Neural blastocyst complementation enables mouse forebrain organogenesis.

Area of Science:

• Neuroscience
• Developmental Biology
• Genetics

Background:

• Traditional methods for generating genetically modified mice, like ES cell microinjection, require extensive breeding for germline transmission.
• Blastocyst complementation, previously used for immune system and organ studies, has not been adapted for brain development.
• Studying gene function in the brain is challenging due to the complexity of forebrain development.

Purpose of the Study:

• To develop a novel method, neural blastocyst complementation (NBC), for studying forebrain development and function.
• To create a system for analyzing gene functions essential for forebrain development using mouse models.
• To establish a rapid and efficient approach for generating complex forebrain organogenesis models.

Main Methods:

• Targeted ablation of host-derived dorsal telencephalic progenitors using diphtheria toxin subunit A in host embryos.
• Injection of donor embryonic stem (ES) cells into blastocysts with forebrain-specific targeting.
• Utilizing CRISPR-Cas9 to generate gene-deficient ES cells (e.g., doublecortin-deficient) for complementation studies.

Main Results:

• NBC successfully created a vacant forebrain niche, leading to agenesis of the cerebral cortex and hippocampus in host embryos.
• Donor ES cells populated the vacant niche, resulting in morphologically and neurologically normal neocortices and hippocampi.
• NBC chimeras with doublecortin-deficient ES cells accurately recapitulated the phenotype of conventional knockout mice, validating the model.

Conclusions:

• Neural blastocyst complementation (NBC) is a rapid and efficient technique for generating complex mouse models of forebrain development and function.
• NBC facilitates the study of genes required for forebrain organogenesis and neurological function.
• This blastocyst complementation strategy holds potential for broader applications in studying organogenesis.