Metabolic Regulation of Neuronal Activity and Affective Behavior in the Basolateral Amygdala Public
O'Flaherty, Brendan (Fall 2019)
Published
Abstract
The prevalence of major depressive disorder (MDD), one of the most common disorders in the United States, is doubled in patients with type 2 diabetes mellitus (T2DM). Emerging evidence suggests that MDD and T2DM may share a common etiology: dysregulated metabolism. Dysregulated metabolism could disrupt neuronal activity in key limbic areas, contributing to MDD. The "master" metabolic regulator AMP-activated protein kinase (AMPK) may mediate this relationship. AMPK has been shown to regulate neuronal activity in the hypothalamus. AMPK may also regulate BLA activity and mediate the relationship between metabolic dysfunction and depression. However, AMPK's role in the control and "depressed" BLA remains unknown. The central hypothesis of this study was that the metabolic regulatory molecule AMPK regulates neuronal excitability in the control BLA, and that AMPK becomes dysregulated in rats fed a high-fructose diet (HFrD; a model of metabolic syndrome) leading to BLA hyperexcitability and anxiety-/depression-like behavior.
Firstly, we characterized the effects of a HFrD in male Sprague Dawley rats. We found that as previously reported in Wistar rats, a HFrD increased body fat mass in Sprague Dawley rats, but we did not replicate the reported anxiety-/depression-like phenotype. We also did not observe increased BLA excitability HFrD-fed rats. We then repeated these experiments in a cohort of Wistar rats. Once again we found that a HFrD increased body fat mass, but did not increase anxiety- or depression-like behavior in male or female Wistar rats. Although limited by low statistical power, our experiments in Wistar rats suggest that environmental differences between laboratories best explain the discrepancy between our results and previous studies. These data show that researchers should be extremely careful to control for environmental variables when designing experiments. We also characterized the role of AMPK regulating neuronal excitability in the control BLA. We found that the AMPK inactivator compound C increased BLA excitability, but the AMPK activator AICAR had no effect. We also found that the antidepressant rolipram increased AMPK excitability in an AMPK-dependent manner, suggesting that rolipram's antidepressant effects might also be AMPK-dependent. These experiments identified AMPK as a regulator of BLA excitability and a possible antidepressant target. Collectively, these data lay the groundwork for understanding the relationship between metabolism and mood in both health and disease.
Table of Contents
1 General Introduction . . . . . . . . . . . . . . . . . . . . . . . . . 1
1.1 Major Depressive Disorder . . . . . . . . . . . . . . . . . . . . . . . . 1
1.2 The Amygdala in Major Depressive Disorder . . . . . . . . . . . . . . 4
1.2.1 The Basolateral Complex of the Amygdala . . . . . . . . . . . 4
1.2.2 Inter-Regional Connectivity of the BLC . . . . . . . . . . . . . 16
1.2.3 Role of Monoamines in MDD and BLC Excitability . . . . . . 19
1.2.4 Role of Stress Hormones in MDD and BLC Excitability . . . . 27
1.2.5 Monoamine and Stress Hormone Signaling Converge onto the AMP-Activated Protein Kinase Pathway . . . . . . . . . . . . 30
1.3 AMP-Activated Protein Kinase. . . . . . . . . . . . . . . . . . . . . . 33
1.3.1 The Cellular Role of AMP-Activated Protein Kinase. . . . . . 34
1.3.2 Dietary Fructose and AMPK . . . . . . . . . . . . . . . . . . . 39
1.3.3 AMP-Activated Protein Kinase in the Brain . . . . . . . . . . 40
1.3.4 The Potential Role of AMP-Activated Protein Kinase in Major Depressive Disorder . . . . . . . . . . . . . . . . . . . . . . . . 44
1.4 Experimental Rationale . . . . . . . . . . . . . . . . . . . . . . . . . . 47
2 High-fructose Diet Initiated During Adolescence does not Affect Basolateral Amygdala Excitability or Affective-like Behavior in Sprague Dawley Rats . . . . . . . . . . . . 49
2.1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50
2.2 Methods. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 51
2.2.1 General Housing . . . . . . . . . . . . . . . . . . . . . . . . . . 51
2.2.2 Diet . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 52
2.2.3 Behavior. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53
2.2.4 Metabolic Outcomes . . . . . . . . . . . . . . . . . . . . . . . . 55
2.2.5 Slice Electrophysiology. . . . . . . . . . . . . . . . . . . . . . . 55
2.2.6 Statistical Analysis . . . . . . . . . . . . . . . . . . . . . . . . . 57
2.3 Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 57
2.3.1 Peri-Adolescent High-Fructose Diet Increases Visceral Fat Mass but not Metabolic Efficiency . . . . . . . . . . . . . . . . . . . 57
2.3.2 HFrD Decreases Fasting Blood Glucose . . . . . . . . . . . . . 58
2.3.3 HFrD Did Not Alter Motor Behavior or Affective-like Behaviors 58
2.3.4 HFrD Did Not Affect Membrane Excitability of BLA Principal Neurons . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60
2.4 Discussion. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60
2.4.1 Genetic Strain Differences Could Affect Response to Dietary Fructose . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 64
2.4.2 Laboratory Environment Could Affect Response to Dietary Fructose. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65
2.4.3 HFrD does not change BLA Electrophysiology . . . . . . . . . 66
2.4.4 A HFrD Changes Fat Pad Mass, but not Fasting Blood Glucose or Metabolic Efficiency in Sprague Dawley Rats . . . . . . . . 66
2.4.5 Overall Conclusions . . . . . . . . . . . . . . . . . . . . . . . . 68
3 High-fructose Diet Initiated During Adolescence does not Change Affective-like Behavior in Wistar Rats . . 69
3.1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70
3.2 Methods. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
3.2.1 General Housing . . . . . . . . . . . . . . . . . . . . . . . . . . 71
3.2.2 Diet . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72
3.2.3 Behavior. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73
3.2.4 Metabolic Outcomes . . . . . . . . . . . . . . . . . . . . . . . . 73
3.2.5 Statistical Analysis . . . . . . . . . . . . . . . . . . . . . . . . . 74
3.3 Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 74
3.3.1 Peri-Adolescent High-Fructose Diet Increases Visceral Fat Mass, but does not Change Metabolic Efficiency . . . . . . . . . . . . 74
3.3.2 HFrD Did Not Alter Fasting Blood Glucose . . . . . . . . . . . 75
3.3.3 HFrD Did Not Alter Motor or Affective-like Behaviors. . . . . 79
3.4 Discussion. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 85
3.4.1 Litter Effects and Statistical Power . . . . . . . . . . . . . . . . 85
3.4.2 Effects of a HFrD on Metabolism and Behavior are Similar for Sprague Dawley and Wistar Rats. . . . . . . . . . . . . . . . . 86
3.4.3 Overall Conclusions . . . . . . . . . . . . . . . . . . . . . . . . 88
4 The Role of AMP-Activated Protein Kinase Regulating the Excitability of Control Basolateral Amygdala Neurons . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 90
4.1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 91
4.2 Methods. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 94
4.2.1 General Housing . . . . . . . . . . . . . . . . . . . . . . . . . . 94
4.2.2 Slice Electrophysiology. . . . . . . . . . . . . . . . . . . . . . . 94
4.2.3 Pharmacology. . . . . . . . . . . . . . . . . . . . . . . . . . . . 97
4.2.4 Statistical Analysis . . . . . . . . . . . . . . . . . . . . . . . . . 97
4.3 Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 98
4.3.1 Rolipram Increases Stimulation-Evoked Neuronal Activity . . 98
4.3.2 Rolipram-Enhanced Negative Peak Correlates with Evoked Action Potential Firing . . . . . . . . . . . . . . . . . . . . . . . . 98
4.3.3 Compound C Mimics Rolipram-Induced Increase in Evoked Stimulation Response . . . . . . . . . . . . . . . . . . . . . . . . . . 100
4.3.4 AICAR Blocks Rolipram-Induced Increase in Evoked Stimulation Response . . . . . . . . . . . . . . . . . . . . . . . . . . . . 101
4.4 Discussion. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 101
4.4.1 Rolipram Increases BLA Excitability. . . . . . . . . . . . . . . 101
4.4.2 Compound C Mimics Rolipram's Effect . . . . . . . . . . . . . 103
4.4.3 AICAR Blocks Rolipram's Effect . . . . . . . . . . . . . . . . . 104
4.4.4 Overall Conclusions . . . . . . . . . . . . . . . . . . . . . . . . 105
5 General Discussion and Future Directions . . . . . . . . . 107
5.1 The Effects of a High-Fructose Diet are Variable . . . . . . . . . . . . 108
5.2 The Role of AMPK in the Basolateral Amygdala. . . . . . . . . . . . 110
5.3 Potential Solutions to the "Rolipram Paradox" . . . . . . . . . . . . . 112
5.4 Future Directions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 113
Bibliography . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 116
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