The BMP antagonist Gremlin1 contributes to the development of cortical excitatory neurons, motor balance and fear responses
Introduction: Bone morphogenetic protein (BMP) signaling is required for early forebrain development, however the role of this signalling pathway in later cortical patterning awaits careful mechanistic investigation. How the endogenous modulators of BMP signaling regulate the structural and functional maturation of the developing brain remains unclear.  Aim: To investigate the expression and function of the BMP antagonist Gremlin1 (Grem1) in the developing mouse brain.  Methods: The role of Grem1 in neuronal differentiation was assessed using neural stem cell (NSC) and progenitor cell culture ex vivo. Lineage tracing of Grem1 expressing cells in the fetal brain was examined by administration of tamoxifen to pregnant Grem1creERT; Rosa26LSLTdtomato mice at 13.5 days post coitum (dpc), followed by collection of embryos later in gestation. In addition, at 14.5 dpc cells from the dorsal telencephalon were FACS sorted into Grem1 positive and negative cells and bulk mRNA seq analysis of differentially expressed transcripts between the two cell populations was performed. We also generated Emx1-cre mediated Grem1 conditional knockout mice (Emx1-Cre;Grem1flox/flox, Grem1Emx1 cKO, in which the Grem1 gene was deleted specifically in the dorsal telencephalon and assessed brain histology and mouse behavior.   Results: Grem1 positive cells were located in the lower cortical plate and subplate at E14.5 and migrated towards the lateral ventricle. Grem1 cells expressed immature neuron markers, self-renewed and gave rise to layer ? and ? excitatory glutamatergic neurons. GREM1 expression also identifies a similar population of excitatory neurons in the developing human brain. Recombinant Grem1 treatment induced neural differentiation of NSCs in vitro, whilst Grem1Emx1 cKO animals had reduced thickness of the cortex, especially layers ? and ?. Behavioural tests revealed Grem1Emx1 cKO mice have impaired motor balance and fear sensitivity compared to littermate controls.  Conclusion: Grem1 expression marks an early neuronal progenitor that gives rise to excitatory neurons in the embryonic mouse brain. This BMP antagonist functions to promote neural progenitor proliferation and neuronal differentiation. Overall design: Grem1 traced mouse brains were harvested at 24 hours post tamoxifen induction. Grem1 positive and negative population were separated via FACS. RNA seq were then performed on these two populations. Differential gene expression of Grem 1 positive vs Grem1 negative populations were then performed, and subsequently network analyses on Grem1 positive population to reveal the distinct biological activity.