Functional imaging of mucociliary phenomena: High-speed digital reflection contrast microscopy

We present a technique for the investigation of mucociliary phenomena on trachea explants under conditions resembling those in the respiratory tract. Using an enhanced reflection contrast, we detect simultaneously the wave-like modulation of the mucus surface by the underlying ciliary activity and the transport of particles embedded in the mucus layer. Digital recordings taken at a speed of 500 frames per second are analyzed by a set of refined data processing algorithms. The simultaneously extracted data include not only ciliary beat frequency and its surface distribution, but also space-time structure of the mucociliary wave field, wave velocity and mucus transport velocity. Furthermore, we propose the analysis of the space and time evolution of the phase of the mucociliary oscillations to be the most direct way to visualize the coordination of the cilia. In particular, this analysis indicates that the synchronization is restricted to patches with varying directions of wave propagation, but the transport direction is strongly correlated with the mean direction of waves. The capabilities of the technique and of the data-processing algorithms are documented by characteristic data obtained from mammalian and avine trachea

Supplementary Material for: Ciliary Beating Plane and Wave Propagation in the Bovine Oviduct

The uterine tube is an essential conduit for the gametes and zygote during reproduction. The necessary bidirectional conveyance occurs through peristalsis and ciliary activity, but unlike in respiratory tract, little is known about mucociliary transport in the uterine tube, and the direction of transport and the alignment of oviductal cilia have not been conclusively characterized. This study aimed to determine the uniformity in the axonemal orientation of motile cilia in the bovine uterine tube, to identify the direction of mucociliary transport and to relate the presumptive beating plane and the mucociliary transport direction to the long axis of the uterine tube. The angular spread of oviductal motile cilia was determined by electron microscopy, and by maintaining the accurate alignment of the samples throughout the processing steps, axonemal orientation was determined relative to the long axis of the oviduct. The direction of the effective mucociliary transport was determined by the analysis of video microscopic data recorded on explants. Vector-based analysis of electron micrographs yielded the mean angle of deviation between the ‘effective ciliary stroke', as derived from axonemal orientation, and the tubal longitudinal axis pointing towards the uterus to be 0.8°, with a standard deviation of 35.2°. The corresponding angular deviation of the short-wave propagation was -6.8° (SD 34.6°). These results show that oviductal motile cilia are rigorously aligned, that the beating plane of the cilia is parallel to the long axis of the uterine tube and that the ‘effective stroke' and mucociliary transport are directed towards the uterus