EXPOSURE TO ETHANOL INDUCES METABOLIC AND bioenergetic CHANGES IN INDUCED PLURIPOTENT STEM CELL-DERIVED ASTROCYTES

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Kärkkäinen O1, 2, 4, Koskuvi M2, Puttonen KA2, Gao Y2, Vaurio O1, Ojansuu I1, Repo-Tiihonen E1, Koistinaho J2, Tiihonen J1,3, Lehtonen S2

1 Department of Forensic Psychiatry, University of Eastern Finland, Niuvanniemi Hospital, Kuopio, Finland
2 Department of Neurobiology, A.I. Virtanen Institute for Molecular Sciences, University of Eastern Finland, Kuopio, Finland
3 Clinical Neuroscience, Karolinska Institutet, Stockholm, Sweden
4 School of Pharmacy, University of Eastern Finland, Kuopio, Finland

Alcohol has a profound effect on the cells of the central nervous system. Alcohol abuse causes several metabolic and neurological disorders and eventually leads to structural damage of the brain. Pharmacological therapies are mainly directed to molecular pathways in neurons; however, glial cells are also markedly affected by exposure to alcohol. Astrocytes are the most abundant glial cell type in brain tissue that maintains homeostasis and synaptic transmission. Little is known about the contribution or response of astrocytes to alcohol intake. Here we have generated induced pluripotent stem cell (iPSC) lines from violent substance abusers and healthy controls. Selected iPSC lines were differentiated into mature astrocytes which were exposed to medium with or without 33 mM ethanol prior to analyses of metabolic activity, mitochondrial respiration and ROS production. Astrocyte cultures of violent substance abusers lines manifested increased GFAP levels (p=0.014) while S100b levels were unchanged when compared to corresponding cultures derived from controls. In astrocytes derived from violent substance abusers, non-targeted profiling showed for example altered choline metabolism as demonstrated by increased cellular levels of choline (p=0.044) and N-methylethanolamine phosphate (p=0.033), and increased medium levels of glycine betaine (p=0.030). On the other hand, ethanol exposure further altered amino acid metabolism. For example, ethanol exposure decreased serotonin levels (p=0.038) in the astrocytes of violent substance abuser lines and elevated medium levels of another tryptophan metabolite indoleacetaldehyde (p<0.001) in both study groups. In addition, pre-exposure of astrocytes to ethanol resulted in reduction of mitochondrial maximal respiration that was induced by uncoupler FCCP (p=0.016). Moreover, pre-exposure to ethanol increased ROS production in the astrocytes (p=0.028) further supporting the ethanol-induced mitochondrion dysfunction in these cells. To conclude, our data shows the great potential of using iPSC-derived astrocytes in studies of alcohol use. The metabolic and bioenergetic changes were observed in pure astrocyte cultures, suggesting that among violent substance abusers ethanol exposure affects astrocytes in a different way compared to healthy controls, independent of neurons. Additional studies addressing the possible alterations in astrocyte-neuron interaction in violent substance abuser lines might help to understand mechanisms linking abnormal astrocyte function and neurophysiologic disturbances in alcohol abuse.

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