The role of calcineurin in the recovery of cognitive function following isoflurane anesthesia Open Access

Speigel, Iris (2017)

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General anesthesia, while indispensable for modern healthcare, is suspected as a factor in cognitive dysfunction. Elevated hippocampal surface expression of the α5β3γ2 GABAA receptor (GABAAR) subtype is implicated in memory and synaptic plasticity impairments that persist over a week beyond a brief exposure to isoflurane anesthesia in rodent models. The exact mechanism is unknown, but neuroinflammation has been implicated.

Calcineurin (CN), a calcium-dependent phosphatase that is active during isoflurane exposure, has been mechanistically linked to neurodegeneration and neurotoxicity. CN is also directly involved in synaptic plasticity as a regulator of GABAAR trafficking as well as activity-dependent changes in gene expression and protein synthesis. Calcineurin inhibitors such as cyclosporine A (CSA) are standard of care for immunosuppressive therapy, but the potential for chronic use to affect recovery from anesthesia is unclear.

We explored the potential impact of CSA on cognitive outcomes following isoflurane anesthesia. In an in vivo model, mice given thirty minutes of isoflurane within a chronic CSA treatment paradigm have aberrations in the production and surface expression of α5 containing receptors in their hippocampus, and behaviorally have significant deficits in visuospatial learning. Further analysis of protein expression also revealed a decrease in GAD-67 and α5 total protein, suggesting that changes in inhibitory signaling occur alongside altered GABAAR surface expression. In vitro experiments in primary hippocampal neurons measuring GABAAR surface expression in response to isoflurane demonstrate no acute effect with a one-hour exposure, regardless of CSA pretreatment, suggesting that in vivo memory-impairing aberrations in surface expression manifest post-operatively (e.g during or after emergence).

Our results confirm the work of others that isoflurane induces changes to inhibitory network function and exclude CN inhibition as an intervention. Rather, these findings identify calcineurin inhibitors as a previously unknown risk-factor for the development of post-operative cognitive dysfunction, and identify patients receiving CSA for immunosuppression (e.g. prior to organ transplantation) as a vulnerable group.

Table of Contents

Table of Contents


Chapter 1: Introduction 1

1.1 - Introduction to neuroscience research in anesthesia 2

1.2 - The study of consciousness: specific challenges and experimental approaches 3

1.2.1 - Anesthetics and theories of consciousness 9

1.2.2 - A history of clinical anesthesia 13

1.2.3 - A history of anesthetic theories and mechanisms 16

1.2.4 - Neurophysiology of anesthetic loss of consciousness 18

1.3 - Anesthetic considerations for surgery 22

1.3.1 - The overlap between sleep, coma and anesthesia as EEG Patterns 23

1.3.2 - Problems of modern anesthesia include intraoperative awareness and POCD 27

1.4 - Anesthetic agents modulate ion channels in the CNS 31

1.5 - GABAA receptor diversity and distribution 32

1.6 - Intrasynaptic and extrasynaptic GABAARs generate phasic and tonic current 38

1.7 - Neurophysiologic consequences of specific GABAAR subunit compositions 42

1.8 - Specific GABAAR subtypes in neuroanatomical regions important to anesthesia 43

1.9 - GABAAR surface expression is controlled by intracellular trafficking 45

1.9.1 - Receptor biosynthesis, exocytosis and subcellular localization 46

1.9.2 - Endocytosis and post-endocytotic sorting: recycling vs. degradation 48

1.9.3 - Anesthetic effects on signal transduction pathways that regulate trafficking 50

1.10 - Effect of Anesthetics on GABAAR Surface Expression 55

1.11 - Anesthetics and Inflammation 56

1.12 - Summary of background information and rationale of dissertation 59

Chapter 2: Effects of isoflurane on GABAA receptor trafficking 62

2.1 - Introduction 63

2.2 - Materials and Methods 64

2.3 - Results 73

2.4 - Discussion 88

Chapter 3: Chronic calcineurin inhibition impairs visuospatial learning after 95 isoflurane anesthesia

3.1 - Introduction 96

3.2 - Materials and Methods 97

3.3 - Results 106

3.4 - Discussion 127

Chapter 4: Discussion 131

4.1 - General Overview 132

4.2 - Calcineurin 135

4.3 - Calcineurin in plasticity and learning 136

4.4 - Role of α5 receptors in the hippocampus and functional implications of protein expression changes following anesthesia 140

4.5 - Implications of depressed GAD-67 protein expression 143

4.6 - Calcineurin in anesthetic dysregulation of neuronal function and toxicity 145

4.7 - The study of working memory outcomes following anesthesia: interpreting animal models within context of clinical knowledge 149

4.8 - Pharmacology of Cyclosporine A and implications for anesthetic management 152

4.9 - Conclusions: Summary and perspectives 153

Chapter 5: Extended Methods for isoflurane exposure in vitro, in vivo, and ex vivo techniques 159

5.1 - Anesthetic delivery for in vitro systems 160

5.2 - Anesthetic delivery for in vivo systems 173

References 175

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