Secretion of Reelin by blood vessels affects the formation of the cerebral cortex

The developing forebrain is surrounded by blood vessels before the first neurons are generated. Vascular networks subsequently invade the embryonic forebrain as newly born neurons migrate to form the six layers that will comprise the adult cerebral cortex. While it was previously believed that blood vessels only have a supportive role during development, it is now widely accepted that they provide instructive signals to regulate the generation and migration of cortical neurons. More recently, defective blood vessel development has been suggested to contribute to cortical pathologies of neurodevelopmental origin. We have found that blood vessels secrete Reelin, a protein which has a crucial role in the formation and connectivity of the rodent and human cerebral cortex, and which has also been linked to neurodevelopmental disorders in humans. While Reelin has been detected in human and rodent blood plasma, little is known about its significance and function in the formation of the cerebral cortex. Here, we propose to use genetic strategies in mice to remove Reelin specifically from blood vessels and analyse how the formation of the cerebral cortex is affected. Further, we will use mouse models in a strategy aimed to shed light on the molecular mechanisms by which blood vessels secrete Reelin and communicate with neurons in the developing brain. Thus, the proposed research programme will explore novel mechanisms that underlie the secretion of a key molecule, Reelin, by blood vessels and the communication between vasculature and neurons during the formation of the cerebral cortex. This work is likely to enhance our understanding of the aetiologies of developmental disorders of this area of the brain such as schizophrenia and autism.

Reelin is a secreted extracellular matrix protein which is known to play a crucial role in neuronal migration, layer formation and connectivity in the developing cerebral cortex. Reeler mouse mutants show severe defects in cortical lamination and synapse formation which underlie their marked motor deficits. While it was previously thought that Cajal-Retzius cells and some interneurons are the sole sources of this protein in the developing cortex, a recent study has suggested that cortical blood vessels express Reelin and its receptors. This is consistent with the detection of Reelin in rodent and human blood serum and with the finding of elevated levels in the plasma of patients with neuropsychiatric disorders. However, the function of circulating Reelin in the formation of the cortex remains unknown. Prompted by preliminary data which showed secretion by cortical endothelial cells, we propose to investigate the function of vascular-Reelin in corticogenesis by using a genetic ablation strategy to specifically remove this protein from endothelial cells in vivo. Moreover, we have recently found that blood vessels are enriched in members of the family of SNARE proteins, including Snap25 which mediates regulated exocytosis in neurons. We propose to use a genetic conditional Cre-Lox strategy to test whether Snap25 mediates vascular-exocytosis and whether Reelin is released by this process. Other candidate SNARE members which are enriched in blood vessels and which may underlie Reelin secretion will be identified by RNA sequencing experiments, and candidates tested by knocking down specific SNARE members in vascular endothelial cells in vitro using siRNA. These experiments will be combined with biochemical studies with the aim to identify the molecular partners underlying Reelin secretion and, more generally, neurovascular coupling. Thus, the proposed studies will elucidate how blood vessels communicate with neurons to regulate the formation of the cerebral cortex.

Firstly, success of this application will have a positive impact on our laboratory which has provided a fertile ground for training of students, postdoctoral fellows and visiting scientists from around the world for the past three decades. Most of them have since gone to become independent and productive scientists. This grant, if successful, will allow us to investigate the novel hypothesis that aberrant secretion of Reelin from blood vessels in the developing brain affects its formation and may contribute to the aetiology of neurodevelopmental disorders. The wide range of techniques included in the project comprise highly transferable skills for people in the laboratory to use in other projects here or other institutions in the future.

The proposed programme of research aims to investigate the complex mechanisms by which blood vessels communicate with neurons in the brain through the secretion of a repertoire of molecules including Reelin. This line of work complements our present and past interests, and those of other groups worldwide, on the cell and molecular mechanisms involved in the formation of the brain, especially of the cerebral cortex. Although I expect the short-term benefit of this research to be academic, a long-term positive effect on public health and economy is likely as it will increase the understanding of mechanisms that may underlie neuropsychiatric disorders of developmental origin, especially of schizophrenia. Understanding such mechanisms may contribute to therapeutic approaches for these disorders. In addition to disseminating our experimental findings to the basic science and medical communities, I will also strive to include our research as part of my undergraduate and postgraduate teaching.