EphB2-dependent signaling promotes neuronal excitotoxicity and inflammation in the acute phase of ischemic stroke

Local cerebral hypoperfusion causes ischemic stroke while driving multiple cell-specific responses including inflammation, glutamate-induced neurotoxicity mediated via NMDAR, edema formation and angiogenesis. Despite the relevance of these pathophysiological mechanisms for disease progression and outcome, molecular determinants controlling the onset of these processes are only partially understood. In this context, our study intended to investigate the functional role of EphB2, a receptor tyrosine kinase that is crucial for synapse function and binds to membrane-associated ephrin-B ligands. Cerebral ischemia was induced in Ephb2−/− mice by transient middle cerebral artery occlusion followed by different times (6, 12, 24 and 48 h) of reperfusion. Histological, neurofunctional and transcriptome analyses indicated an increase in EphB2 phosphorylation under these conditions and attenuated progression of stroke in Ephb2−/− mice. Moreover, while infiltration of microglia/macrophages and astrocytes into the peri-infarct region was not altered, expression of the pro-inflammatory mediators MCP-1 and IL-6 was decreased in these mice. In vitro analyses indicated that binding of EphB2 to astrocytic ephrin-B ligands stimulates NF-κB-mediated cytokine expression via the MAPK pathway. Further magnetic resonance imaging of the Ephb2−/− ischemic brain revealed a lower level of cytotoxic edema formation within 6 h upon onset of reperfusion. On the mechanistic level, absence of neuronal EphB2 decreased the mitochondrial Ca2+ load upon specific activation of NMDAR but not during synaptic activity. Furthermore, neuron-specific loss of ephrin-B2 reduced the extent of cerebral tissue damage in the acute phase of ischemic stroke. Collectively, EphB2 may promote the immediate response to an ischemia-reperfusion event in the central nervous system by (i) pro-inflammatory activation of astrocytes via ephrin-B-dependent signaling and (ii) amplification of NMDA-evoked neuronal excitotoxicity

Neuron-astrocyte metabolic coupling facilitates spinal plasticity and maintenance of inflammatory pain

Long-lasting pain stimuli can trigger maladaptive changes in the spinal cord, reminiscent of plasticity associated with memory formation. Metabolic coupling between astrocytes and neurons has been implicated in neuronal plasticity and memory formation in the central nervous system, but neither its involvement in pathological pain nor in spinal plasticity has been tested. Here we report a form of neuroglia signalling involving spinal astrocytic glycogen dynamics triggered by persistent noxious stimulation via upregulation of the Protein Targeting to Glycogen (PTG) in spinal astrocytes. PTG drove glycogen build-up in astrocytes, and blunting glycogen accumulation and turnover by Ptg gene deletion reduced pain-related behaviours and promoted faster recovery by shortening pain maintenance in mice. Furthermore, mechanistic analyses revealed that glycogen dynamics is a critically required process for maintenance of pain by facilitating neuronal plasticity in spinal lamina 1 neurons. In summary, our study describes a previously unappreciated mechanism of astrocyte-neuron metabolic communication through glycogen breakdown in the spinal cord that fuels spinal neuron hyperexcitability

Inositol-1,4,5-trisphosphate receptor-mediated Ca2+ waves in pyramidal neuron dendrites propagate through hot spots and cold spots

We studied inositol-1,4,5-trisphosphate (IP3) receptor-dependent intracellular Ca2+ waves in CA1 hippocampal and layer V medial prefrontal cortical pyramidal neurons using whole-cell patch-clamp recordings and Ca2+ fluorescence imaging. We observed that Ca2+ waves propagate in a saltatory manner through dendritic regions where increases in the intracellular concentration of Ca2+ ([Ca2+]i) were large and fast (‘hot spots’) separated by regions where increases in [Ca2+]i were comparatively small and slow (‘cold spots’). We also observed that Ca2+ waves typically initiate in hot spots and terminate in cold spots, and that most hot spots, but few cold spots, are located at dendritic branch points. Using immunohistochemistry, we found that IP3 receptors (IP3Rs) are distributed in clusters along pyramidal neuron dendrites and that the distribution of inter-cluster distances is nearly identical to the distribution of inter-hot spot distances. These findings support the hypothesis that the dendritic locations of Ca2+ wave hot spots in general, and branch points in particular, are specially equipped for regenerative IP3R-dependent internal Ca2+ release. Functionally, the observation that IP3R-dependent [Ca2+]i rises are greater at branch points raises the possibility that this novel Ca2+ signal may be important for the regulation of Ca2+-dependent processes in these locations. Futhermore, the observation that Ca2+ waves tend to fail between hot spots raises the possibility that influences on Ca2+ wave propagation may determine the degree of functional association between distinct Ca2+-sensitive dendritic domains