Lrrc8aa and lrrc8ab play redundant roles in facilitating the inflation of the brain ventricle. (A) The smaller DMv phenotype in lrrc8aa-MO e2i2 or lrrc8ab-MO e2i2 morphants can be rescued by overexpression of either of the lrrc8a paralog mRNA. (B) The csad-MO i2e3 morphants show a smaller brain ventricle phenotype compared to the untreated embryo at 32 hpf. The arrows indicate the brain ventricle. Scale bars: 200 µm. (C–E) The smaller DMv phenotype in lrrc8aa-MO e2i2 or lrrc8ab-MO e2i2 morphants can be rescued by culture in embryo medium supplement with taurine. The DMv area size was increased in morphant culture with taurine (C). The images are representative median values of the DMv area of each group. Arrows indicate DMv area. Scale bars: 200 µm. The lrrc8aa-MO e2i2 (D) or lrrc8ab-MO e2i2 (E) morphants were cultured in embryo medium supplement with different amounts of taurine. The smaller DMv phenotype of lrrc8aa-MO e2i2 or lrrc8ab-MO e2i2 morphants could be rescued by supplementation of 50 mM or 6.25 mM taurine in embryo medium, respectively. (F) The lrrc8a MOs were injected into the csad transgenic embryos. The smaller DMv phenotypes were partially rescued in the csad transgenic background compared to the wild-type background. Overexpression of csad mRNA into csad transgenic embryo could fully rescued the smaller DMv phenotype. Different letters on top of the histograms represent different statistical groups (P<0.05). The lower normal boundary is shown as a red dotted line. The sample size (n) of each group is indicated on top of the histogram. cMO, control MO; aMO, lrrc8aa-MO e2i2; bMO, lrrc8ab-MO e2i2; mRNA-a, lrrc8aa-IRES-eGFP mRNA 100 pg; mRNA-a++, lrrc8aa-IRES-eGFP mRNA 200 pg; mRNA-b+, lrrc8ab-IRES-eGFP mRNA 150 pg; mRNA-b++, lrrc8ab-IRES-eGFP mRNA 200 pg; untreated embryo, UN; WT, wild-type; mRNA-Csad, Csad-IRES-eGFP mRNA 150 pg.
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