Mouse neural tube organoids self-organize floorplate through BMP-mediated cluster competition

Neural tube organoid generation: a robust and reproducible protocol from single mouse embryonic stem cells

“…NTOs can be used to study aspects of floorplate self-organization such as reported by Krammer et al 2024 19 . This work demonstrates the formation of organizers, and it challenges the previous notion that embryonic induction relies solely on a controlling structure, such as the notochord.…”

“…-Robust epithelialization within first 3 days, -Relatively small spherical tissues (~200 μm) with a single lumen, -Clonal growth from single cells, -Serum-free, chemically defined culture medium, -Gradual neural differentiation from pluripotent cultures within 6 days, -'By-default' anterior midbrain identity by day 6, -Dorsoventrally patterned hindbrain/ cervical spinal cord-like structures within 6 days 19 .…”

“…-Antibody staining can be performed as outlined in Meinhardt et al 17 and Krammer et al 19 -Block/ permeabilization solution (1x PBS with 1% BSA and 0.5% Triton-X) is kept at 4 °C and adapted to room temperature before use.…”

“…Until now, many laboratories have developed neural organoids, mostly of mouse or human origin, that mimic different parts of the neural tube and its development, as summarized in 18 . However, to our knowledge, the protocol established by Meinhardt et al 17 is the only method where NTOs reliably form from a single cell, self-organize and efficiently pattern into dorsoventral fates following a single, global pulse of retinoic acid (RA) without the addition of other ventralizing factors such as Sonic Hedgehog Agonist (SAG) 19 , or the usage of microfluidic chambers that provide spatial information due to gradient formation. The protocol provided here was adapted from Meinhardt et al 17 as provided in Krammer et al 19 .…”

“…However, to our knowledge, the protocol established by Meinhardt et al 17 is the only method where NTOs reliably form from a single cell, self-organize and efficiently pattern into dorsoventral fates following a single, global pulse of retinoic acid (RA) without the addition of other ventralizing factors such as Sonic Hedgehog Agonist (SAG) 19 , or the usage of microfluidic chambers that provide spatial information due to gradient formation. The protocol provided here was adapted from Meinhardt et al 17 as provided in Krammer et al 19 . In this protocol, single mouse ESCs are seeded in Matrigel and exposed to defined neurodifferentiation conditions.…”