Figure 4

Trpc1 knock down reduces the expression of Trpc1 in the ectoderm and abolishes the increase in intracellular Ca²⁺ generated following activation of Ca_V1.2 channels in animal cap explants. (A–D) Embryos at the 2-cell stage were injected with (A–C) control-MO or (B–D) TRPC1-MO1 into both blastomeres, and the expression of Trpc1 was revealed by immunostaining at the blastula stage. (A,B) Confocal view of sagittal section taken through the entire embryo. (C,D) Confocal view at the level of the ectoderm. trpc1 knock-down impaired the expression of Trpc1. The nuclei (in blue) were labelled with ToPro3. Scale bars are 350 µm in (A,B) and 40 µm in (C,D). (E,F) Relative changes in fluorescence (F/F0) revealing changes in intracellular Ca²⁺ generated in single animal caps loaded with the Ca²⁺-indicator Fluo4 and isolated from embryos injected with either (E) control-MO or (F) TRPC1-MO1. The data are plotted as the mean of F/F0 (red traces) + SEM (black bars) from 7 (E) or 10 (F) randomly selected fields within a single animal cap. Noggin (2 µg/mL) was added (blue arrows) within the first 10 min after the start of data acquisition. Additional data are provided in Figure S3.