Evolutionarily conserved roles of foxg1a in the developing subpallium of zebrafish embryos.

The vertebrate telencephalic lobes consist of the pallium (dorsal) and subpallium (ventral). The subpallium gives rise to the basal ganglia, encompassing the pallidum and striatum. The development of this region is believed to depend on Foxg1/Foxg1a functions in both mice and zebrafish. This study aims to elucidate the genetic regulatory network controlled by foxg1a in subpallium development using zebrafish as a model. The expression gradient of foxg1a within the developing telencephalon was examined semi-quantitatively in initial investigations. Utilizing the CRISPR/Cas9 technique, we subsequently established a foxg1a mutant line and observed the resultant phenotypes. Morphological assessment revealed that foxg1a mutants exhibit a thin telencephalon together with a misshapen preoptic area (POA). Notably, accumulation of apoptotic cells was identified in this region. In mutants at 24 h postfertilization, the expression of pallium markers expanded ventrally, while that of subpallium markers was markedly suppressed. Concurrently, the expression of fgf8a, vax2, and six3b was shifted ventrally, causing anomalous expression in regions typical of POA formation in wild-type embryos. Consequently, the foxg1a mutation led to expansion of the pallium and disrupted the subpallium and POA. This highlights a pivotal role of foxg1a in directing the dorsoventral patterning of the telencephalon, particularly in subpallium differentiation, mirroring observations in mice. Additionally, reduced expression of neural progenitor maintenance genes was detected in mutants, suggesting the necessity of foxg1a in preserving neural progenitors. Collectively, these findings underscore evolutionarily conserved functions of foxg1 in the development of the subpallium in vertebrate embryos.

Involvement of nr2f genes in brain regionalization and eye development during early zebrafish development.

Nuclear receptor subfamily 2 group F (Nr2f) proteins are essential for brain development in mice, but little is known about their precise roles and their evolutionary diversification. In the present study, the expression patterns of major nr2f genes (nr2f1a, nr2f1b, and nr2f2) during early brain development were investigated in zebrafish. Comparisons of their expression patterns revealed similar but temporally and spatially distinct patterns after early somite stages in the brain. Frameshift mutations in the three nr2f genes, achieved using the CRISPR/Cas9 method, resulted in a smaller telencephalon and smaller eyes in the nr2f1a mutants; milder forms of those defects were present in the nr2f1b and nr2f2 mutants. Acridine orange staining revealed enhanced cell death in the brain and/or eyes in all nr2f homozygous mutants. The expression of regional markers in the brain did not suggest global defects in brain regionalization; however, shha expression in the preoptic area and hypothalamus, as well as fgf8a expression in the anterior telencephalon, was disturbed in nr2f1a and nr2f1b mutants, potentially leading to a defective telencephalon. Specification of the retina and optic stalk was also significantly affected. The overexpression of nr2f1b by injection of mRNA disrupted the anterior brain at a high dose, and the expression of pax6a in the eyes and fgf8a in the telencephalon at a low dose. The results of these loss- and gain-of-function approaches showed that nr2f genes regulate the development of the telencephalon and eyes in zebrafish embryos.