Alcohol transcriptome changes in mice microglia log2FC
Description:
Microglia are fundamentally important immune cells within the central nervous system (CNS) that respond to environmental challenges to maintain normal physiological processes. Alterations in steady-state cellular function and over-activation of microglia can facilitate the initiation and progression of neuropathological conditions such as Alzheimer’s disease, Multiple Sclerosis, and Major Depressive Disorder. Alcohol consumption disrupts signaling pathways including both innate and adaptive immune responses that are necessary for CNS homeostasis. Coordinate expression of these genes is not ascertained from an admixture of CNS cell-types, underscoring the importance of examining isolated cellular populations to reveal systematic gene expression changes arising from mature microglia. Unbiased RNA-Seq profiling was used to identify gene expression changes in isolated prefrontal cortical microglia in response to recurring bouts of voluntary alcohol drinking behavior. The voluntary ethanol paradigm utilizes long-term consumption ethanol that results in escalated alcohol intake and altered cortical plasticity that is seen in humans. Gene coexpression analysis identified a coordinately regulated group of genes, unique to microglia, that collectively are associated with alcohol consumption. Genes within this group are involved in toll-like receptor signaling and transforming growth factor beta signaling. Network connectivity of this group identified Siglech as a putative hub gene and highlighted the potential importance of proteases in the microglial response to chronic ethanol. In conclusion, we identified a distinctive microglial gene expression signature for neuroimmune responses related to alcohol consumption that provides valuable insight into microglia-specific changes underlying the development of substance abuse, and possibly other CNS disorders.
Authors:
Gizelle M McCarthy, Sean P Farris, Yuri A Blednov, R Adron Harris, R Dayne Mayfield
Alcohol transcriptome changes in mice microglia total homogenate log2FC
Description:
Microglia are fundamentally important immune cells within the central nervous system (CNS) that respond to environmental challenges to maintain normal physiological processes. Alterations in steady-state cellular function and over-activation of microglia can facilitate the initiation and progression of neuropathological conditions such as Alzheimer’s disease, Multiple Sclerosis, and Major Depressive Disorder. Alcohol consumption disrupts signaling pathways including both innate and adaptive immune responses that are necessary for CNS homeostasis. Coordinate expression of these genes is not ascertained from an admixture of CNS cell-types, underscoring the importance of examining isolated cellular populations to reveal systematic gene expression changes arising from mature microglia. Unbiased RNA-Seq profiling was used to identify gene expression changes in isolated prefrontal cortical microglia in response to recurring bouts of voluntary alcohol drinking behavior. The voluntary ethanol paradigm utilizes long-term consumption ethanol that results in escalated alcohol intake and altered cortical plasticity that is seen in humans. Gene coexpression analysis identified a coordinately regulated group of genes, unique to microglia, that collectively are associated with alcohol consumption. Genes within this group are involved in toll-like receptor signaling and transforming growth factor beta signaling. Network connectivity of this group identified Siglech as a putative hub gene and highlighted the potential importance of proteases in the microglial response to chronic ethanol. In conclusion, we identified a distinctive microglial gene expression signature for neuroimmune responses related to alcohol consumption that provides valuable insight into microglia-specific changes underlying the development of substance abuse, and possibly other CNS disorders.
Authors:
Gizelle M McCarthy, Sean P Farris, Yuri A Blednov, R Adron Harris, R Dayne Mayfield
The current study used two inbred mouse strains, C57BL/6 J and A/J, to investigate the genetics of behavioral responses to fentanyl. Mice were tested for conditioned place preference and fentanyl-induced locomotor activity. C57BL/6J mice formed a conditioned place preference to fentanyl injections and fentanyl increased their activity. Neither effect was noted in A/J mice. We conducted RNA-sequencing on the nucleus accumbens of mice used for fentanyl-induced locomotor activity. Surprisingly, we noted few differentially expressed genes using treatment as the main factor. However many genes differed between strains.
Authors:
Samuel J Harp, Mariangela Martini, Will Rosenow, Larry D Mesner, Hugh Johnson, Charles R Farber, Emilie F Rissman
The current study used two inbred mouse strains, C57BL/6 J and A/J, to investigate the genetics of behavioral responses to fentanyl. Mice were tested for conditioned place preference and fentanyl-induced locomotor activity. C57BL/6J mice formed a conditioned place preference to fentanyl injections and fentanyl increased their activity. Neither effect was noted in A/J mice. We conducted RNA-sequencing on the nucleus accumbens of mice used for fentanyl-induced locomotor activity. Surprisingly, we noted few differentially expressed genes using treatment as the main factor. However many genes differed between strains.
Authors:
Samuel J Harp, Mariangela Martini, Will Rosenow, Larry D Mesner, Hugh Johnson, Charles R Farber, Emilie F Rissman
The current study used two inbred mouse strains, C57BL/6 J and A/J, to investigate the genetics of behavioral responses to fentanyl. Mice were tested for conditioned place preference and fentanyl-induced locomotor activity. C57BL/6J mice formed a conditioned place preference to fentanyl injections and fentanyl increased their activity. Neither effect was noted in A/J mice. We conducted RNA-sequencing on the nucleus accumbens of mice used for fentanyl-induced locomotor activity. Surprisingly, we noted few differentially expressed genes using treatment as the main factor. However many genes differed between strains.
Authors:
Samuel J Harp, Mariangela Martini, Will Rosenow, Larry D Mesner, Hugh Johnson, Charles R Farber, Emilie F Rissman
DEG PFC in adolescent D2 mice 24hr post treatment_pvalue
Description:
DBA/2J males and females (n = 24/sex) were orally dosed with 4 g/kg ethanol (25% w/v in water by gavage) or water intermittently (2 days on/2 days off) on PND 29, 30, 33, 34, 37, 38, 41, and 42. Tissue was collected for gene expression studies at PND 43 (n = 22) and PND 66 (n = 19). Behaviorally naïve tissue from the PFC was collected 24 h (at PND 43) and 3 weeks (at PND 66) after the last ethanol binge (dose). Total RNA was analyzed for gene-level expression differences using Mouse Transcriptome Arrays v1.0. Two complementary analyses were conducted to interrogate differential gene expression at each age. Gene Ontology over-representation analysis identified six categories involved in oligodendrocyte development and myelination as the primary Biological Processes altered by adolescent binge ethanol. For transcript IDs significant for the interaction between sex and adolescent treatment, Gene Ontology analysis only identified two over-represented cellular components: ER chaperone component and smooth ER. When comparing gene expression between adolescent males vs. females, most of the differentially expressed genes either resided on the Y chromosome (Ddx3y, Eif2s3y, Kdm5d, Uty), or are known to escape X-inactivation (Ddx3x, Eif2s3x, Kdm5c, Kdm6a) in mice (Yang et al., 2010). Over-represented Gene Ontology categories (Supplementary Table 2) reflect their processes, such as histone demethylase activity, angiotensin catabolic processes in blood, cell adhesion and regulation of gap junction assembly. Genes in this geneset are all significantly altered as a main effect of treatment, sex, or the interaction between treatment and sex (p < 0.01).
Authors:
Jennifer T Wolstenholme, Tariq Mahmood, Guy M Harris, Shahroze Abbas, Michael F Miles
DEG PFC in adolescent D2 mice 24hr post treatment_logFC
Description:
DBA/2J males and females (n = 24/sex) were orally dosed with 4 g/kg ethanol (25% w/v in water by gavage) or water intermittently (2 days on/2 days off) on PND 29, 30, 33, 34, 37, 38, 41, and 42. Tissue was collected for gene expression studies at PND 43 (n = 22) and PND 66 (n = 19). Behaviorally naïve tissue from the PFC was collected 24 h (at PND 43) and 3 weeks (at PND 66) after the last ethanol binge (dose). Total RNA was analyzed for gene-level expression differences using Mouse Transcriptome Arrays v1.0. Two complementary analyses were conducted to interrogate differential gene expression at each age. Gene Ontology over-representation analysis identified six categories involved in oligodendrocyte development and myelination as the primary Biological Processes altered by adolescent binge ethanol. For transcript IDs significant for the interaction between sex and adolescent treatment, Gene Ontology analysis only identified two over-represented cellular components: ER chaperone component and smooth ER. When comparing gene expression between adolescent males vs. females, most of the differentially expressed genes either resided on the Y chromosome (Ddx3y, Eif2s3y, Kdm5d, Uty), or are known to escape X-inactivation (Ddx3x, Eif2s3x, Kdm5c, Kdm6a) in mice (Yang et al., 2010). Over-represented Gene Ontology categories (Supplementary Table 2) reflect their processes, such as histone demethylase activity, angiotensin catabolic processes in blood, cell adhesion and regulation of gap junction assembly. Genes in this geneset are all significantly altered as a main effect of treatment, sex, or the interaction between treatment and sex (p < 0.01).
Authors:
Jennifer T Wolstenholme, Tariq Mahmood, Guy M Harris, Shahroze Abbas, Michael F Miles
Adolescent D2 transcripts sig. altered in PFC using S-score analysis at FDR < 0.05 (EtOH vs control)
Description:
DBA/2J males and females (n = 24/sex) were orally dosed with 4 g/kg ethanol (25% w/v in water by gavage) or water intermittently (2 days on/2 days off) on PND 29, 30, 33, 34, 37, 38, 41, and 42. Tissue was collected for gene expression studies at PND 43 (n = 22) and PND 66 (n = 19). Behaviorally naïve tissue from the PFC was collected 24 h (at PND 43) and 3 weeks (at PND 66) after the last ethanol binge (dose). Total RNA was analyzed for gene-level expression differences using Mouse Transcriptome Arrays v1.0. We performed an analysis using the S-score probe-level algorithm which we have previously shown to have increased sensitivity for differential expression analysis (Zhang et al., 2002; Kennedy et al., 2006). For this analysis, data was collapsed over sex to increase the power to detect differences between ethanol treatment versus controls and to focus on lasting differences following binge ethanol. To assess genes that were persistently regulated long-term following adolescent binge ethanol, we intersected the S-score analysis gene list significantly altered by ethanol in adolescents with the list obtained from adults.
Authors:
Jennifer T Wolstenholme, Tariq Mahmood, Guy M Harris, Shahroze Abbas, Michael F Miles
DEG female mouse forebrain PND1 morphine vs saline_pvalue
Description:
To examine forebrain transcriptomic changes that might elucidate mechanisms of withdrawal, delayed development, and any long-term behavior changes, we generated transcriptomic signatures following our “3-trimester” exposure model (3-Tri). In addition, we also examined transcriptomes from animals that received opioids only during the gestational period (PND1) or only during the last trimester from PND 1–14 (PND 14). We sought to determine whether transcriptomic signatures vary based on the window of exposure, perhaps contributing to the discrepancies in the literature regarding acute and long-term outcomes. Brains were dissected from PND 1 pups 6 h after discovery. Brains were dissected from post-natal exposure only (PND 14) or 3-trimester exposure (3-tri) 6 h after the last morphine or saline injection. The number of animals per group was similar (N = 5–7 animals, male and female C57Bl/6NTac mice), and the quality controls, library construction and sequence parameters were also identical across all groups. Libraries were sequenced on a NovaSeq 6000 at a depth of 30 million total reads/sample using paired-end sequencing of 150 base pairs (PE150), to a depth of 30 million total reads/sample. Reads were then mapped to the mouse reference genome (Mus Musculus, GRCm38/mm10) using HISAT2 (version 2.2.1), and duplicated fragments were removed using Picard MarkDuplicates. Differential expression analysis between two conditions (e.g., Morphine and Saline) was performed in R (version 4.1.1) with DESeq2 (v1.32.0) package. Genes were assigned by the authors as differentially expressed if the (adjusted) (nominal) p-value < 0.05. All genes/scores are presented here.
Authors:
Amelia D Dunn, Shivon A Robinson, Chiso Nwokafor, Molly Estill, Julia Ferrante, Li Shen, Crystal O Lemchi, Jordi Creus-Muncunill, Angie Ramirez, Juliet Mengaziol, Julia K Brynildsen, Mark Leggas, Jamie Horn, Michelle E Ehrlich, Julie A Blendy
DEG male mouse forebrain PND1 morphine vs saline_pvalue
Description:
To examine forebrain transcriptomic changes that might elucidate mechanisms of withdrawal, delayed development, and any long-term behavior changes, we generated transcriptomic signatures following our “3-trimester” exposure model (3-Tri). In addition, we also examined transcriptomes from animals that received opioids only during the gestational period (PND1) or only during the last trimester from PND 1–14 (PND 14). We sought to determine whether transcriptomic signatures vary based on the window of exposure, perhaps contributing to the discrepancies in the literature regarding acute and long-term outcomes. Brains were dissected from PND 1 pups 6 h after discovery. Brains were dissected from post-natal exposure only (PND 14) or 3-trimester exposure (3-tri) 6 h after the last morphine or saline injection. The number of animals per group was similar (N = 5–7 animals, male and female C57Bl/6NTac mice), and the quality controls, library construction and sequence parameters were also identical across all groups. Libraries were sequenced on a NovaSeq 6000 at a depth of 30 million total reads/sample using paired-end sequencing of 150 base pairs (PE150), to a depth of 30 million total reads/sample. Reads were then mapped to the mouse reference genome (Mus Musculus, GRCm38/mm10) using HISAT2 (version 2.2.1), and duplicated fragments were removed using Picard MarkDuplicates. Differential expression analysis between two conditions (e.g., Morphine and Saline) was performed in R (version 4.1.1) with DESeq2 (v1.32.0) package. Genes were assigned by the authors as differentially expressed if the (adjusted) (nominal) p-value < 0.05. All genes/scores are presented here.
Authors:
Amelia D Dunn, Shivon A Robinson, Chiso Nwokafor, Molly Estill, Julia Ferrante, Li Shen, Crystal O Lemchi, Jordi Creus-Muncunill, Angie Ramirez, Juliet Mengaziol, Julia K Brynildsen, Mark Leggas, Jamie Horn, Michelle E Ehrlich, Julie A Blendy
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