Microfluidic Control of Dorsal-Ventral Patterning Within a Single Forebrain Organoid.

Area of Science:

• Neuroscience
• Developmental Biology
• Bioengineering

Background:

• Understanding brain regionalization is crucial for developmental neuroscience.
• Current in vitro models using fused organoids introduce variability, hindering regionalization studies.
• A need exists for precise methods to control tissue identity development in vitro.

Purpose of the Study:

• To develop a microfluidic platform for co-developing distinct tissue identities within a single 3D culture.
• To investigate the role of controlled morphogen delivery in driving spatial regionalization.
• To offer a scalable and practical alternative to organoid fusion for studying brain development.

Main Methods:

• A microfluidic platform enabling stable perfusion and real-time imaging was designed.
• Controlled delivery of Sonic hedgehog pathway agonist (SAG) was applied to mouse forebrain organoids.
• Spatially segregated molecular transport and tissue development were tracked using fluorescence imaging.

Main Results:

• The platform supported co-development of distinct tissue identities within a unified 3D culture domain.
• Spatiotemporal delivery of SAG induced segregated ventral (Nkx2.1+) and dorsal (Pax6+) domains.
• Controlled morphogen delivery drove region-specific fate specification without organoid fusion.

Conclusions:

• The microfluidic platform provides a novel method for studying tissue regionalization in vitro.
• Controlled morphogen gradients are sufficient to pattern distinct brain regions within a single organoid.
• This approach offers a scalable and reproducible alternative for modeling early brain development.