Chemokine CX3CL1/metabolism , Dendritic Spines/physiology , Dendritic Spines/ultrastructure , Disks Large Homolog 4 Protein , Excitatory Postsynaptic Potentials , Guanylate Kinases/analysis , Long-Term Synaptic Depression , Membrane Proteins/analysis , Mice , Mice, Knockout , Miniature Postsynaptic Potentials , Neuronal Plasticity , Patch-Clamp Techniques , Pyramidal Cells/physiology , Receptors, Chemokine/genetics , Receptors, Chemokine/metabolism , Receptors, Cytokine/genetics , Receptors, Cytokine/metabolism , Receptors, HIV/genetics , Receptors, HIV/metabolism , Signal Transduction , Synaptosomal-Associated Protein 25/analysisABSTRACT
We have examined how the chemokine fractalkine/CX(3)CL1 influences long-term potentiation (LTP) in CA1 mouse hippocampal slices. Field potentials (fEPSPs) were recorded upon electrical stimulation of Schaffer collaterals. It was found that application of CX(3)CL1 inhibits LTP when present during the critical induction period. LTP impairment (i) failed to occur in CX(3)CR1 deficient mice (CX(3)CR1(GFP/GFP)) and in the presence of okadaic acid (OA); (ii) required the activation of adenosine receptor 3 (A(3)R), since it was prevented in A(3)R-deficient mice or by MRS1523, a selective A(3)R antagonist. Together, these findings indicate that CX(3)CL1 inhibits hippocampal LTP through A(3)R activity.
