Upregulated GIRK2 Counteracts Ethanol-Induced Changes in Excitability and Respiration in Human Neurons.

Genome-wide association studies (GWAS) of electroencephalographic endophenotypes for alcohol use disorder (AUD) has identified noncoding polymorphisms within the KCNJ6 gene. KCNJ6 encodes GIRK2, a subunit of a G-protein-coupled inwardly rectifying potassium channel that regulates neuronal excitability. We studied the effect of upregulating KCNJ6 using an isogenic approach with human glutamatergic neurons derived from induced pluripotent stem cells (male and female donors). Using multielectrode arrays, population calcium imaging, single-cell patch-clamp electrophysiology, and mitochondrial stress tests, we find that elevated GIRK2 acts in concert with 7-21 d of ethanol exposure to inhibit neuronal activity, to counteract ethanol-induced increases in glutamate response, and to promote an increase intrinsic excitability. Furthermore, elevated GIRK2 prevented ethanol-induced changes in basal and activity-dependent mitochondrial respiration. These data support a role for GIRK2 in mitigating the effects of ethanol and a previously unknown connection to mitochondrial function in human glutamatergic neurons.

Tissue Microarray Lipidomic Imaging Mass Spectrometry Method: Application to the Study of Alcohol-Related White Matter Neurodegeneration.

Central nervous system (CNS) white matter pathologies accompany many diseases across the lifespan, yet their biochemical bases, mechanisms, and consequences have remained poorly understood due to the complexity of myelin lipid-based research. However, recent advances in matrix-assisted laser desorption/ionization-imaging mass spectrometry (MALDI-IMS) have minimized or eliminated many technical challenges that previously limited progress in CNS disease-based lipidomic research. MALDI-IMS can be used for lipid identification, semi-quantification, and the refined interpretation of histopathology. The present work illustrates the use of tissue micro-arrays (TMAs) for MALDI-IMS analysis of frontal lobe white matter biochemical lipidomic pathology in an experimental rat model of chronic ethanol feeding. The use of TMAs combines workload efficiency with the robustness and uniformity of data acquisition. The methods described for generating TMAs enable simultaneous comparisons of lipid profiles across multiple samples under identical conditions. With the methods described, we demonstrate significant reductions in phosphatidylinositol and increases in phosphatidylcholine in the frontal white matter of chronic ethanol-fed rats. Together with the use of a novel rapid peak alignment protocol, this approach facilitates reliable inter- and intra-group comparisons of MALDI-IMS data from experimental models and could be extended to human disease states, including using archival specimens.