Cell Types in the Bed Nucleus of the Stria Terminalis: Effectors for Stress Modulation of Anxiety Open Access

Daniel, Sarah Elizabeth (2016)

Permanent URL: https://etd.library.emory.edu/concern/etds/7p88cg64w?locale=en
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Abstract

The anterolateral group of the bed nucleus of the stria terminalis (BNSTALG) is a complex brain structure that plays a crucial role in regulating anxiety. It contains multiple sub-regions composed of neurons expressing a range of neuropeptides and receptors. The heterogeneous nature of the BNSTALG, in which sub-regions such as the oval BNST (ovBNST) and anterodorsal BNST (adBNST) modulate anxiety in opposing ways, creates two opposing circuits. Understanding the intrinsic circuitry of the BNSTALG and how this circuitry may be affected by stress will be key to understanding anxiety in both a normal and pathological state. Previous work has defined three distinct cell types in the BNSTALG, namely Type I, Type II, and Type III cells, based on their electrophysiological response to hyperpolarizing and depolarizing current injections and mRNA expression profile. However, little is known about how these neurons contribute to the opposing pathways that facilitate and attenuate anxiety behavior. In this dissertation, we extend the characterization of the cell types in the BNSTALG with the goal of learning how these groups of neurons act together to affect anxiety behavior.

First, we explore differences in electrophysiological cell types across species and regions in the BNSTALG. Type I-III cells were first described in rats, but no study has examined the electrophysiological properties of the cells in the mouse or primate BNSTALG. To this end, we compare the electrophysiological and morphological properties of BNSTALG neurons in the mouse, rat, and rhesus macaque. The mouse and primate BNSTALG contain cells that closely fit the description of Type I-III cells in the rat, however they are observed in significantly different proportions and do not all fit into these classification schemes. These data suggest there may be significant differences in the organization of the BNSTALG across species, however future studies will need to examine these differences further. The BNSTALG is composed of multiple regions, including the ovBNST and adBNST that have differential effects on anxiety. In the third chapter, we compare the cell types found in these two regions and show differential expression of cell types and electrophysiological properties between the ovBNST and adBNST.

Finally, the fourth chapter seeks to explore the effects of chronic shock stress on the cell types in the BNSTALG. Although different cell types are thought to play opposing roles in the anxiety circuit, the effects of stress are often investigated with out an attempt to distinguish separate cell types. Here, we show multiple effects of stress on the electrophysiological and mRNA expression profile of cells in the BNSTALG, however, no effect of stress was observed in all cell types. Specifically Type III cells represent a population of corticotropin releasing factor (CRF) neurons that are uniquely sensitive to chronic stress. These results support the study of individual cell types to gain a better understanding of the circuits within the BNSTALG. We can then use the understanding of these circuits to explore better therapeutic interventions for pathological anxiety.

Table of Contents

Chapter 1: Stress modulation of opposing circuits in the bed nucleus of the stria terminalis
Abstract........................................................................................................................... 1
Introduction..................................................................................................................... 2
Interaction of Stress with Neuromodulators................................................................... 4

Corticotropin Releasing Factor...................................................................................... 4
Norepinephrine........................................................................................................... 15
Dopamine................................................................................................................... 24
Serotonin.................................................................................................................... 32
Stress Modulation of Synaptic Plasticity....................................................................... 44
Clinical Implications...................................................................................................... 52
Future Research Directions........................................................................................... 54
Chapter 2: Comparison of neurons in the BNSTALG in the mouse, rat, and rhesus macaque 57
Introduction................................................................................................................... 58
Methods......................................................................................................................... 60

Animal Subjects.......................................................................................................... 60
Preparation of BNST slices.......................................................................................... 61
General patch clamp recording procedures................................................................. 62
Neuronal morphology................................................................................................ 64
Statistical Analysis....................................................................................................... 65
Results............................................................................................................................ 66
Comparison of this rat sample with previous published samples.................................. 66
Distribution of cell types in the mouse........................................................................ 69
Distribution of cell types in the rhesus macaque.......................................................... 71
Type I cells................................................................................................................. 74
Type II cells................................................................................................................ 75
Type III cells.............................................................................................................. 77
Morphology and Input Resistance............................................................................... 78
Discussion....................................................................................................................... 80
Chapter 3: Comparison of cell types in the oval and anterodorsal BNST................. 87
Introduction................................................................................................................... 88
Methods......................................................................................................................... 89

Animal Subjects.......................................................................................................... 89
Preparation of BNST slices.......................................................................................... 90
General patch clamp recording procedures................................................................. 90
Neuronal morphology................................................................................................ 93
Single cell RT-PCR..................................................................................................... 94
Test of Thy1-Cre mouse line....................................................................................... 95
Statistical analysis........................................................................................................ 96
Results............................................................................................................................ 97
Comparison of cell types in the oval and anterodorsal BNST in the rat........................ 97
Characterization of a novel subpopulation of neurons in Thy1-YFP transgenic mice. 101
Discussion..................................................................................................................... 107
Chapter 4: Chronic stress differentially affects Type III neurons in the bed nucleus of the stria terminalis 121
Introduction................................................................................................................. 122
Methods....................................................................................................................... 124

Animal Subjects........................................................................................................ 124
Chronic stress and behavioral testing......................................................................... 124
Electrophysiology..................................................................................................... 128
Single-cell quantitative PCR...................................................................................... 134
Statistical Analysis..................................................................................................... 137
Results.......................................................................................................................... 138
Context fear and anxiety-like behavior..................................................................... 139
Effect of stress on other electrophysiological properties of BNST neurons................ 140
AMPA-to-NMDA ratio and AMPA rectification........................................................ 144
Effect of stress on CP-AMPA receptors in Type III and non-Type III cells............... 145
Effect of stress on gene expression............................................................................ 147
Discussion..................................................................................................................... 151
Effect of CSS on context fear and anxiety-like behavior........................................... 152
Effect of stress on electrophysiological properties and genetic expression in Type III cells 154
AMPA-to-NMDA ratio............................................................................................. 160
AMPA Rectification and CP-AMPARs...................................................................... 161
Differences in expression of NMDA receptor subunits.............................................. 164
Conclusion................................................................................................................ 166
Chapter 5: Conclusion and Future Directions........................................................... 169
Literature Cited............................................................................................................. 181
List of Figures

Figure 1.1 ………………………………………………………………………………...2
Figure 1.2………………………………………………………………………………..13
Figure 2.1………………………………………………………………………………..61
Figure 2.2………………………………………………………………………………..68
Figure 2.3………………………………………………………………………………..70
Figure 2.4………………………………………………………………………………..72
Figure 2.5………………………………………………………………………………..74
Figure 2.6………………………………………………………………………………..80
Figure 3.1………………………………………………………………………………..97
Figure 3.2………………………………………………………………………………..98
Figure 3.3………………………………………………………………………………100
Figure 3.4………………………………………………………………………………102
Figure 3.5………………………………………………………………………………103
Figure 3.6………………………………………………………………………………105
Figure 3.7………………………………………………………………………………106
Figure 3.8………………………………………………………………………………109
Figure 4.1………………………………………………………………………………139
Figure 4.2………………………………………………………………………………141
Figure 4.3………………………………………………………………………………143
Figure 4.4………………………………………………………………………………145
Figure 4.5………………………………………………………………………………146
Figure 4.6………………………………………………………………………………147
Figure 4.7………………………………………………………………………………149
Figure 4.8………………………………………………………………………………151
Figure 4.9………………………………………………………………………………162
List of Tables
Table 1.1……………………………………………………………………………….…6
Table 1.2………………………………………………………………………………...15
Table 1.3………………………………………………………………………………...33
Table 1.4………………………………………………………………………………...42
Table 2.1…………………………………………………………………………………67
Table 3.1…………………………………………………………………………………99
Table 3.2………………………………………………………………………………. 103
Table 3.3………………………………………………………………………………..104
Table 4.1………………………………………………………………………………..134
Supplementary Material
Supplementary Table 4.1……………………………………………………………..168

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