Celsr1 suppresses WNT5A-mediated chemoattraction to prevent incorrect rostral migration of facial branchiomotor neurons
Authors:   , Hummel Devynn S. 
Published: September 2022

As the nervous system develops, neurons often migrate from where they originate to their functional locations prior to them contributing to the circuits that drive cognitive and motor function. Within the vertebrate hindbrain, facial branchiomotor (FBM) neurons migrate caudally from rhombomere 4 (r4) to r6 and assemble into the circuits that drive facial and jaw movements. While several components of the Wnt/PCP (Planar Cell Polarity) pathway have been investigated based on their necessary role in initiating FBM neuron migration, much less is known regarding the mechanisms that determine directionality. However, our lab discovered that in mice lacking the Wnt/PCP component Celsr1, many FBM neurons inappropriately migrate rostrally. Tissue-specific knockouts indicate that Celsr1 is functioning non-cell autonomously within the ventricular zone rostral to r4 in order to prevent rostral migration. Intriguingly, Celsr1 and potential chemoattractant Wnt5a are expressed in overlapping domains within the rostral hindbrain. Based on these findings, we hypothesize that under normal conditions, Celsr1 suppresses Wnt5a activity rostral to r4 to block the inappropriate rostral migration of FBM neurons. Therefore, if WNT5A functions as the primary chemoattractive cue lending to the rostral migration observed among Celsr1 mutants, then rostral migration should be suppressed in Celsr1; Wnt5a double knockouts. Indeed, FBM neurons in Celsr1; Wnt5a double knockout embryos never migrate rostrally. Furthermore, if WNT5A acts as the source of chemoattraction in the rostral hindbrain of Celsr1 knockouts, then rostral migration should observed in wildtype hindbrains with WNT5A-coated beads placed in r3. We found that FBM neurons migrated rostrally toward WNT5A beads in [less than] 80 percent of wildtype explants, demonstrating that excess WNT5A can allow for FBM neurons to overcome the endogenous effects of Celsr1. Importantly, rostral migration towards beads is significantly reduced in Dvl2 mutants, suggesting that the Wnt5a-mediated chemoattraction of FBM neurons is dependent on Wnt5a-Dvl2 signaling. Lastly, our model predicts that rostral migration of FBM neurons should be enhanced in Celsr1 mutants overexpressing Wnt5a in r3. As expected, when a Wnt5a gain-of-function allele is used to overexpress Wnt5a in r3 of Celsr1 mutants, the proportion of rostrally migrating FBM neurons is substantially greater than in control mutants, further supporting the chemoattractive model. These results reveal a novel role for a Wnt/PCP component regulating neuronal migration through suppressing chemoattraction (Chapter 3). In other studies, we used transgenic zebrafish that express various calcium indicators in order to characterize calcium dynamics and neural activity, as calcium has been shown to allow for neuron migration as well as circuit establishment spanning multiple neuronal contexts. Here we show that FBM neurons exhibit fluctuations in intracellular calcium (Cai) among both cell bodies and neurite processes during their caudal migration. We also found that beginning at 60 hpf (hours post fertilization), synchronized patterns of neuronal activity extend to most hindbrain neurons, as well as caudal-most midbrain neurons (Chapter 4). In separate studies, we used a transgenic line expressing nitroreductase in BM neurons to evaluate the ability of BM neurons to regenerate following metronidazole-mediated ablation. Surprisingly, BM neurons were not regenerated even after 1 week of recovery. Moreover, treatments shown to stimulate the regenerative response in other contexts failed to elicit BM neuron regeneration. These results suggest that different neuronal populations in zebrafish vary in their regenerative potential (Chapter 5).