Fig. 2: Application of the decidual-ECM-mimicking 3D culture system.

A Representative images showing Ca²⁺ dynamics in IVC day 9 embryos, before and after treatment with the β-adrenergic agonist isoproterenol (1 μM). The Ca²⁺-binding dye Fluo8-AM was used to visualize calcium fluxes, highlighting changes in intracellular calcium concentrations. Scale bars, 50 μm. n = 8 independent experiments. B Graphical representation of Ca²⁺ signal changes (F/F0) in IVC day 9 embryos, captured before and after isoproterenol treatment. The traces depict a clear enhancement in calcium transients post-treatment, indicative of increased cardiac activity. C Heart rate (HR) response to isoproterenol: Quantification of the beating rate of embryos, illustrating a significant increase in HR (beats per minute, BPM) following isoproterenol treatment. Data were plotted as mean ± s.d., n = 8. P-values were calculated using unpaired, two-sided Student’s t-test, ****P < 0.0001. D Time-lapse light-field imaging of E3.5 embryo development: Sequential light-field images showing the development of an E3.5 embryo over a nine-day IVC period. Images were acquired every two hours during culture, capturing the progression of embryonic development from the blastocyst stage to more advanced stages. See also Supplementary Movie 9 for the complete time-lapse sequence. Scale bars, 500 μm. n = 3.