Local synchronization of cilia and tissue-scale cilia alignment are sufficient for global metachronal waves

Abstract

Motile cilia are hair-like cell extensions present in multiple organs of the body. How cilia coordinate their regular beat in multiciliated epithelia to efficiently displace fluids remains elusive. Here, we propose the zebrafish nose as an accessible model system to study ciliary dynamics, due to its conserved properties with other ciliated tissues and its high availability for non-invasive imaging. We reveal that cilia are locally synchronized, and that the size of local synchronization domains increases with the viscosity of the surrounding medium. Despite this merely local synchronization, we observe global patterns of traveling metachronal waves across the multiciliated epithelium. Intriguingly, these global wave direction patterns are conserved across individual fish, but different for left and right nose, revealing a chiral asymmetry of metachronal coordination. In conclusion, we show that local synchronization together with tissue-scale cilia alignment shape global wave patterns in multiciliated epithelia.

Measurement of ciliary beating and fluid flow in the zebrafish adult telencephalon

“…To analyze ciliary beating frequency (CBF), we suggest two methods using a MATLAB script or an ImageJ/Fiji plugin (FreQ). The experimenters can choose their method of choice because both methods adapt a similar Fourier-based principle to analyze CBF ( Figure 4 A) ( Haward et al., 2021 ; Reiten et al., 2017 ; Ringers et al., 2021 ). However, we recommend the experimenters to always use the same method for their analysis, since there are slight differences in their output results, as indicated in the limitations section and in Figure 10 .…”

“…Hence it is crucial to make sure that there are no drifts in x and y axis prior to the analysis. If you observe drift in x,y, you may try to correct those with an alignment code prior to the CBF analysis described earlier ( Reiten et al., 2017 ; Ringers et al., 2021 ). However, it is important that you check manually, frame by frame that, after correction, the sample is no longer drifting in the image.…”

“…This protocol provides the methods to measure ciliary beating and CSF flow in the telencephalon of an adult zebrafish brain explant. Combining light transmission microscope and fast Fourier transform-based analysis ( D'Gama et al., 2021 ; Olstad et al., 2019 ; Reiten et al., 2017 ; Ringers et al., 2021 ) allows experimenters to examine the extent and frequency of ciliary beating. We also describe how to measure CSF flow upon injection of fluorescent particles into the brain ventricle and tracking via fluorescent imaging.…”

“…This protocol is also applicable to other ciliated tissues. For instance, we used identical analytical tools to measure ciliary beating of multiciliated cells in the zebrafish nose ( Reiten et al., 2017 ; Ringers et al., 2021 ), mouse tracheal epithelial cultures ( Haward et al., 2021 ), and acute slices of the murine lung (unpublished data). However, it may be difficult to apply this approach for the analysis of monociliated tissues because it is hard to detect ciliary beating of mono cilia by a light transmission microscope.…”

Computational Modeling of Motile Cilia-Driven Cerebrospinal Flow in the Brain Ventricles of Zebrafish Embryo

“…It is also reported that the increasing cilia length provides an increased directional flow rate, and a smaller cilia length with a random beating pattern results in increased flow mixing [ 54 ]. Besides, the way ciliary beating is synchronized and how synchronization modulates fluid pumping remain poorly understood [ 55 ]. Notably, metachronal waves, which refer to cilia beating with a time delay between neighbors [ 1 , 56 ], have been commonly described for multi-ciliated cells and are suggested to improve fluid pumping [ 55 , 57 , 58 , 59 , 60 , 61 ].…”