Requirement for the zebrafish mid-hindbrain boundary in midbrain polarisation, mapping and confinement of the retinotectal projection

The organizer at the midbrain-hindbrain boundary (MHB organizer) has been p roposed to induce and polarize the midbrain during development, We investig ate the requirement for the MHB organizer in acerebellar mutants, which lac k a MHB and cerebellum, but retain a tectum, and are mutant for fgf8, a can didate inducer and polarizer. We examine the retinotectal projection in the mutants to assay polarity in the tectum, In mutant tecta, retinal ganglion cell (RGC) axons form overlapping termination fields, especially in the ve ntral tectum, and along both the anterior-posterior and dorsal-ventral axis of the tectum, consistent with a MHB requirement in generating midbrain po larity. However, polarity is not completely lost in the mutant tecta, in sp ite of the absence of the MHB, Moreover, graded expression of the ephrin fa mily ligand Ephrin-A5b is eliminated, whereas Ephrin-A2 and Ephrin-A5a expr ession is leveled in acerebellar mutant tecta, showing that ephrins are dif ferentially affected by the absence of the MHB, Some RGC axons overshoot be yond the mutant tectum, suggesting that the MHB also serves a barrier funct ion for axonal growth, By transplanting whole eye primordia, we show that m apping defects and overshooting largely, but not exclusively, depend on tec tal, but not retinal genotype, and thus demonstrate an independent function for Fgf8 in retinal development. The MHB organizer, possibly via Fgf8 itse lf, is thus required for midbrain polarisation and for restricting axonal g rowth, but other cell populations may also influence midbrain polarity.