Cell type-specific assessment of cholesterol distribution in models of neurodevelopmental disorders

Abstract

Most nervous system disorders manifest through alterations in neuronal signaling based on abnormalities in neuronal excitability, synaptic transmission, and cell survival. However, such neuronal phenotypes are frequently accompanied or even caused by metabolic dysfunctions in neuronal or nonneuronal cells. The tight packing and highly heterogenous properties of neural, glial and vascular cell types pose significant challenges to dissecting metabolic aspects of brain disorders. Perturbed cholesterol homeostasis has recently emerged as key parameter associated with sub-sets of neurodevelopmental disorders. However, approaches for tracking and visualizing endogenous cholesterol distribution in the brain have limited capability of resolving cell type-specific differences. We here develop tools for genetically-encoded sensors that report on cholesterol distribution in the mouse brain with cellular resolution. We apply these probes to examine sub-cellular cholesterol accumulation in two genetic mouse models of neurodevelopmental disorders, Npc1 and Ptchd1 knock-out mice. While both genes encode proteins with sterol-sensing domains that have been implicated in cholesterol transport, we uncover highly selective and cell type-specific phenotypes in cholesterol homeostasis. The tools established in this work should facilitate probing sub-cellular cholesterol distribution in complex tissues like the mammalian brain and enable capturing cell type-specific alterations in cholesterol flow between cells in models of brain disorders.

Visualization of accessible cholesterol using a GRAM domain-based biosensor

“…In vitro, D4 binds less efficiently to membranes containing anionic lipids compared to membranes containing neutral lipids [60][61][62] . While substitution of aspartic acid at the 434th position of PFO-D4 to serine (D434S) (i.e., PFO-D4H) generally improves the binding of cytosolic PFO-D4 to the PM 63 , even PFO-D4H fails to bind to the PM in some cell types at steady state 62,[64][65][66][67] . Further, both ALO-D4 and PFO-D4H are known to inhibit the movement of accessible cholesterol [68][69][70] .…”