The role of excitatory neurotransmission in the induction of homeostatic synaptic plasticity Open Access

Fong, Ming-Fai (2014)

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Homeostatic plasticity encompasses a family of compensatory mechanisms that help maintain stability within neural circuits. Synaptic scaling is a form of homeostatic plasticity characterized by a coordinated strengthening or weakening of all synaptic inputs onto a neuron by a common factor as a compensatory response to altered activity. While synaptic scaling has been widely observed both in vitro and in vivo, how neural circuits sense altered activity in order to trigger the scaling process remains unclear. Because prolonged blockade of spiking robustly leads to upward scaling, a leading hypothesis is that neurons monitor their own firing rates to induce scaling. However, chronic blockade of AMPA-type glutamate receptors (AMPARs) also leads to upward scaling, suggesting that reduced excitatory neurotransmission triggers the scaling process. Spiking and excitatory transmission are highly correlated at the circuit level, so distinguishing between reductions in firing rate and reductions in transmission as triggers for scaling presents a unique challenge. In this dissertation, we systematically investigated the independent roles of reduced firing rate versus reduced AMPAergic transmission in the induction of homeostatic synaptic scaling. To test the importance of firing rate in scaling, we used multi-electrode recordings to continuously monitor spiking activity in cultured cortical networks during perturbations that trigger upward scaling. While each perturbation reduced spiking activity to some degree, there was no correlation between the severity of the reduction in firing rate and the degree of scaling observed. Next, we independently manipulated firing rate and AMPAergic transmission using two complementary strategies. First, we blocked AMPARs while restoring normal levels of spiking using closed-loop optogenetic stimulation. Second, we blocked spiking while partially restoring AMPAergic transmission using a pharmacological AMPAR modulator. In both cases, we found that the induction of upward scaling was driven by reductions in AMPAergic transmission, rather than reductions in firing rate. These results provide strong evidence that excitatory neurotransmission is the activity signal sensed by neural circuits in order to trigger homeostatic synaptic scaling. Our findings highlight the role of synaptic activity in the maintenance of circuit stability and raise important questions about the role of scaling in learning, development, and disease.

Table of Contents

Abstract ii

Acknowledgements iv

List of Tables xii

List of Figures xiii

List of Abbreviations xvi

1 Introduction 1

1.1 Activity homeostasis in neural circuits 2

1.2 Homeostatic regulation of intrinsic neuronal excitability 4

1.2.1 Evidence for intrinsic homeostasis 5

1.2.2 Potential triggers for intrinsic homeostasis 8

1.3 Homeostatic regulation of neurotransmitter release 10

1.3.1 Evidence for transmitter release homeostasis 10

1.3.2 Potential triggers for transmitter release homeostasis 14

1.4 Homeostatic regulation of postsynaptic response 15

1.4.1 Evidence for postsynaptic homeostasis 16

1.4.2 Potential triggers for postsynaptic homeostasis 21

1.5 Challenges for studying homeostatic plasticity 23

1.6 Thesis Goals and Organization 24

2 Characterization of spiking activity during chronic AMPA receptor blockade 26

2.1 Introduction 27

2.1.1 Background 27

2.1.2 Micro-electrode arrays 28

2.1.3 Chapter summary 33

2.2 Methods 33

2.2.1 Cell culture 33 Plating 33 MEA Cleaning 35 MEA Sterilization 35

2.2.2 Chronic treatments 35

2.2.3 Microelectrode array electrophysiology 36 Recordings 37 Extracellular unit analysis 37 Firing rate analysis 38 Bursting analysis 39

2.2.4 Patch clamp electrophysiology 40 mEPSC recordings 40 mEPSC analysis 41

2.3 Results 41

2.3.1 Network-wide spiking activity persists during AMPAergic transmission blockade 41

2.3.2 Firing of individual extracellular units persists during AMPAergic blockade 48

2.3.3 Relationship between reductions in spiking and homeostatic changes in synaptic strength 50

2.4 Discussion 54

3 Reductions in AMPAergic transmission directly trigger homeostatic synaptic scaling 66

3.1 Introduction 67

3.1.1 Background 67

3.1.2 Feedback control of neural activity 72

3.1.3 Chapter summary 74

3.2 Methods 74

3.2.1 Transfections 75

3.2.2 Chronic treatments 76

3.2.3 Optical stimulation 77

3.3 Results 79

3.3.1 Optogenetic feedback system for controlling ring rates 79 Stimulus selection 81 Clamping ring rate 81 Compensating for glutamatergic blockade 84

3.3.2 Closed-loop stimulation restores spiking activity during an AMPAergic transmission blockade 86 Network-wide spiking activity 87 Individual unit ring activity 88 Activity patterns within burst 91 Summary 96

3.3.3 Reductions in AMPAergic transmission are sucient to trigger synaptic scaling 96

3.4 Discussion 98

4 Reductions in AMPA receptor activation are required to trigger upward synaptic scaling 107

4.1 Introduction 108

4.1.1 Background 108

4.1.2 Chapter Summary 110

4.2 Methods 111

4.2.1 Patch clamp electrophysiology 111

4.2.2 Chronic treatments 111

4.3 Results 112

4.3.1 Cyclothiazide partially restores AMPAR activation during a spiking blockade 112

4.3.2 Partially restoring AMPAR activation attenuates TTX-induced synaptic scaling 115

4.4 Discussion 119

5 Discussion 125

5.1 Transmission-dependent synaptic scaling: relationships to other forms of synaptic plasticity 126

5.1.1 Local homeostatic regulation of synaptic strength 126

5.1.2 Hebbian plasticity 130 Hebbian plasticity and cell-wide homeostasis 131 Hebbian plasticity and dendrite-wide homeostasis 132 Hebbian plasticity and synapse-specic homeostasis 133

5.1.3 Metaplasticity 134

5.2 Synaptic scaling in the living nervous system 136

5.2.1 Sensory systems 137

5.2.2 Hippocampus 139

5.2.3 Motor systems 140

5.3 Future Outlook 141

5.3.1 Monitoring and manipulating neurotransmission 142

5.3.2 Monitoring and manipulating calcium 144

5.4 Concluding remarks 146

Appendices 148

A Spike Sorting 149

A.1 Approach 149

A.2 Analysis 150

A.3 Conclusion 151

B Compensatory changes in NMDA and GABA synaptic currents help recover synchronous activity during chronic AMPAergic blockade 153

B.1 Motivation 153

B.2 Results 155

B.2.1 Inward GABAergic currents emerge during AMPAergic blockade 155

B.2.2 NMDAergic transmission drives population bursting 157

B.3 Summary 161

Bibliography 165

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