Origins and Migration of the Octavolateral Efferent and Facial Branchiomotor Neurons in Zebrafish

The vertebrate hindbrain is a segmented structure that displays broad morphological and molecular conservation across vertebrate species. Efferent neurons within the hindbrain project to peripheral targets such as muscles and sensory organs, providing essential control over functions of the vertebrate head. Their axons project often along highly stereotyped routes, exiting the brainstem as the cranial nerves. This thesis will focus on the development and migration of two types of cranial efferent neurons in zebrash: the facial branchiomotor neurons (FBMNs) of the VIIth (facial) nerve and the octavolateral efferent neurons (OENs) of the VIIIth (vestibuloacoustic) nerve. Very little is known about the OENs, in part due to the lack of molecular markers which distinguish them from the better-characterized FBMNs. Two clusters of OENs have been identied in the hindbrain, consisting of the rostral efferent neurons (RENs) in r6 and the caudal efferent neurons (CENs) in r7. Both RENs and CENs migrate tangentially in a rostrocaudal manner along the same route as the FBMNs before clustering in r6 and r7. However, the segmental origins of the OENs, timing of their migration and axon outgrowth, and their interactions with the FBMNs have not been characterized. Here, I employ high-resolution imaging techniques to investigate early stages of OEN development. In Chapter 2, I use single-plane illumination microscopy (SPIM) to backtrack individual neurons to their birthplaces over 12 hours of developmental time. I demonstrate that OENs are born in the ventral neuroepithelium, close to the medial floor plate. RENs are born with the FBMNs in r4 between 11 and 16 hpf, but CENs have a more posterior origin within r5 and their births occur later, between 15 and 16 hpf. Both FBMNs and OENs generally remain ipsilateral to their mothers, while their sisters integrate contralaterally into the neuroepithelium. In Chapter 3, I use a new photoconvertible transgenic line to characterize the migration of the OENs and investigate their interactions with FBMNs. I find that OENs migrate alongside the FBMNs between 18 and 48 hpf. RENs rely on proper function of hoxb1a to migrate caudally out of r4 and to innervate the otic vesicle, while CEN identity is independent of hoxb1a. Neither RENs nor CENs rely on the planar cell polarity molecule, Pk1b, to migrate, unlike the FBMNs which remain in r4 in its absence. In a series of cell ablation experiments, I also investigate interactions between the FBMNs, RENs, and CENs during their migration, determining that although FBMN/RENs make contact with CENs across the r4/5 border, CENs do not lead migration through r5. These results uncover important differences between OEN and FBMN migration and suggest that the origins of OENs in the zebrash are more complex than previously assumed. Taken together, my findings offer a better understanding of this rarely studied cell type, and raise important questions about its relationship to other cranial efferent populations in the vertebrate brainstem