Observation of nodal cilia movement and measurement of nodal flow.

Mammalian left-right determination is a good example of how multiple cell biological processes coordinate in the formation of a basic body plan, but until recently its mechanism was totally elusive. In the past 10 years, molecular genetic studies of kinesin and dynein motor proteins, live-cell imaging techniques, and theoretical studies of fluid mechanics revealed unexpected mechanisms of left-right determination. The leftward movement of fluid at the ventral node, called nodal flow, is the central process in symmetry breaking on the left-right axis. Nodal flow is autonomously generated by the rotation of posteriorly tilted cilia that are built by transport via the KIF3 motor on cells of the ventral node. Recent evidence suggests that nodal flow transports sheathed lipidic particles, called nodal vesicular parcels (NVPs), to the left edge of the node, which results in the activation of the noncanonical Hedgehog signaling pathway, an asymmetric elevation in intracellular Ca(2+), and changes in gene expression. This chapter reviews techniques for the observation of nodal cilia movement and nodal flow in living vertebrate embryos.