The dorsal habenulae (dHb) is a prominent left/right asymmetric structure in the zebrafish epithalamus. The development of such asymmetry is dependent on the temporal regulation of neurogenesis by a series of intrinsic and extrinsic factors. While dbx1b and irx7 are believed to delineate the progenitors of habenular neurons, their specific roles in the development and lateralisation of dHb remain elusive. Here, a rapid reverse genetics strategy leveraging multiplexed CRISPR/Cas9 unveiled the requirement of dbx1b for the growth and asymmetry of dHb, while irx7 potentially independently regulates habenular and visceral laterality. Phenotypes of dbx1b crispants were explored in detail. Quantifying the volumetric growth of dHb via the expression of a gng8:EGFP transgene revealed delayed and significantly reduced habenular growth in dbx1b crispants. The developmental delay may underlie the observed reduced degree of habenular asymmetry in the expression of an asymmetric differentiation marker, kctd12.1. Reduced differentiation rather than proliferation is thought to cause the delay in dbx1b crispants, as well as their habenular size reduction and sustained Cachd1-positive progenitor domain. Together, these findings suggest a dual role for dbx1b in regulating differentiation into and from dHb progenitors, possibly mediated through distinct actions of Wnt and/or Notch signalling pathways. The absence of this mechanism in dbx1b crispants may necessitate additional time to produce progenitors, resulting in the observed developmental delay and laterality defects, whilst also inhibiting the production of terminally differentiated habenular neurons. Overall, this study identifies dbx1b as a crucial player in dHb development, with potential implications for differentiation and brain asymmetry.
I would like to express my sincere gratitude to my supervisors, Dr. Gareth Powell, for his patient teaching and exceptional guidance throughout this project, and Prof. Steve Wilson, for the opportunity to conduct research in his laboratory. I also thank members and staffs of the Wilson Lab, Fish Floor Labs, and Fish Facility at UCL for providing essential support and constructive suggestions. In particular, Dr. Aaron Scott, Dr. Chintan Trivedi, Dr. Manuela Lahne, and Gema Rodriguez Valentin. Finally, although thousands of miles away, I would like to thank my parents for their support, encouragement and love.