High-Speed Video Microscopy of Ependymal Cilia in Brain Organotypic and Cell Culture Models

The wall of the ventricular system within the neuraxis is lined almost entirely by E1 ependymal cells, each of which projects multiple motile cilia from their apical surface into the cerebrospinal fluid (CSF). This specialized layer of E1 cells constitutes the border between the CSF and the brain interstitial fluid (BIF), and by controlling influx and efflux across the CSF to BIF interface, it is increasingly recognized to play an integral role in modulating and maintaining the brain microenvironment. The motile cilia have been shown to be responsive to changes in the CSF microenvironment, and while the physiological role of this mechanism remains incompletely understood, manipulating this control mechanism may influence the brain microenvironment potentially opening a new frontier in therapeutic intervention.In this paper, we describe our techniques for preparing organotypic slices from the murine brain parenchyma and establishing cell cultures of multiciliated ependymal cells from mouse and rat neonatal brain tissue. Our methodology generates a functional readout of ciliary function, specifically high-speed video microscopy (HSVM) enables the quantification of ciliary beat frequency (CBF), and characterization of ciliary beat pattern.