We propose to investigate how signals in the amniotic fluid (AF) and nascent cerebrospinal fluid (CSF) instruct neural stem cell behavior during early brain developmental stages surrounding the time of neural tube closure (E8.5-E10.5). The rapidly changing and growing population of neural stems at these early stages will give rise to all neurons and glia in the adult brain, yet comparatively few studies exist on the intrinsic genetc programs or the extrinsic fluid-based signals involved in driving these early stages of development, largely due to technical limitations. We and others have demonstrated broad influences of CSF and vascular fluid niches in instructing later stages of brain development. Yet virtually nothing is known about the protein composition of AF and early CSF during early brain development, at a time when the choroid plexus and vasculature have yet to form. Our overarching hypothesis is that secreted signals in the AF and nascent CSF are critical in instructing and synchronizing the proliferation and fate of embryonic E8.5-E10.5 neural stem cells bathed in these fluids. We will first test this hypothesis by comparing the effects of AF, early CSF and buffered media on stem cell explants at ages E8.5 and E10.5. We predict that, as in later development, early explants only develop normally when bathed in the age-appropriate fluid, suggesting that changes in the AF/CSF proteome are mirrored by changes in expression of associated receptors at the apical surface of neural stem cells. As such, we expect that normal stem cell function should be impaired by genetic perturbations to the cilia and membrane of neural stems cells at their apical surface, which is in direct contact with the AF/CSF. A near- complete list of CSF proteins and their associated receptors on apical membranes will then be deduced using quantitative mass-spec and RNAseq technologies, together with immunostaining. We will then determine the contributions of CSF-LIF and other top candidate proteins in instructing specific aspects of E10.5 stem cell behavior, and the embryonic sources of these CSF signals. This proposal has important clinical significance: We currently have relatively little understanding of the origins of early developmental disorders (defects due to errant neural tube closure, hydrocephalus, and infiltration by teratogens), and scant capacities for early diagnosis or intervention. This proposal should provide a foundation for asking how these early perturbations ultimately derail some or all aspects of normal brain development, and for developing minimally invasive AF/CSF sampling and replacement strategies for disease diagnosis and for reprogramming of neural stem cells in order to bring the development brain back on track.
We propose to investigate how signals in the amniotic fluid and early cerebrospinal fluid instruct neural stem cell behavior during pivotal but understudied early stages of brain development. The proposed research has important clinical implications for the understanding and diagnosis of common disorders of early development, such as neural tube closure defects, congenital hydrocephalus. This work will also provide clues as to how harmful teratogens gain access to the brain during early development. Our approach should also guide neural stem cell reprogramming strategies using minimally invasive AF/CSF replacement for nervous system repair and maintenance.
|Mathew, Rebecca S; Mullan, Hillary; Blusztajn, Jan Krzysztof et al. (2016) Comment on ""Multiple repressive mechanisms in the hippocampus during memory formation"". Science 353:453|
|Chang, Jessica T; Lehtinen, Maria K; Sive, Hazel (2016) Zebrafish cerebrospinal fluid mediates cell survival through a retinoid signaling pathway. Dev Neurobiol 76:75-92|
|Chau, Kevin F; Springel, Mark W; Broadbelt, Kevin G et al. (2015) Progressive Differentiation and Instructive Capacities of Amniotic Fluid and Cerebrospinal Fluid Proteomes following Neural Tube Closure. Dev Cell 35:789-802|
|Lun, Melody P; Monuki, Edwin S; Lehtinen, Maria K (2015) Development and functions of the choroid plexus-cerebrospinal fluid system. Nat Rev Neurosci 16:445-57|
|Lun, Melody P; Johnson, Matthew B; Broadbelt, Kevin G et al. (2015) Spatially heterogeneous choroid plexus transcriptomes encode positional identity and contribute to regional CSF production. J Neurosci 35:4903-16|