Endothelial Dab1 signaling orchestrates neuro-glia-vessel communication in the central nervous system

Developing the bloodbrain barrier During development, signals need to be dynamically integrated by endothelial cells, neurons, and glia to achieve functional neuro-glia-vascular units in the central nervous system. During cortical development, neuronal Dab1 and ApoER2 receptors respond to a guidance cue called reelin. Studying mice, Segarra et al. found that Dab1 and ApoER2 are also expressed in endothelial cells (see the Perspective by Thomas). The integration of reelin signaling in endothelial cells and neurons facilitates the communication between vessels, glia, and neurons that is necessary for the correct positioning of neurons during cortical development. This integration is also important for correct communication at the neurovascular unit required for blood-brain barrier integrity in the mature brain. Science, this issue p. eaao2861; see also p. 754 A single molecule, reelin, can orchestrate both neuronal migration and the vascularization of the central nervous system. INTRODUCTION The function of the brain relies on communication among the complex network of cells that constitute this organ. Vascularization of the central nervous system (CNS) ensures adequate delivery of oxygen and nutrients to build up and maintain homeostasis of neuronal networks. Thus, it is not surprising that blood vessels and neuronal cells share multiple parallelisms orchestrating their development in synchrony and in a mutually dependent manner in the CNS. Despite the essential role of the endothelium in brain function, the means by which signaling at the interface of endothelial cells, glial cells, and neurons is integrated temporally and spatially for proper brain development has remained largely unexplored. RATIONALE Integration of signaling pathways and cellular responses among endothelial cells, glial cells, and neurons is needed to ensure proper architecture of the brain. Reelin (Reln), a large secreted glycoprotein, induces Disabled 1 (Dab1)–dependent responses in neurons to guide their migration in all layered brain structures. Secretion of reelin by Cajal-Retzius cells in the marginal zone of the cortex timely coincides with active sprouting of pial vessels ingrowing perpendicularly into the marginal zone and forming a complex vascular network needed to support brain development and function. Therefore, reelin might be in the perfect position to perform a bivalent function to timely and spatially orchestrate both neuronal migration and CNS vascularization. We reasoned that blood vessels might instruct the process of neuronal migration by a cell-autonomous function of Dab1 on endothelial cells. To investigate this, we deleted the expression of vascular Dab1 in mice and investigated the effects on CNS vascularization, neuroglial organization, and neurovascular unit function. RESULTS We found that reelin/Dab1 signaling is conserved in endothelial cells and exerts potent proangiogenic effects in the developing vasculature of the CNS by controlling endothelial cell proliferation and active filopodia extension of the vascular network. The interaction of the reelin receptor ApoER2 (apolipoprotein E receptor 2) and VEGFR2 (vascular endothelial growth factor receptor 2) mediated the proangiogenic roles of Dab1 in endothelial cells. Surprisingly, deletion of Dab1 exclusively in the vascular system induced changes in the position of postmitotic pyramidal neurons in the cortical layers of the cerebral cortex. At the cellular level, depletion of vascular Dab1 reduced the docking of the radial glia processes to the pial surface at embryonic and postnatal stages and altered the differentiation of glial cells to astrocytes. The defects in neuronal migration persisted in adult mutant animals, where stereotypical attachment of the astrocytes to penetrating vessels in the glia limitans superficialis was also found to be aberrant. The functionality of the neurovascular unit [blood-brain barrier (BBB) integrity] was also affected in reelin knockout animals, and we could attribute those defects to the lack of Dab1 signaling exclusively in endothelial cells. The increased BBB permeability was again associated with an insufficient coverage of the brain vasculature by astrocytic endfeet. Mechanistically, we determined that the astroglial attachment to the vasculature is mediated by reelin-induced deposition of laminin-α4 by endothelial cells to the extracellular compartment, which in turn enables the binding of the glial processes to the CNS vasculature via the activation of integrin-β1 in glial cells. CONCLUSION Our results shed new light on the function of the vasculature in CNS development and homeostasis—in particular, how signals from the endothelium orchestrate the communication among vessels, glial cells, and neurons, and how specific changes in the molecular signature of the endothelium affect a plethora of processes such as CNS vascularization, extracellular matrix composition, neuroglial cytoarchitecture, and BBB development. Instructive functions of vascular Dab1 in the neurovascular interface. (A) ApoER2 and Dab1 control endothelial cell proliferation and tip cell filopodia extension during CNS vascularization by cross-talking to the VEGFR2 pathway. (B and C) Vascular Dab1 also instructs radial glia organization and neuronal migration in the developing cerebral cortex (B) and the development of the blood-brain barrier (C). In both cases, Dab1 signaling in the vasculature regulates the deposition of laminin-α4 and, in turn, the activation of integrin-β1 in glial cells. The architecture of the neurovascular unit (NVU) is controlled by the communication of neurons, glia, and vascular cells. We found that the neuronal guidance cue reelin possesses proangiogenic activities that ensure the communication of endothelial cells (ECs) with the glia to control neuronal migration and the establishment of the blood-brain barrier in the mouse brain. Apolipoprotein E receptor 2 (ApoER2) and Disabled1 (Dab1) expressed in ECs are required for vascularization of the retina and the cerebral cortex. Deletion of Dab1 in ECs leads to a reduced secretion of laminin-α4 and decreased activation of integrin-β1 in glial cells, which in turn control neuronal migration and barrier properties of the NVU. Thus, reelin signaling in the endothelium is an instructive and integrative cue essential for neuro-glia-vascular communication.