Stephen C. Noctor - US grants

DESCRIPTION (provided by applicant): This project, if successful, has the prospect of fundamentally revolutionizing our understanding of the mechanisms that regulate prenatal development of the cerebral cortex. We have discovered a novel mechanism for regulating cell production that may profoundly alter our basic understanding of mechanisms that regulate neocortical development and our appreciation for how maternal immune activation may alter cortical development and lead to psychiatric disorders. Regulation of precursor cell proliferation is central for ensuring that cell production does not exceed the requirements of the developing cerebral cortex. Unrestrained cell production during prenatal brain development would have profoundly negative consequences for brain organization and function. However, mechanisms that restrain cell proliferation are poorly understood. Microglial cells colonize the cerebral cortx during prenatal development. But, despite recent progress elucidating the functions of microglia in the developing CNS and a wealth of knowledge on microglial function in the mature brain, the functional roles of microglia during prenatal cortical development are not well understood. This project simultaneously addresses gaps in our knowledge on mechanisms that regulate the number of precursor cells and the functional role of microglia in the developing cerebral cortex. Our preliminary data show that 1) microglia colonize the neurogenic proliferative zones of the prenatal rat cerebral cortex; 2) microglia phagocytose neocortical precursor cells, particularly during late stages of cortical neurogenesis; 3) maternal immune activation (MIA) in pregnant rats activates microglia in the fetal cerebral cortex; and 4) increased microglia activation is associated with a significant decrease in the number of neural precursor cells in the neocortex. Our preliminary data lead to the testable hypotheses that microglia regulate the number of neural precursor cells in the cerebral cortex through phagocytosis, and that MIA will decrease the number of neural precursor cells and alter the number of neurons and glia in the cerebral cortex. This project will test our central hypothesis, Microglia regulate the number of neural precursor cells in the developing cerebral cortex. Specific Aim 1 will quantify changes in fetal microglial activation state and precursor cell number after inducing MIA, treating with Dox, or eliminating microglia from the cortex, and quantify the number of neurons and glia in the mature cerebral cortex of animals in each group. Specific Aim 2 will test the effect of M-CSF expression on microglial colonization of the neural proliferative zones in the developing cortex, and test the effect of exogenous microglia and cytokines on the expression of 'find me' signals by neural precursor cells. Specific Aim 3 will test the capacity of pharmacological agents to attenuate the detrimental impact of MIA on the developing neocortex.

SUMMARY / ABSTRACT The outbreak of Zika virus (ZIKV) and the alarming rise in fetal brain malformations highlight ZIKV as an urgent public health concern. ZIKV has recently met Shephard's criteria for teratogenic classification because of the brain anomalies reported. This application addresses the direct relationship between ZIKV infection and fetal brain development using our established fetal primate model of intrauterine pathogenesis. The studies outlined in this application leverage our prior discoveries and collaborative investigations on neural precursor cell function in relation to microglia in the fetal primate brain, and our expertise in primate development, imaging, virology, and immunology. Our track record and experienced team provides the means to pursue the goals of this proposal?understanding the mechanism(s) of ZIKV teratogenesis?and the steps necessary to progress to studies focused on interventions. Our goal is to determine how ZIKV alters cortical development by capitalizing on our team's essential expertise and research experiences to address this urgent public health concern rapidly and effectively through the following Specific Aims: (1) Define the impact of fetal ZIKV on neural precursor cells and cortical development, and (2) Determine the impact of fetal ZIKV infection on fetal and maternal inflammation and assess if inflammation is predictive of abnormalities in cortical development. These studies will provide new insights into the underpinnings of ZIKV teratogenesis in a primate model with similar neurodevelopmental features when compared to humans, and by efficiently leveraging an existing primate model of fetal viral infection. Overall, these investigations focus on fetal developmental outcomes associated with direct ZIKV infection and will provide the necessary mechanistic understanding and outcome metrics to assess intervention strategies that protect the fetus and newborn from the devastating consequences of ZIKV infection and congenital disease.

Zika virus (ZIKV) has been identified as a neuroteratogen similar to the TORCH agents cytomegalovirus (CMV) and rubella. Neurotropic teratogenic viruses such as CMV cause a wide range of sequelae in the infected human fetus and infant, yet we have little definitive information about the range of outcomes related to congenital Zika syndrome. These questions are difficult to study, largely due to the rapid emergence of Zika virus and our incomplete understanding of the virus's natural history and mechanisms of pathogenesis. However, our previous research on congenital CMV in the rhesus monkey model offers a precedent and template for improved appreciation of Zika virus pathogenic outcomes, and for the rational design of diagnostic, preventative, and interventional strategies for this rapidly spreading global teratogen. The studies in this proposal capitalize on our considerable expertise with the rhesus CMV (RhCMV) in utero infection model, which defined the pathology associated with disease and has been used to investigate the safety of new vaccines and protective interventions. It has become increasingly apparent that environmental factors such as infectious teratogens and inflammation together exert a persistent or even lifelong impact on neurodevelopment and immune function. The objectives of these studies include the following Specific Aims: (1) Determine the fetal immunologic effects of placental ZIKV and test if immune activation is associated with the extent of central nervous system (CNS) infection, neural precursor cell loss, and brain malformations. In these investigations we will test the hypothesis that Zika virus infection results in maternal cell ingress, alteration of decidual cytokine production, increased microchimerism, and fetal/neonatal systemic inflammation. (2) Evaluate the impact of neonatal ZIKV infection on immune activation, immune ontogeny, and neurodevelopment. In these studies we will test the hypothesis that in neonates, Zika virus infection has a significant correlated influence on immune function and neurodevelopment. Overall, these investigations will provide new insights into the developmental perturbations resulting from Zika virus infection, and will leverage our experiences in developing outcome metrics and interventions that can protect the fetus and infant from the devastating consequences of congenital disease.

PROJECT SUMMARY / ABSTRACT Microglial cells are resident innate immune cells in the brain and exhibit dynamic changes in morphology and distribution during gestation. They can regulate diverse programs that are essential for normal brain development and function, and recent studies have shown that in the developing brain microglia regulate key processes including synapse development, axonal path finding, and cortical layer formation. We have shown that microglia regulate cell production through phagocytosis of neural precursor cells. However, it is currently not known how microglial cell lineage(s) correlate with colonization of the brain or the cellular function of microglial cells in the developing brain, particularly in relation to neural precursor cells. The studies proposed in this application are designed to address significant gaps in our knowledge on the trafficking and seeding of microglial cell populations during critical windows of nonhuman primate brain development. These studies will generate new insights into normal development that are critical to our understanding of ontogeny and will directly address the goals of PAS-18-483 in a translational nonhuman primate model through the following Specific Aims: (1) Evaluate the role of yolk sac precursor cells in contributing to the microglial cells that populate the fetal rhesus monkey cerebral cortex; (2) Determine the role of clonal lineage(s) in regional colonization of the cerebral cortex and function of cortical microglia; and (3) Define temporal dynamics that impact the distribution and function of clonal microglia. Together the proposed studies will address fundamental gaps in our understanding of factors that regulate colonization of the cerebral cortex by microglial cells, and the impact microglial cell function has on the developing brain.

PROJECT SUMMARY / ABSTRACT Microglial cells are the resident innate immune cell in the brain and exhibit a range of distinct morphologies and distribution during gestation. Studies in the past decade have shown that microglia regulate key processes in the developing brain including synapse development, axonal path finding, and cortical layer formation. We recently showed that microglia regulate cell production in the prenatal cerebral cortex through phagocytosis of neural precursor cells (NPCs), but the full range of functions performed by fetal microglial cells are not well understood. We have discovered a novel population of periventricular microglia that make extensive contact with NPCs undergoing division at the ventricular lumen in the fetal rhesus monkey telencephalon. The studies proposed in this application are designed to address significant gaps in our knowledge on the function of fetal microglial cells during critical windows of nonhuman primate brain development. This proposal will generate new insights into normal development that are critical to our understanding of ontogeny through the following Specific Aims: (1) Define the role of fetal periventricular microglia in NPC detachment through live time-lapse videomicroscopy in cultured explants; and (2) Determine the role of purinergic signaling in regulating periventricular microglia contact with NPCs and NPC detachment in the fetal monkey telencephalon. The proposed studies will enhance our understanding of factors that regulate development of the cerebral cortex and further define the impact microglial cell function has on the developing fetal brain.