Oligodendrocyte precursor cells (OPCs) in the adult brain contribute to white matter homeostasis. After white matter damage, OPCs compensate for oligodendrocyte loss by differentiating into mature oligodendrocytes. However, the underlying mechanisms remain to be fully defined. Here, we test the hypothesis that, during endogenous recovery from white matter ischemic injury, astrocytes support the maturation of OPCs by secreting brain-derived neurotrophic factor (BDNF). For in vitro experiments, cultured primary OPCs and astrocytes were prepared from postnatal day 2 rat cortex. When OPCs were subjected to chemical hypoxic stress by exposing them to sublethal CoCl 2 for 7 d, in vitro OPC differentiation into oligodendrocytes was significantly suppressed. Conditioned medium from astrocytes (astro-medium) restored the process of OPC maturation even under the stressed conditions. When astro-medium was filtered with TrkB-Fc to remove BDNF, the BDNF-deficient astro-medium no longer supported OPC maturation. For in vivo experiments, we analyzed a transgenic mouse line (GFAP cre /BDNF wt/fl ) in which BDNF expression is downregulated specifically in GFAP ϩ astrocytes. Both wild-type (GFAP wt /BDNF wt/fl mice) and transgenic mice were subjected to prolonged cerebral hypoperfusion by bilateral common carotid artery stenosis. As expected, compared with wild-type mice, the transgenic mice exhibited a lower number of newly generated oligodendrocytes and larger white matter damage. Together, these findings demonstrate that, during endogenous recovery from white matter damage, astrocytes may promote oligodendrogenesis by secreting BDNF.

Cav2.1 channels, which conduct P͞Q-type Ca 2؉ currents, were expressed in superior cervical ganglion neurons in cell culture, and neurotransmission initiated by these exogenously expressed Ca 2؉ channels was measured. Deletions in the synaptic protein interaction (synprint) site in the intracellular loop between domains II and III of Cav2.1 channels reduced their effectiveness in synaptic transmission. Surprisingly, this effect was correlated with loss of presynaptic localization of the exogenously expressed channels. Cav1.2 channels, which conduct L-type Ca 2؉ currents, are ineffective in supporting synaptic transmission, but substitution of the synprint site from Cav2.1 channels in Cav1.2 was sufficient to establish synaptic transmission initiated by L-type Ca 2؉ currents through the exogenous Cav1.2 channels. Substitution of the synprint site from Cav2.2 channels, which conduct N-type Ca 2؉ currents, was even more effective than Cav2.1. Our results show that localization and function of exogenous Ca 2؉ channels in nerve terminals of superior cervical ganglion neurons require a functional synprint site and suggest that binding of soluble NSF attachment protein receptor (SNARE) proteins to the synprint site is a necessary permissive event for nerve terminal localization of presynaptic Ca 2؉ channels. E lectrophysiological and pharmacological studies have defined a diverse array of native Ca 2ϩ currents having different functions in neurons (1, 2). Voltage-gated Ca 2ϩ channels are complexes of a pore-forming ␣ 1 subunit with associated ␣ 2 ␦, ␤, and ␥ subunits (3-5). Ca v 2.1 channels that conduct P͞Q-type Ca 2ϩ currents and Ca v 2.2 channels that conduct N-type Ca 2ϩ currents are the primary initiators of fast synaptic transmission in vertebrate neurons (1, 6-12). These Ca 2ϩ channels bind directly to soluble NSF attachment protein (SNAP) receptor (SNARE) proteins involved in neurotransmitter release through a synaptic protein interaction (synprint) site in the large intracellular loop connecting domains II and III (L II-III ) of their ␣ 1 subunits (13-15). Disruption of this interaction by peptide inhibitors injected into presynaptic neurons reduces the efficiency of Ca 2ϩ entry in stimulating exocytosis (16,17). These results implicate the interaction of SNARE proteins with the synprint site in determining the efficiency of fast synaptic transmission, possibly by organizing docked synaptic vesicles close to the site of Ca 2ϩ entry.The molecular basis for the specific role of Ca v 2 channels in initiation of fast neurotransmission is not well understood. It may involve specific localization of Ca v 2 channels in nerve terminals, specific interactions with SNARE proteins or other proteins in the nerve terminal, or both. Results presented in the accompanying paper (18) show that exogenous Ca v 2.1 channels can be functionally expressed in superior cervical ganglion neurons (SCGNs) and can reconstitute synaptic transmission in neurons whose endogenous Ca v 2.2 channels have been blocked by -conotoxin GVIA. Here w...

Background and Purpose Pentraxin 3 (PTX3) is released upon inflammatory responses in many organs. However, roles of PTX3 in brain are still mostly unknown. Here we asked whether and how PTX3 contributes to blood-brain barrier (BBB) dysfunction during the acute phase of ischemic stroke. Methods In vivo, spontaneously hypertensive rats were subjected to focal cerebral ischemia by transient middle cerebral artery occlusion. At day 3, brains were analyzed to evaluate the cellular origin of PTX3 expression. Correlations with BBB breakdown were assessed by IgG staining. In vitro, rat primary astrocytes and rat brain endothelial RBE.4 cells were cultured to study the role of astrocyte-derived PTX3 on VEGF-mediated endothelial permeability. Results During the acute phase of stroke, reactive astrocytes in the peri-infarct area expressed PTX3. There was negative correlation between gradients of IgG leakage and PTX3-positive astrocytes. Cell culture experiments showed that astrocyte-conditioned media increased levels of tight junction proteins and reduced endothelial permeability under normal conditions. Removing PTX3 from astrocyte-conditioned media by immunoprecipitation increased endothelial permeability. PTX3 strongly bound VEGF in vitro and was able to decrease VEGF-induced endothelial permeability. Conclusions Astrocytes in peri-infarct areas upregulate PTX3 which may support BBB integrity by regulating VEGF-related mechanisms. This response in astrocytes may comprise a compensatory mechanism for maintaining BBB function after ischemic stroke.