University of Calgary PRISM: University of Calgary's Digital Repository Graduate Studies The Vault: Electronic Theses and Dissertations 2020-04-27 The role of the mesocortical dopaminergic pathway in the processing of chronic pain signals Huang, Shuo Huang, S. (2020). The role of the mesocortical dopaminergic pathway in the processing of chronic pain signals (Unpublished doctoral thesis). University of Calgary, Calgary, AB. http://hdl.handle.net/1880/111917 doctoral thesis University of Calgary graduate students retain copyright ownership and moral rights for their thesis. You may use this material in any way that is permitted by the Copyright Act or through licensing that has been assigned to the document. For uses that are not allowable under copyright legislation or licensing, you are required to seek permission. Downloaded from PRISM: https://prism.ucalgary.ca UNIVERSITY OF CALGARY The role of the mesocortical dopaminergic pathway in the processing of chronic pain signals by Shuo Huang A THESIS SUBMITTED TO THE FACULTY OF GRADUATE STUDIES IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY GRADUATE PROGRAM IN NEUROSCIENCE CALGARY, ALBERTA APRIL, 2020 © Shuo Huang 2020 Abstract Chronic pain is a debilitating condition which is prevalent in terminal diseases and aged populations. Pain medications are frequently ineffective for chronic use due to resistance to treatment. This is because the pathophysiology, especially cerebral mechanisms of chronic pain is not fully understood. The processing of chronic pain signals is mainly through the cortical areas, the limbic system, and the nucleus accumbens in the brain, which outputs affect downstream targets exerting top-down control. These brain areas mediate emotional and salience-related processing of pain signals, forming the ‘pain matrix’. The ‘pain matrix’ refers to the brain regions mediating different functions such as valance, salience, emotion, and memory that are able to interact with each other to allow pain perception to emerge. The ‘pain matrix’ also process reward information. Signals from pain and reward converge in the ‘pain matrix’and dopamine modulates the emotional and salience aspects of both. The medial prefrontal cortex (mPFC) is a cortical region that controls many executive functions such as attention, working memory, and learning. The mPFC is involved in pain perception, and undergoes plasticity during development of chronic pain. The PFC receives dopaminergic inputs from the ventral tegmental area (VTA), forming the mecoscortical pathway. The mesocortical circuit modulates neuronal plasticity in the mPFC. This modulation has been shown to affect working memory and aversion; however, whether and how the VTA-mPFC dopaminergic inputs are involved in chronic pain remains incompletely understood. This PhD dissertation examines the hypothesis that VTA dopaminergic neurons undergo plasticity during chronic pain states, and projections from these neurons to the mPFC modulate chronic pain-associated behaviours. Dopaminergic subpopulations of both the lateral and medial VTA were defined by action potential firing patterns. However, plasticity induced by neuropathic chronic pain only resides in specific i dopaminergic subpopulations. In addition, dopaminergic subpopulations of lateral and medial VTA are differentially altered after induction of neuropathic pain. Using optogenetic approaches to selectively target dopaminergic inputs to the mPFC, we found that phasic activation of VTA- mPFC dopaminergic inputs reduced mechanical hypersensitivity during neuropathic pain states. Photostimulation of dopamine input to the mPFC also induced a preference for photostimulation- paired context only in mice with neuropathic pain. Fiber photometry imaging of calcium signals demonstrated that dopamine enhances the activity of mPFC neurons projecting to the ventrolateral periaquductal gray, a crucial downstream target for top-down regulation of pain states. Altogether, this study indicates an important modulatory role of mesocortical dopamine in cerebral chronic pain signaling. ii Acknowledgements I would like to first thank my supervisor Dr. Gerald Zamponi for your encouragement and support to my scientific ideas, and for providing me an excellent platform to conduct research. Without you I would not be able to develop independent and critical thinking. Thank you for all your advices throughout my PhD. I am also very thankful to have Dr. Stephanie Borgland to be my co-supervisor. Thank you for always supporting me with your expertise and all the insightful inputs. The critical attitude to science and warm heart to people are what I learned from you and will carry on in the rest of my career. I am very grateful to my committee Dr. Jaideep Bains and Dr. Tuan Trang for your constructive suggestions for this project, and your generous advices on my academic and non- academic problems. I also felt very lucky to know Dr. William Cole who was an external examiner of my candidacy, but gave me invaluable support till the end of my PhD. Many thanks to Dr. Zizhen Zhang, Eder Gambeta, Nathan Godfrey, Shi Chen Xu, Lina Chen, Catherine Thomas, Dr. Said M'Dahoma, Dr. Vinicius Gadotti, and other lab members from the Zamponi and Borgland lab for your contribution to this study, and to Dr. Tamas Fuzesi and Dr. Leonardo Molina from the HALO Optogenetics Platform for your technical support. Without you this study would not happen. I would like to acknowledge the Hotchkiss Brain Institute and my funding agencies Alberta Innovates-Health Solutions and University of Calgary Eyes High Strategy for supporting through my PhD study, and providing me precious academic opportunities. Finally, thanks to all the obstacles and failures, without which I could not become a better scientist. iii Dedication To my parents, who gave me wings and let me fly. Thank you for always being there, with unconditional love. To Zixiang, who shared all the up and down times with me through the end of my PhD. iv Acknowledging collaborators This work involves collaboration with Dr. Zizhen Zhang, Eder Gambeta, Nathan Godfrey, Shi Chen Xu, Lina Chen, Catherine Thomas, Dr. Said M'Dahoma, Dr. Vinicius Gadotti and other lab members from the Zamponi and Borgland lab. ZZ participated in designing the fiber photometry experiments, and conducted PAG microinjections. EG performed the CPP experiments, analyzed the data, and drafted the method section for CPP. Voltammetry recordings were conducted by SCX. Voltammetry data were analyzed by SCX and organized by CT and SH. CT provided essential technical support and drafted the method section for voltammetry. NG did voltage-clamp recordings for validation of ChR2 function in the VTA, and analyzed the data. LC performed double immunochemistry for c- fos and CamKII. SM contributed to RT-PCR and data analysis. VG performed Complete Freund's Adjuvant and PBS injections. SH performed all other experiments. *This thesis contains three manuscripts (with permissions from the publishers): 1. Dopaminergic modulation of pain signals in the medial prefrontal cortex: challenges and perspectives. Neurosci. Lett. 2018 Nov 29. PMID: 30503912, Huang S, Borgland SL, Zamponi GW. (Introduction, with modifications) 2. Peripheral nerve injury-induced alterations in VTA neuron firing properties. Molecular Brain. PMID: 31685030. Huang, S, Borgland, SL, Zamponi, GW. (Chapter Three) 3. Dopaminergic inputs from the ventral tegmental area into the medial prefrontal cortex modulate neuropathic pain associated behaviors in mice. Cell Reports. Submitted. Huang S, Zhang Z, Gambeta E, Xu SC, Thomas C, Godfrey N, Chen L, M'Dahoma S, Borgland SL, and Zamponi GW. (Chapter Four) v Abbreviations ACC: anterior cingulate cortex Amy: amygdala aCSF: artificial cerebrospinal fluid AHP: afterhyperpolarization potential AP (figures and table only): action potential AP (text-for injection locations): anterior posterior ATP: adenosine triphosphate BDNF: brain-derived neurotrophic factor BLA: basolateral amygdala cAMP: cyclic adenosine monophosphate Cav: voltage-gated calcium channel CaMKII: calcium/calmodulin-dependent protein kinase II CCI: chronic constriction injury ChR2: channelrhodopsin-2 CFA: complete Freund's Adjuvant Con (in the figures): contralateral CPA: conditioned place aversion CPP: conditioned place preference CTB: cholera toxin B subunit D1R: dopamine D1 like receptors D2R: dopamine D2 like receptors DAT: dopamine transporter vi DIO: double-floxed inverted open reading frame dLight: fluorescent dopamine sensor DPA: dynamic plantar aesthesiometer DREADDs: designer receptors exclusively activated by designer drugs DV: dorsal ventral EEG: electroencephalogram EPSC: excitatory postsynaptic current fMRI: functional magnetic resonance imaging F-I: frequency-current FSCV: fast-scan cyclic voltammetry GABA: gamma aminobutyric acid GABAR: gamma aminobutyric acid receptor GCaMP: genetically encoded green fluorescent protein-based calcium sensor Gs: stimulative regulative G-protein Gi/o: inhibitory regulative G-protein Hipp: hippocampus HCN: hyperpolarization-activated cyclic nucleotide–gated Ih: hyperpolarization-activated current INaP: persistent sodium current IPSC: inhibitory postsynaptic current Ips (in the figures): ipsilateral Kir: inward-rectifier potassium ion channel LHb: lateral habenula vii ML: medial lateral mPFC: medial prefrontal cortex mRNA: messenger RNA NAc: nucleus accumbens NAcc: nucleus accumbens core NMDA: N-Methyl-d-aspartic acid or N-Methyl-d-aspartate NMDAR: N-Methyl-d-aspartic