Probes for INS

Sonic hedgehog (SHH) is an essential morphogenetic signal dictating cell fate decisions in several developing organs in mammals. In vitrodata suggest that SHH is required to specify LHX3+/LHX4+ Rathke's pouch (RP) progenitor identity. However, in vivo studies have failed to reveal such a function, supporting instead, a critical role for SHH in promoting proliferation of these RP progenitors and for differentiation of pituitary cell types. Here, we have used a genetic approach to demonstrate that activation of the SHH pathway is necessary to induce LHX3+/LHX4+ RP identity in mouse embryos. First, we show that conditional deletion of Shh in the anterior hypothalamus results in a fully penetrant phenotype characterised by a complete arrest of RP development, with lack of Lhx3/Lhx4 expression in RP epithelium at 9.0 dpc (days post coitum) and total loss of pituitary tissue by 12.5 dpc. Conversely, over-activation of the SHH pathway by conditional deletion of Ptch1 in RP progenitors leads to severe hyperplasia and enlargement of the Sox2+ve stem cell compartment by the end of gestation.

The unique mental abilities of humans are rooted in the immensely expanded and folded neocortex, which reflects the expansion of neural progenitors, especially basal progenitors including basal radial glia (bRGs) and intermediate progenitor cells (IPCs). We found that constitutively active Sonic hedgehog (Shh) signaling expanded bRGs and IPCs and induced folding in the otherwise smooth mouse neocortex, whereas the loss of Shh signaling decreased the number of bRGs and IPCs and the size of the neocortex. SHH signaling was strongly active in the human fetal neocortex but Shh signaling was not strongly active in the mouse embryonic neocortex, and blocking SHH signaling in human cerebral organoids decreased the number of bRGs. Mechanistically, Shh signaling increased the initial generation and self-renewal of bRGs and IPC proliferation in mice and the initial generation of bRGs in human cerebral organoids. Thus, robust SHH signaling in the human fetal neocortex may contribute to bRG and IPC expansion and neocortical growth and folding.

Mitochondrial oxidative phosphorylation (OXPHOS) and substrate utilization critically regulate the function of hypothalamic proopiomelanocortin (POMC)-expressing neurons. Here, we demonstrate that inactivation of apoptosis-inducing factor (AIF) in POMC neurons mildly impairs mitochondrial respiration and decreases firing of POMC neurons in lean mice. In contrast, under diet-induced obese conditions, POMC-Cre-specific inactivation of AIF prevents obesity-induced silencing of POMC neurons, translating into improved glucose metabolism, improved leptin, and insulin sensitivity, as well as increased energy expenditure in AIFΔPOMC mice. On a cellular level, AIF deficiency improves mitochondrial morphology, facilitates the utilization of fatty acids for mitochondrial respiration, and increases reactive oxygen species (ROS) formation in POMC neurons from obese mice, ultimately leading to restored POMC firing upon HFD feeding. Collectively, partial impairment of mitochondrial function shifts substrate utilization of POMC neurons from glucose to fatty acid metabolism and restores their firing properties, resulting in improved systemic glucose and energy metabolism in obesity.

Endosomal sorting complex required for transport (ESCRT) complex proteins regulate biogenesis and release of extracellular vesicles (EVs), which enable cell-to-cell communication in the nervous system essential for development and adult function. We recently showed human loss-of-function (LOF) mutations in ESCRT-III member CHMP1A cause autosomal recessive microcephaly with pontocerebellar hypoplasia, but its mechanism was unclear. Here, we show Chmp1a is required for progenitor proliferation in mouse cortex and cerebellum and progenitor maintenance in human cerebral organoids. In Chmp1a null mice, this defect is associated with impaired sonic hedgehog (Shh) secretion and intraluminal vesicle (ILV) formation in multivesicular bodies (MVBs). Furthermore, we show CHMP1A is important for release of an EV subtype that contains AXL, RAB18, and TMED10 (ART) and SHH. Our findings show CHMP1A loss impairs secretion of SHH on ART-EVs, providing molecular mechanistic insights into the role of ESCRT proteins and EVs in the brain.