Mechanisms of plasticity in sympathetic preganglionic and postganglionic neurons of the chick embryo Open Access

Abstract

Throughout development, the nervous system is faced with countless challenges to which the cells in the system must have the ability to respond. The autonomic nervous system (ANS), in particular, is designed to respond to internal and environmental changes, to alter the function of different organs and tissues, ultimately regulating body temperature, heart rate, muscle tone, and numerous other important functions. In order to accomplish this, the ANS must be able to transiently sense a change, for example in external temperature, and respond by influencing the tissue appropriately, in this example blood vessel dilation or constriction. However, it is important that once the ideal temperature is reached, and target tissue activation is no longer needed, the system must be able to return to some setpoint of firing rate for the output from the autonomic nervous system, also known as baseline autonomic tone. It is not well known, however, what kinds of cellular mechanisms establish this setpoint, which likely occurs early in development. As we know, there are instances where baseline autonomic tone is chronically imbalanced, such as in hypertension. Therefore, understanding the cellular mechanisms for setting a healthy set point for autonomic tone has important implications for human health. Here, we test whether mechanisms of plasticity, which may play a role in setting up the trajectory for autonomic tone, are expressed during embryonic development of the sympathetic nervous system (SNS).

Table of Contents

Contents

1 Introduction 1

1.1 Background . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1

1.2 Characterization of the sympathetic preganglionic neurons . . . . . . 5

1.3 Characterization of the sympathetic postganglionic neurons . . . . . . 11

1.4 Nicotinic acetylcholine receptor neurotransmission . . . . . . . . . . . 13

1.5 Muscarinic acetylcholine receptor neurotransmission . . . . . . . . . . 17

1.6 The lumbosacral segments of the sympathetic nervous system . . . . 18

1.7 Synaptic transmission at the SPN - PGN synapse . . . . . . . . . . . 19

1.8 Disease and disorder in the autonomic nervous system . . . . . . . . . 22

1.9 Homeostatic plasticity . . . . . . . . . . . . . . . . . . . . . . . . . . 26

1.9.1 Synaptic scaling . . . . . . . . . . . . . . . . . . . . . . . . . . 28

1.9.2 Presynaptic homeostatic plasticity . . . . . . . . . . . . . . . . 30

1.9.3 Homeostatic intrinsic plasticity . . . . . . . . . . . . . . . . . 32

1.9.4 Homeostatic plasticity in the chick embryo spinal cord . . . . 34

1.10 Embryonic development of the sympathetic nervous system . . . . . . 40

1.11 Research objectives of this dissertation . . . . . . . . . . . . . . . . . 44

2 Plasticity in preganglionic and postganglionic neurons of the sympathetic

nervous system during embryonic development 47

2.1 Abstract . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47

2.2 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48

2.3 Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50

2.3.1 Chicken embryos . . . . . . . . . . . . . . . . . . . . . . . . . 50

2.3.2 Tissue isolation . . . . . . . . . . . . . . . . . . . . . . . . . . 51

2.3.3 Retrograde labeling of SPNs and MNs . . . . . . . . . . . . . 51

2.3.4 Calcium imaging of SPNs . . . . . . . . . . . . . . . . . . . . 52

2.3.5 Chloride imaging . . . . . . . . . . . . . . . . . . . . . . . . . 53

2.3.6 Extracellular electrophysiology in PGNs . . . . . . . . . . . . 54

2.3.7 Intracellular electrophysiology . . . . . . . . . . . . . . . . . . 54

2.3.8 Experimental and Statistical Analysis . . . . . . . . . . . . . . 55

2.4 Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 56

2.4.1 Chloride-mediated scaling in sympathetic preganglionic neurons

(SPNs) . . . . . . . . . . . . . . . . . . . . . . . . . . . . 56

2.4.2 Cellular excitability decreased in PGNs following 2-day synaptic

blockade in ovo. . . . . . . . . . . . . . . . . . . . . . . . . 60

2.4.3 No homeostatic adjustment to synaptically-evoked PGN discharge

following acute nicotinic blockade . . . . . . . . . . . . 67

2.5 Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67

3 A late muscarinic response in PGNs following SPN stimulation may

be homeostatic 74

3.1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 74

3.2 Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 77

3.2.1 Chick embryos . . . . . . . . . . . . . . . . . . . . . . . . . . 77

3.2.2 Tissue isolation . . . . . . . . . . . . . . . . . . . . . . . . . . 78

3.2.3 Extracellular recordings . . . . . . . . . . . . . . . . . . . . . 78

3.2.4 Pharmacology . . . . . . . . . . . . . . . . . . . . . . . . . . . 79

3.2.5 Statistical analysis . . . . . . . . . . . . . . . . . . . . . . . . 80

3.3 Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 80

3.4 Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 85

4 Conclusions and Future Directions 90

4.1 Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 90

4.2 Future Directions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 95

4.2.1 Identify critical periods and determine the timeline of expression 96

4.2.2 Further characterization of SPN and PGN plasticity . . . . . . 98

4.2.3 Uncover mechanisms of plasticity in additional componenets of

the SNS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99

4.2.4 Examine effect of spontaneous activity on PGNs throughout

development . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99

4.2.5 Elucidating the impact of nicotinic blockade in vivo . . . . . . 100

4.2.6 Determine whether the plasticity mechanisms are species-specific101

Appendix A Further analyses and approaches for reference 103

A.1 Novel methods for long-term tracking of embryonic limb movements in

ovo . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 103

A.2 Nicotinic and muscarinic blockade: outcomes for excitability of PGNs 109

Bibliography 111

About this Dissertation

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School	Laney Graduate School
Department	Biological and Biomedical Sciences
Subfield / Discipline	Neuroscience
Degree	Ph.D.
Submission	Dissertation
Language	English
Research Field	Biology, Neuroscience
Keyword	Plasticity Development Spinal Cord
Committee Chair / Thesis Advisor	Pete Wenner, Emory University
Committee Members	Marie-Claude Perreault, Emory University Shawn Hochman, Emory University Nicholas Au Yong, Emory University Astrid Prinz, Emory University

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