Traumatic Stress, Chronic Ethanol Exposure, or the Combination, Alter Cannabinoid System Components in Reward and Limbic Regions of the Mouse Brain

doi:10.3390/molecules26072086

. 2021 Apr 6;26(7):2086.

doi: 10.3390/molecules26072086.

Traumatic Stress, Chronic Ethanol Exposure, or the Combination, Alter Cannabinoid System Components in Reward and Limbic Regions of the Mouse Brain

Veronica M Piggott^{1

2}, Scott C Lloyd^{1

2}, James I Matchynski^{1

2

3}, Shane A Perrine^{1

2

3}, Alana C Conti^{1

2

3}

Affiliations

¹ Research & Development Service, John D. Dingell VA Medical Center, Detroit, MI 48201, USA.
² Department of Psychiatry and Behavioral Neurosciences, Wayne State University School of Medicine, Detroit, MI 48201, USA.
³ Translational Neuroscience Program, Wayne State University School of Medicine, Detroit, MI 48201, USA.

PMID: 33917316
PMCID: PMC8038692
DOI: 10.3390/molecules26072086

Traumatic Stress, Chronic Ethanol Exposure, or the Combination, Alter Cannabinoid System Components in Reward and Limbic Regions of the Mouse Brain

Veronica M Piggott et al. Molecules. 2021.

. 2021 Apr 6;26(7):2086.

doi: 10.3390/molecules26072086.

Authors

Veronica M Piggott^{1

2}, Scott C Lloyd^{1

2}, James I Matchynski^{1

2

3}, Shane A Perrine^{1

2

3}, Alana C Conti^{1

2

3}

Affiliations

¹ Research & Development Service, John D. Dingell VA Medical Center, Detroit, MI 48201, USA.
² Department of Psychiatry and Behavioral Neurosciences, Wayne State University School of Medicine, Detroit, MI 48201, USA.
³ Translational Neuroscience Program, Wayne State University School of Medicine, Detroit, MI 48201, USA.

PMID: 33917316
PMCID: PMC8038692
DOI: 10.3390/molecules26072086

Abstract

The cannabinoid system is independently affected by stress and chronic ethanol exposure. However, the extent to which co-occurrence of traumatic stress and chronic ethanol exposure modulates the cannabinoid system remains unclear. We examined levels of cannabinoid system components, anandamide, 2-arachidonoylglycerol, fatty acid amide hydrolase, and monoacylglycerol lipase after mouse single-prolonged stress (mSPS) or non-mSPS (Control) exposure, with chronic intermittent ethanol (CIE) vapor or without CIE vapor (Air) across several brain regions using ultra-high-performance liquid chromatography tandem mass spectrometry or immunoblotting. Compared to mSPS-Air mice, anandamide and 2-arachidonoylglycerol levels in the anterior striatum were increased in mSPS-CIE mice. In the dorsal hippocampus, anandamide content was increased in Control-CIE mice compared to Control-Air, mSPS-Air, or mSPS-CIE mice. Finally, amygdalar anandamide content was increased in Control-CIE mice compared to Control-Air, or mSPS-CIE mice, but the anandamide content was decreased in mSPS-CIE compared to mSPS-Air mice. Based on these data we conclude that the effects of combined traumatic stress and chronic ethanol exposure on the cannabinoid system in reward pathway regions are driven by CIE exposure and that traumatic stress affects the cannabinoid components in limbic regions, warranting future investigation of neurotherapeutic treatment to attenuate these effects.

Keywords: 2-arachidonoylglycerol; anandamide; chronic intermittent ethanol; fatty acid amide hydrolase; limbic system; monoacylglycerol lipase; mouse single-prolonged stress; post-traumatic stress disorder; reward pathway.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

**Figure 1**
Anandamide (AEA) and 2-arachidonoylglycerol (2-AG) contents, fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAG-L) levels in the prefrontal cortex (PFC) after mSPS/Control or Air/CIE exposures. (A) Schematic of coronal slice where PFC tissue punches were taken bilaterally (adapted from [58]). Average (B) AEA content (Control-Air: n = 4; Control CIE: n = 7; mSPS-Air: n = 6; mSPS-CIE: n = 8) (C) FAAH levels (Control-Air: n = 6; Control CIE: n = 4; mSPS-Air: n = 5; mSPS-CIE: n = 8) (Inset: representative immunoblotting sample images; kDa kilodaltons) (D) 2-AG content (Control-Air: n = 4; Control CIE: n = 7; mSPS-Air: n = 6; mSPS-CIE: n = 8), and average (E) MAG-L levels (Control-Air: n = 7; Control CIE: n = 4; mSPS-Air: n = 4; mSPS-CIE: n = 7) in the PFC (Inset: representative immunoblotting sample images) did not change among groups after mSPS/Control or Air/CIE exposures. Data are mean ± SEM.

**Figure 2**
Anandamide (AEA) and 2-arachidonoylglycerol (2-AG) contents, fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAG-L) levels in the anterior striatum (STR) after mSPS/Control or Air/CIE exposures. (A) Schematic of coronal slice where anterior STR tissue punches were taken bilaterally (adapted from [58]). Average (B) AEA content (Control-Air: n = 6; Control CIE: n = 7; mSPS-Air: n = 6; mSPS-CIE: n = 8) in the anterior STR was significantly increased in mSPS-CIE mice compared to mSPS-Air mice (* p < 0.05). However, average (C) FAAH level (Control-Air: n = 7; Control CIE: n = 4; mSPS-Air: n = 5; mSPS-CIE: n = 8) did not change among groups (Inset: representative immunoblotting sample images). Average (D) 2-AG content (Control-Air: n = 6; Control CIE: n = 6; mSPS-Air: n = 5; mSPS-CIE: n = 8) in the anterior STR was significantly increased in mSPS-CIE mice compared to mSPS-Air mice (* p < 0.05), but the average (E) MAG-L levels (Control-Air: n = 7; Control CIE: n = 4; mSPS-Air: n = 5; mSPS-CIE: n = 8) did not change among groups (Inset: representative immunoblotting sample images). Data are mean ± SEM.

**Figure 3**
Anandamide (AEA) and 2-arachidonoylglycerol (2-AG) contents, fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAG-L) levels in the nucleus accumbens (NAc) after mSPS/Control or Air/CIE exposures. (A) Schematic of coronal slice where NAc tissue punches were taken bilaterally (adapted from [58]). Average (B) AEA content (Control-Air: n = 3; Control CIE: n = 5; mSPS-Air: n = 6; mSPS-CIE: n = 6) and (C) FAAH levels (Control-Air: n = 7; Control CIE: n = 4; mSPS-Air: n = 5; mSPS-CIE: n = 8) (Inset: representative immunoblotting sample images) did not change among groups after mSPS/Control or Air/CIE exposures. The average (D) 2-AG content (Control-Air: n = 3; Control CIE: n = 6; mSPS-Air: n = 6; mSPS-CIE: n = 7) and the average (E) MAG-L levels (Control-Air: n = 7; Control CIE: n = 4; mSPS-Air: n = 5; mSPS-CIE: n = 8) (Inset: representative immunoblotting sample images) did not change among groups after mSPS/Control or Air/CIE exposures. Data are mean ± SEM.

**Figure 4**
Anandamide (AEA) and 2-arachidonoylglycerol (2-AG) contents, fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAG-L) levels in the dorsal hippocampus (HC) after mSPS/Control or Air/CIE exposures. (A) Schematic of coronal slice where dorsal HC tissue punches were taken bilaterally (adapted from [58]). Average (B) AEA content (Control-Air: n = 6; Control CIE: n = 7; mSPS-Air: n = 6; mSPS-CIE: n = 8) increased in Control-CIE mice compared to Control-Air, mSPS-Air, or mSPS-CIE mice after mSPS/Control or Air/CIE exposures (* p < 0.05). However, the average (C) FAAH levels (Control-Air: n = 6; Control CIE: n = 4; mSPS-Air: n = 5; mSPS-CIE: n = 8) (Inset: representative immunoblotting sample images) did not change among groups after mSPS/Control or Air/CIE exposures. In addition, average (D) 2-AG (Control-Air: n = 6; Control CIE: n = 7; mSPS-Air: n = 6; mSPS-CIE: n = 8) and (E) MAG-L levels (Control-Air: n = 6; Control CIE: n = 4; mSPS-Air: n = 5; mSPS-CIE: n = 8) (Inset: representative immunoblotting sample images) did not change among groups after mSPS/Control or Air/CIE exposures. Data are mean ± SEM.

**Figure 5**
Anandamide (AEA) and 2-arachidonoylglycerol (2-AG) contents, fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAG-L) levels in the amygdala (AMYG) after mSPS/Control or Air/CIE exposures. (A) Schematic of coronal slice where AMYG tissue punches were taken bilaterally (adapted from [58]). Average (B) AEA content (Control-Air: n = 6; Control CIE: n = 6; mSPS-Air: n = 6; mSPS-CIE: n = 8) increased in Control-CIE mice (* p < 0.05) compared to Control-Air or mSPS-CIE mice after mSPS/Control or Air/CIE exposures. However, the average (C) FAAH levels (Control-Air: n = 7; Control CIE: n = 4; mSPS-Air: n = 5; mSPS-CIE: n = 8) (Inset: representative immunoblotting sample images) did not change among groups after mSPS/Control or Air/CIE exposures. In addition, average (D) 2-AG content (Control-Air: n = 6; Control CIE: n = 6; mSPS-Air: n = 6; mSPS-CIE: n = 7) and (E) MAG-L levels (Control-Air: n = 6; Control CIE: n = 3; mSPS-Air: n = 5; mSPS-CIE: n = 8) (Inset: representative immunoblotting sample images) did not change among groups. Data are mean ± SEM.

**Scheme 1**
Schematic overview of (A) the mouse single prolonged stress (mSPS) paradigm, (B) mSPS-CIE consecutive cycles paradigm. After mSPS paradigm and 7 days incubation, mice were exposed to 4 consecutive cycles of CIE exposure.

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References

1. Roberts A.L., Gilman S.E., Breslau J., Breslau N., Koenen K.C. Race/Ethnic Differences in Exposure to Traumatic Events, Development of Post-Traumatic Stress Disorder, and Treatment-Seeking for Post-Traumatic Stress Disorder in the United States. Psychol. Med. 2011;41:71–83. doi: 10.1017/S0033291710000401. - DOI - PMC - PubMed
1. Breslau N., Peterson E.L., Poisson L.M., Schultz L.R., Lucia V.C. Estimating Post-Traumatic Stress Disorder in the Community: Lifetime Perspective and the Impact of Typical Traumatic Events. Psychol. Med. 2004;34:889–898. doi: 10.1017/S0033291703001612. - DOI - PubMed
1. Breslau N., Wilcox H.C., Storr C.L., Lucia V.C., Anthony J.C. Trauma Exposure and Posttraumatic Stress Disorder: A Study of Youths in Urban America. J. Urban Health: Bull. N. Y. Acad. Med. 2004;81:530–544. doi: 10.1093/jurban/jth138. - DOI - PMC - PubMed
1. Perkonigg A., Kessler R.C., Storz S., Wittchen H.U. Traumatic Events and Post-Traumatic Stress Disorder in the Community: Prevalence, Risk Factors and Comorbidity. Acta Psychiatr. Scand. 2000;101:46–59. doi: 10.1034/j.1600-0447.2000.101001046.x. - DOI - PubMed
1. Kessler R.C., Berglund P., Demler O., Jin R., Merikangas K.R., Walters E.E. Lifetime Prevalence and Age-of-Onset Distributions of Dsm-Iv Disorders in the National Comorbidity Survey Replication. Arch. Gen. Psychiatry. 2005;62:593–602. doi: 10.1001/archpsyc.62.6.593. - DOI - PubMed

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[1] Roberts A.L., Gilman S.E., Breslau J., Breslau N., Koenen K.C. Race/Ethnic Differences in Exposure to Traumatic Events, Development of Post-Traumatic Stress Disorder, and Treatment-Seeking for Post-Traumatic Stress Disorder in the United States. Psychol. Med. 2011;41:71–83. doi: 10.1017/S0033291710000401. - DOI - PMC - PubMed

[2] Roberts A.L., Gilman S.E., Breslau J., Breslau N., Koenen K.C. Race/Ethnic Differences in Exposure to Traumatic Events, Development of Post-Traumatic Stress Disorder, and Treatment-Seeking for Post-Traumatic Stress Disorder in the United States. Psychol. Med. 2011;41:71–83. doi: 10.1017/S0033291710000401. - DOI - PMC - PubMed

[3] Breslau N., Peterson E.L., Poisson L.M., Schultz L.R., Lucia V.C. Estimating Post-Traumatic Stress Disorder in the Community: Lifetime Perspective and the Impact of Typical Traumatic Events. Psychol. Med. 2004;34:889–898. doi: 10.1017/S0033291703001612. - DOI - PubMed

[4] Breslau N., Peterson E.L., Poisson L.M., Schultz L.R., Lucia V.C. Estimating Post-Traumatic Stress Disorder in the Community: Lifetime Perspective and the Impact of Typical Traumatic Events. Psychol. Med. 2004;34:889–898. doi: 10.1017/S0033291703001612. - DOI - PubMed

[5] Breslau N., Wilcox H.C., Storr C.L., Lucia V.C., Anthony J.C. Trauma Exposure and Posttraumatic Stress Disorder: A Study of Youths in Urban America. J. Urban Health: Bull. N. Y. Acad. Med. 2004;81:530–544. doi: 10.1093/jurban/jth138. - DOI - PMC - PubMed

[6] Breslau N., Wilcox H.C., Storr C.L., Lucia V.C., Anthony J.C. Trauma Exposure and Posttraumatic Stress Disorder: A Study of Youths in Urban America. J. Urban Health: Bull. N. Y. Acad. Med. 2004;81:530–544. doi: 10.1093/jurban/jth138. - DOI - PMC - PubMed

[7] Perkonigg A., Kessler R.C., Storz S., Wittchen H.U. Traumatic Events and Post-Traumatic Stress Disorder in the Community: Prevalence, Risk Factors and Comorbidity. Acta Psychiatr. Scand. 2000;101:46–59. doi: 10.1034/j.1600-0447.2000.101001046.x. - DOI - PubMed

[8] Perkonigg A., Kessler R.C., Storz S., Wittchen H.U. Traumatic Events and Post-Traumatic Stress Disorder in the Community: Prevalence, Risk Factors and Comorbidity. Acta Psychiatr. Scand. 2000;101:46–59. doi: 10.1034/j.1600-0447.2000.101001046.x. - DOI - PubMed

[9] Kessler R.C., Berglund P., Demler O., Jin R., Merikangas K.R., Walters E.E. Lifetime Prevalence and Age-of-Onset Distributions of Dsm-Iv Disorders in the National Comorbidity Survey Replication. Arch. Gen. Psychiatry. 2005;62:593–602. doi: 10.1001/archpsyc.62.6.593. - DOI - PubMed

[10] Kessler R.C., Berglund P., Demler O., Jin R., Merikangas K.R., Walters E.E. Lifetime Prevalence and Age-of-Onset Distributions of Dsm-Iv Disorders in the National Comorbidity Survey Replication. Arch. Gen. Psychiatry. 2005;62:593–602. doi: 10.1001/archpsyc.62.6.593. - DOI - PubMed

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Traumatic Stress, Chronic Ethanol Exposure, or the Combination, Alter Cannabinoid System Components in Reward and Limbic Regions of the Mouse Brain

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Traumatic Stress, Chronic Ethanol Exposure, or the Combination, Alter Cannabinoid System Components in Reward and Limbic Regions of the Mouse Brain

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