Background: The apolipoprotein E (APOE) gene exists in three isoforms in humans: APOE2, APOE3 and APOE4.
APOE4 causes structural and functional alterations in normal brains, and is the strongest genetic risk factor of the
sporadic form of Alzheimer’s disease (LOAD). Research on APOE4 has mainly focused on the neuronal damage
caused by defective cholesterol transport and exacerbated amyloid-β and Tau pathology. The impact of APOE4 on
non-neuronal cell functions has been overlooked. Astrocytes, the main producers of ApoE in the healthy brain, are
building blocks of neural circuits, and Ca2+ signaling is the basis of their excitability. Because APOE4 modifies
membrane-lipid composition, and lipids regulate Ca2+ channels, we determined whether APOE4 dysregulates
Ca2+signaling in astrocytes.
Methods: Ca2+ signals were recorded in astrocytes in hippocampal slices from APOE3 and APOE4 gene targeted
replacement male and female mice using Ca2+ imaging. Mechanistic analyses were performed in immortalized
astrocytes. Ca2+ fluxes were examined with pharmacological tools and Ca2+ probes. APOE3 and APOE4 expression
was manipulated with GFP-APOE vectors and APOE siRNA. Lipidomics of lysosomal and whole-membranes were
Results: We found potentiation of ATP-elicited Ca2+responses in APOE4 versus APOE3 astrocytes in male, but not
female, mice. The immortalized astrocytes modeled the male response, and showed that Ca2+ hyperactivity
associated with APOE4 is caused by dysregulation of Ca2+ handling in lysosomal-enriched acidic stores, and is
reversed by the expression of APOE3, but not of APOE4, pointing to loss of function due to APOE4 malfunction.
Moreover, immortalized APOE4 astrocytes are refractory to control of Ca2+ fluxes by extracellular lipids, and present
distinct lipid composition in lysosomal and plasma membranes.
Conclusions: Immortalized APOE4 versus APOE3 astrocytes present: increased Ca2+ excitability due to lysosome
dysregulation, altered membrane lipidomes and intracellular cholesterol distribution, and impaired modulation of
Ca2+ responses upon changes in extracellular lipids. Ca2+ hyperactivity associated with APOE4 is found in astrocytes
from male, but not female, targeted replacement mice. The study suggests that, independently of Aβ and Tau
pathologies, altered astrocyte excitability might contribute to neural-circuit hyperactivity depending on APOE allele,
sex and lipids, and supports lysosome-targeted therapies to rescue APOE4 phenotypes in LOAD.