Pilot Study of iPS-Derived Neural Cells to Examine Biologic Effects of Alcohol on Human Neurons In Vitro
Background Studies of the effects of alcohol on N‐methyl‐d‐aspartate (NMDA) receptor function and gene expression have depended on rodent or postmortem human brain models. Ideally, the effects of alcohol might better be examined in living neural tissue derived from human subjects. In this study, we...
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| Published in | Alcoholism, clinical and experimental research Vol. 36; no. 10; pp. 1678 - 1687 |
|---|---|
| Main Authors | , , , , |
| Format | Journal Article |
| Language | English |
| Published |
Hoboken, NJ
Blackwell Publishing Ltd
01.10.2012
Wiley |
| Subjects | |
| Online Access | Get full text |
| ISSN | 0145-6008 1530-0277 1530-0277 |
| DOI | 10.1111/j.1530-0277.2012.01792.x |
Cover
| Abstract | Background
Studies of the effects of alcohol on N‐methyl‐d‐aspartate (NMDA) receptor function and gene expression have depended on rodent or postmortem human brain models. Ideally, the effects of alcohol might better be examined in living neural tissue derived from human subjects. In this study, we used new technologies to reprogram human subject‐specific tissue into pluripotent cell colonies and generate human neural cultures as a model system to examine the molecular actions of alcohol.
Methods
Induced pluripotent stem (iPS) cells were generated from skin biopsies taken from 7 individuals, 4 alcohol‐dependent subjects, and 3 social drinkers. We differentiated the iPS cells into neural cultures and characterized them by immunocytochemistry using antibodies for the neuronal marker beta‐III tubulin, glial marker s100β, and synaptic marker synpasin‐1. Electrophysiology was performed to characterize the iPS‐derived neurons and to measure the effects of acute alcohol exposure on the NMDA receptor response in chronically alcohol exposed and nonexposed neural cultures from 1 nonalcoholic. Finally, we examined changes in mRNA expression of the NMDA receptor subunit genes GRIN1, GRIN2A, GRIN2B, and GRIN2D after 7 days of alcohol exposure and after 24‐hour withdrawal from chronic alcohol exposure.
Results
Immunocytochemistry revealed positive staining for neuronal, glial, and synaptic markers. iPS‐derived neurons displayed spontaneous electrical properties and functional ionotropic receptors. Acute alcohol exposure significantly attenuated the NMDA response, an effect that was not observed after 7 days of chronic alcohol exposure. After 7 days of chronic alcohol exposure, there were significant increases in mRNA expression of GRIN1, GRIN2A, and GRIN2D in cultures derived from alcoholic subjects but not in cultures derived from nonalcoholics.
Conclusions
These findings support the potential utility of human iPS‐derived neural cultures as in vitro models to examine the molecular actions of alcohol on human neural cells. |
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| AbstractList | Studies of the effects of alcohol on N-methyl-d-aspartate (NMDA) receptor function and gene expression have depended on rodent or postmortem human brain models. Ideally, the effects of alcohol might better be examined in living neural tissue derived from human subjects. In this study, we used new technologies to reprogram human subject-specific tissue into pluripotent cell colonies and generate human neural cultures as a model system to examine the molecular actions of alcohol.BACKGROUNDStudies of the effects of alcohol on N-methyl-d-aspartate (NMDA) receptor function and gene expression have depended on rodent or postmortem human brain models. Ideally, the effects of alcohol might better be examined in living neural tissue derived from human subjects. In this study, we used new technologies to reprogram human subject-specific tissue into pluripotent cell colonies and generate human neural cultures as a model system to examine the molecular actions of alcohol.Induced pluripotent stem (iPS) cells were generated from skin biopsies taken from 7 individuals, 4 alcohol-dependent subjects, and 3 social drinkers. We differentiated the iPS cells into neural cultures and characterized them by immunocytochemistry using antibodies for the neuronal marker beta-III tubulin, glial marker s100β, and synaptic marker synpasin-1. Electrophysiology was performed to characterize the iPS-derived neurons and to measure the effects of acute alcohol exposure on the NMDA receptor response in chronically alcohol exposed and nonexposed neural cultures from 1 nonalcoholic. Finally, we examined changes in mRNA expression of the NMDA receptor subunit genes GRIN1, GRIN2A, GRIN2B, and GRIN2D after 7 days of alcohol exposure and after 24-hour withdrawal from chronic alcohol exposure.METHODSInduced pluripotent stem (iPS) cells were generated from skin biopsies taken from 7 individuals, 4 alcohol-dependent subjects, and 3 social drinkers. We differentiated the iPS cells into neural cultures and characterized them by immunocytochemistry using antibodies for the neuronal marker beta-III tubulin, glial marker s100β, and synaptic marker synpasin-1. Electrophysiology was performed to characterize the iPS-derived neurons and to measure the effects of acute alcohol exposure on the NMDA receptor response in chronically alcohol exposed and nonexposed neural cultures from 1 nonalcoholic. Finally, we examined changes in mRNA expression of the NMDA receptor subunit genes GRIN1, GRIN2A, GRIN2B, and GRIN2D after 7 days of alcohol exposure and after 24-hour withdrawal from chronic alcohol exposure.Immunocytochemistry revealed positive staining for neuronal, glial, and synaptic markers. iPS-derived neurons displayed spontaneous electrical properties and functional ionotropic receptors. Acute alcohol exposure significantly attenuated the NMDA response, an effect that was not observed after 7 days of chronic alcohol exposure. After 7 days of chronic alcohol exposure, there were significant increases in mRNA expression of GRIN1, GRIN2A, and GRIN2D in cultures derived from alcoholic subjects but not in cultures derived from nonalcoholics.RESULTSImmunocytochemistry revealed positive staining for neuronal, glial, and synaptic markers. iPS-derived neurons displayed spontaneous electrical properties and functional ionotropic receptors. Acute alcohol exposure significantly attenuated the NMDA response, an effect that was not observed after 7 days of chronic alcohol exposure. After 7 days of chronic alcohol exposure, there were significant increases in mRNA expression of GRIN1, GRIN2A, and GRIN2D in cultures derived from alcoholic subjects but not in cultures derived from nonalcoholics.These findings support the potential utility of human iPS-derived neural cultures as in vitro models to examine the molecular actions of alcohol on human neural cells.CONCLUSIONSThese findings support the potential utility of human iPS-derived neural cultures as in vitro models to examine the molecular actions of alcohol on human neural cells. Studies of the effects of alcohol on N-methyl-d-aspartate (NMDA) receptor function and gene expression have depended on rodent or postmortem human brain models. Ideally, the effects of alcohol might better be examined in living neural tissue derived from human subjects. In this study, we used new technologies to reprogram human subject-specific tissue into pluripotent cell colonies and generate human neural cultures as a model system to examine the molecular actions of alcohol. Induced pluripotent stem (iPS) cells were generated from skin biopsies taken from 7 individuals, 4 alcohol-dependent subjects, and 3 social drinkers. We differentiated the iPS cells into neural cultures and characterized them by immunocytochemistry using antibodies for the neuronal marker beta-III tubulin, glial marker s100 beta , and synaptic marker synpasin-1. Electrophysiology was performed to characterize the iPS-derived neurons and to measure the effects of acute alcohol exposure on the NMDA receptor response in chronically alcohol exposed and nonexposed neural cultures from 1 nonalcoholic. Finally, we examined changes in mRNA expression of the NMDA receptor subunit genes GRIN1,GRIN2A,GRIN2B, and GRIN2D after 7 days of alcohol exposure and after 24-hour withdrawal from chronic alcohol exposure. Immunocytochemistry revealed positive staining for neuronal, glial, and synaptic markers. iPS-derived neurons displayed spontaneous electrical properties and functional ionotropic receptors. Acute alcohol exposure significantly attenuated the NMDA response, an effect that was not observed after 7 days of chronic alcohol exposure. After 7 days of chronic alcohol exposure, there were significant increases in mRNA expression of GRIN1, GRIN2A, and GRIN2D in cultures derived from alcoholic subjects but not in cultures derived from nonalcoholics. These findings support the potential utility of human iPS-derived neural cultures as in vitro models to examine the molecular actions of alcohol on human neural cells. Studies of the effects of alcohol on N-methyl-d-aspartate (NMDA) receptor function and gene expression have depended on rodent or postmortem human brain models. Ideally, the effects of alcohol might better be examined in living neural tissue derived from human subjects. In this study, we used new technologies to reprogram human subject-specific tissue into pluripotent cell colonies and generate human neural cultures as a model system to examine the molecular actions of alcohol. Induced pluripotent stem (iPS) cells were generated from skin biopsies taken from 7 individuals, 4 alcohol-dependent subjects, and 3 social drinkers. We differentiated the iPS cells into neural cultures and characterized them by immunocytochemistry using antibodies for the neuronal marker beta-III tubulin, glial marker s100β, and synaptic marker synpasin-1. Electrophysiology was performed to characterize the iPS-derived neurons and to measure the effects of acute alcohol exposure on the NMDA receptor response in chronically alcohol exposed and nonexposed neural cultures from 1 nonalcoholic. Finally, we examined changes in mRNA expression of the NMDA receptor subunit genes GRIN1, GRIN2A, GRIN2B, and GRIN2D after 7 days of alcohol exposure and after 24-hour withdrawal from chronic alcohol exposure. Immunocytochemistry revealed positive staining for neuronal, glial, and synaptic markers. iPS-derived neurons displayed spontaneous electrical properties and functional ionotropic receptors. Acute alcohol exposure significantly attenuated the NMDA response, an effect that was not observed after 7 days of chronic alcohol exposure. After 7 days of chronic alcohol exposure, there were significant increases in mRNA expression of GRIN1, GRIN2A, and GRIN2D in cultures derived from alcoholic subjects but not in cultures derived from nonalcoholics. These findings support the potential utility of human iPS-derived neural cultures as in vitro models to examine the molecular actions of alcohol on human neural cells. Background Studies of the effects of alcohol on N‐methyl‐d‐aspartate (NMDA) receptor function and gene expression have depended on rodent or postmortem human brain models. Ideally, the effects of alcohol might better be examined in living neural tissue derived from human subjects. In this study, we used new technologies to reprogram human subject‐specific tissue into pluripotent cell colonies and generate human neural cultures as a model system to examine the molecular actions of alcohol. Methods Induced pluripotent stem (iPS) cells were generated from skin biopsies taken from 7 individuals, 4 alcohol‐dependent subjects, and 3 social drinkers. We differentiated the iPS cells into neural cultures and characterized them by immunocytochemistry using antibodies for the neuronal marker beta‐III tubulin, glial marker s100β, and synaptic marker synpasin‐1. Electrophysiology was performed to characterize the iPS‐derived neurons and to measure the effects of acute alcohol exposure on the NMDA receptor response in chronically alcohol exposed and nonexposed neural cultures from 1 nonalcoholic. Finally, we examined changes in mRNA expression of the NMDA receptor subunit genes GRIN1, GRIN2A, GRIN2B, and GRIN2D after 7 days of alcohol exposure and after 24‐hour withdrawal from chronic alcohol exposure. Results Immunocytochemistry revealed positive staining for neuronal, glial, and synaptic markers. iPS‐derived neurons displayed spontaneous electrical properties and functional ionotropic receptors. Acute alcohol exposure significantly attenuated the NMDA response, an effect that was not observed after 7 days of chronic alcohol exposure. After 7 days of chronic alcohol exposure, there were significant increases in mRNA expression of GRIN1, GRIN2A, and GRIN2D in cultures derived from alcoholic subjects but not in cultures derived from nonalcoholics. Conclusions These findings support the potential utility of human iPS‐derived neural cultures as in vitro models to examine the molecular actions of alcohol on human neural cells. |
| Author | Lieberman, Richard Abreu, Christine Kranzler, Henry R. Covault, Jonathan Levine, Eric S. |
| Author_xml | – sequence: 1 givenname: Richard surname: Lieberman fullname: Lieberman, Richard organization: Graduate Program in Neuroscience, University of Connecticut Health Center, Connecticut, Farmington – sequence: 2 givenname: Eric S. surname: Levine fullname: Levine, Eric S. organization: Department of Neuroscience, University of Connecticut Health Center, Connecticut, Farmington – sequence: 3 givenname: Henry R. surname: Kranzler fullname: Kranzler, Henry R. organization: Department of Psychiatry, University of Pennsylvania and VISN 4 MIRECC, Philadelphia VAMC, Pennsylvania, Philadelphia – sequence: 4 givenname: Christine surname: Abreu fullname: Abreu, Christine organization: Clinical Research Center, University of Connecticut Health Center, Connecticut, Farmington – sequence: 5 givenname: Jonathan surname: Covault fullname: Covault, Jonathan email: jocovault@uchc.edu organization: Alcohol Research Center, Department of Psychiatry, University of Connecticut Health Center, Connecticut., Farmington |
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| Keywords | Human Ethanol iPS Cell Electrophysiology Induced pluripotent stem cell Nervous system Alcohol Glutamate receptor Gene expression Epidemiology In vitro Pilot study Alcoholic beverage Neuron NMDA receptor Public health |
| Language | English |
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Studies of the effects of alcohol on N‐methyl‐d‐aspartate (NMDA) receptor function and gene expression have depended on rodent or postmortem human... Studies of the effects of alcohol on N-methyl-d-aspartate (NMDA) receptor function and gene expression have depended on rodent or postmortem human brain... |
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| SubjectTerms | Alcohol Alcohol Drinking - metabolism Alcohol Drinking - pathology Alcoholism - metabolism Alcoholism - pathology Alcoholism and acute alcohol poisoning Animals Biological and medical sciences Cell Differentiation - drug effects Cell Differentiation - physiology Cells, Cultured Coculture Techniques Electrophysiology Embryonic Stem Cells - drug effects Embryonic Stem Cells - metabolism Ethanol - pharmacology Fibroblasts - drug effects Fibroblasts - metabolism Gene Expression Humans Induced Pluripotent Stem Cells - cytology Induced Pluripotent Stem Cells - drug effects Induced Pluripotent Stem Cells - metabolism iPS Cell Medical sciences Mice Nerve Tissue Proteins - biosynthesis Neurons - drug effects Neurons - metabolism NMDA Receptor Pilot Projects Receptors, N-Methyl-D-Aspartate - biosynthesis Toxicology |
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| Title | Pilot Study of iPS-Derived Neural Cells to Examine Biologic Effects of Alcohol on Human Neurons In Vitro |
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