Graduation date: 1996

No description available.

Remifentanil is an ultra-short-acting phenylpiperidine opioid analgesic that is rapidly metabolized by nonspecific blood and tissue esterases. In clinical practice, remifentanil is now more commonly used during both cardiac and non-cardiac surgery than classic opioid agonists such as morphine, since it can be given in higher doses, is more titratable and enables fast recovery of patients in the postoperative period. Remifentanil preconditioning (RPC), achieved by intravenous remifentanil infusion interspersed with infusion-free periods before indexed ischaemia, attenuates cardiac ischaemia-reperfusion injury (IRI). This is experimentally manifested by reduced postischaemic myocardial infarct size (IS) and diminished markers of cardiac failure and apoptosis, and, clinically, by reduced release of biomarkers of myocardial cellular injury after cardiac surgery. However, the underlying mechanisms by which RPC has a cardioprotective effect need to be further explored. It’s generally considered that the Reperfusion Injury Salvage Kinase (RISK) pathway, triggering the expression of phosphatidylinositol 3-kinase (PI3K) as well as Akt, exerts a pivotal role in both classic ischaemic preconditioning (IPC) and pharmacological preconditioning induced cardioprotection. Moreover, recent studies show that the Survivor Activating Factor Enhancement (SAFE) signalling pathway, which involves signal transducers and activators of transcription-3 (STAT3) and janus activated kinase-2 (JAK2), also has an essential role in IPC. Although cross-talk has been found between the RISK and SAFE pathways, the SAFE pathway can function independently of the RISK to confer cardioprotection. However, the roles of JAK/STAT and PI3K/Akt signalling and, in particular, their relative importance in RPC-mediated cardioprotection have not been studied. I explored whether RPC confers cardioprotection via the JAK/STAT or PI3K/Akt pathway and its relationship with GSK3β inhibition. In first part of my study, I explored relative role of the JAK/STAT and PI3K/Akt which were involved in RPC cardioprotection using JAK2 and PI3K inhibition. Male Sprague-Dawley rats were either sham operated or randomly assigned to receive I/R alone or as well as RPC. Pretreatment with the JAK2 inhibitor AG490 or the PI3K inhibitor wortmannin was induced before ischaemia in rats. RPC reduced myocardial infarction and haemodynamic dysfunction induced by IRI accompanied with increased phosphorylation of STAT3 but not Akt or eNOS phosphorylation. AG490 but not wortmannin cancelled RPC’s cardioprotection. In addition, RPC attenuated hypoxia/reoxygenation induced cardiomyocyte apoptosis while STAT3 knock-out abolished the protective effects of RPC. These findings suggest that RPC confers cardioprotection primarily via activation of the JAK/STAT signalling but not the PI3K/Akt signalling pathway. The second study further investigated the role of GSK3β in RPC cardioprotection using the GSK3β inhibitor SB216763. I found that SB restored the ability of RPC to reduce the extent of myocardial infarction and CK-MB release despite the presence of AG490. The phosphorylation of GSK3β was increased by RPC. In addition, GSK3β gene knock-out with siRNA preserved RPC’s cardioprotection regardless of STAT3 abrogation indicating that GSK3β inhibition plays a critical role as a downstream effector in RPC mediated cardioprotection. Taken together with the evidence from this two part study, I conclude that RPC confers cardioprotection by activating the JAK/STAT and, subsequently, inhibiting GSK3β, a critical downstream effector of RPC cardioprotection. / published_or_final_version / Anaesthesiology / Master / Master of Philosophy

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published_or_final_version / Biochemistry / Master / Master of Philosophy

Substantial evidence shows that activation of glial cells in the spinal cord may promote central sensitization and enhancement of pain. Descending facilitation from the rostroventromedial medulla (RVM) is also recognized as a critical component in the maintenance of chronic pain states, although the precise mechanisms driving this activity are unclear. Here, we investigated the possibility that glial activation in the RVM could promote descending facilitation from the RVM in states of enhanced pain. Peripheral inflammation was induced with carrageenan injected into the plantar aspect of the hindpaw of male Sprague-Dawley rats and behavioral responses to noxious thermal and light tactile stimuli were determined. Microinjection of the glial inhibitors minocycline or fluorocitrate, or of SB 203580, a p38 MAPK inhibitor, produced a significant and time-related reversal of behavioral hypersensitivity resulting from hindpaw inflammation. Moreover, carrageenan-induced inflammation appeared to produce an increase in immunolabeling for activated microglia and astrocytes in the RVM, as well as for phosphorylated p38 MAPK; the latter was localized to both microglia and neurons of the RVM. Microinjection of the glial inhibitors into the RVM appeared to diminish immunofluorescent labeling for activated RVM microglia and astrocytes. Carrageenan-induced inflammation also increased RVM protein levels of Iba1 and GFAP and administration of minocycline or fluorocitrate into the RVM attenuated this effect. To examine a possible mechanism of glial activation, α, β-methylene-ATP was microinjected into the RVM, inducing thermal hyperalgesia, and pre-treatment with the P2X antagonists, PPADS and TNP-ATP, delayed the initiation of ATP-induced hyperalgesia. Post-treatment with the antagonists had no effect on established ATP-induced or carrageenan-induced hypersensitivity. The activation of P2X receptors initiates a signaling cascade leading to the production and release of nociceptive mediators, including BDNF. The RVM microinjection of an anti- BDNF antibody reversed carrageenan-induced thermal hyperalgesia. A model of morphine-induced paradoxical pain was also used to examine the role of glial activation in the RVM. Sustained morphine administration induced tactile allodynia and RVM microinjection of minocycline, but not fluorocitrate, attenuated the behavioral hypersensitivity. Sustained morphine also induced morphological changes in microglia of the RVM, suggesting microglial activation. A third model of enhanced pain used to study medullary glial activation was the spinal nerve ligation (SNL) model of neuropathic pain. The SNL injury induced astrocyte activation within the RVM and microinjection of the astrocyte inhibitor fluorocitrate attenuated the nerve injury-induced tactile allodynia. Minocycline administered to the RVM did not attenuate the behavioral hypersensitivity, suggesting a role for astrocytes, not microglia, in nerve injury-induced enhanced pain. The data show that inflammatory, opioid-induced and neuropathic pain is associated with glial activation in the RVM which likely participates in driving descending pain facilitation via glial-neuronal communication. These findings reveal a novel site of glial modulation of pain.

Exercise is often prescribed for patients with chronic pain, but there is little objective evidence supporting this recommendation. Therefore, we tested the effect of moderate aerobic exercise on the sensory hypersensitivity produced in an animal model of neuropathic pain. Male rats that underwent unilateral ligation of the L5 and L6 spinal nerves (SNL) were divided into exercise-trained or sedentary groups. Exercise training was performed using a treadmill, beginning 7 days after surgery, and continued 5 days a week for 5 weeks. Animals were exercised 30 min/day, at a speed of 14-16 m/min. Sensory testing was performed 23 hours after exercise training. Typical thermal and tactile hypersensitivity developed within 1 week after surgery. Treadmill training reversed thermal and tactile hypersensitivity in injured animals within 4 weeks, but had no effect on sham-operated or non-operated animals. One week after the cessation of exercise training, tactile hypersensitivity returned.The effects of exercise training on SNL-induced sensory hypersensitivity were reversed by the opioid receptor antagonist naloxone. Naloxone or naloxone methiodide reversed the effects of exercise when administered intracerebroventricularly (i.c.v.). Immunohistochemistry revealed increased immunostaining for B-endorphin and met-enkephalin in the periaquaductal grey (PAG) and rostral ventromedial medulla (RVM) regions of exercise-trained animals compared to sedentary animals. An ELISA immunoassay revealed a 31% increase in PAG B-endorphin content in exercise-trained SNL animals. More BDNF was also present in the brain's of exercise-trained animals compared to sedentary, specifically in the ventromedial hypothalamus, hippocampus, and outer rim of the PAG. Administering a BDNF sequestering agent reversed B-endorphin increases in the PAG of exercise-trained animals. Exercise-trained SNL animals treated with 25 ug BDNF sequestering agent (i.c.v.) had lower tactile thresholds compared to the exercise-trained vehicle group.These results support the recommendation of moderate aerobic exercise for patients suffering from neuropathic pain, and suggest that exercise-induced pain reversal results from the upregulation of endogenous opioids in the brainstem. Additionally, increased BDNF with exercise training may play a role in exercise-induced reversal of neuropathic pain by increasing the expression of endogenous opioids, but this needs to be verified further.

Chronic non-cancer pain (CNCP) affects approximately 25% of Canadians. Opioids are medications frequently prescribed for management of patients with CNCP. Concern about addiction, misuse, and diversion for illicit use led the Canadian medical regulatory bodies to release a national guideline on the safe and effective use of opioids in CNCP. This thesis used an online survey to determine how closely the self-reported practices of Canadian family physicians matched the recommendations of the Canadian Guideline. We received 710 responses suitable for analysis. Thirteen percent of respondents did not prescribe strong opioids for CNCP. Practice gaps indentified were infrequently using a management agreement and monitoring pain with a scale; incorrect choice of second line opioid for mild to moderate pain; incorrect choice of first, second, and third line opioids for severe pain, and starting fentanyl incorrectly. Findings provide baseline information for future follow-up to compare physicians’ adherence to the guideline.

Controversy exists concerning the role that dopamine plays in analgesia. In the present studies, dopamine agonists produced analgesia, and D-amphetamine potentiated morphine analgesia, while treatment with 6-hydroxydopamine or mixed or selective D1 and D2 dopamine receptor antagonists attenuated or abolished morphine and D-amphetamine-induced analgesia, in the formalin test. Furthermore, microinjection of morphine into the ventral tegmental area (VTA) and ventral striatum produced analgesia, while intra-VTA microinjection of naloxone methylbromide antagonized the analgesia produced by systemic morphine, in the formalin test. In contrast, similar manipulations of dopamine had little or no effect in the tail flick test. Thus, dopamine appears to play a facilitatory role in formalin test analgesia, and there appear to be fundamental differences between the formalin and tail flick tests and parallels between the role of dopamine in the formalin test and in clinical pain, the vocalization after-discharge test and reward.

In an attempt to investigate the role of the endogenous opioid peptides in substrate utilization and hormonal responses to exercise, eight trained cyclists completed two exercise trials at each of two distinct intensity/duration combinations. Briefly, cyclists completed two trials at 70% VO2max for 90 minutes and two trials at 90% VO2max until exhaustion. Trials were conducted following the administration of the opiate antagonist naloxone (NAL) (0.1 mg-kg-1 bolus + 0.1 mg-kg-1-·hr-1) or volume matched saline (SAL). Serum glucose was maintained at significantly higher levels at 60 and 90 minutes of exercise in 70% NAL vs 70% SAL. Serum glucose was significantly higher at all points during exercise and at 30 and 60 minutes of recovery in 90% NAL vs 90% SAL. Serum insulin was not altered by naloxone administration at either 70% or 90% trials. Serum Cpeptide was significantly higher at 60 and 90 minutes in 70%-NAL vs 70% SAL, and was significantly lower during exercise in 90%-NAL vs 90% SAL. Plasma glucagon was not different during exercise in the 70% trials, but was significantly higher during exercise in 90%-NAL vs 90%-SAL. The glucagon:insulin molar ration was not significantly altered by naloxone administration in any trial. Rating of peceived exertion was significantly higher during exercise in 70%-NAL, but was not different during exercise in the 90% trials. However, time to exhaustion was significantly (18%) reduced in 90%-NAL vs 90%-SAL. No systematic differences were observed in the cardiorespiratory responses to exercise at either intensity, although pulmonary ventilation was modestly (7%) elevated in 90%-NAL. Thus, opiate antagonism prevents the decline in serum glucose seen in prolonged exercise without altering substrate oxidation, and with minimal influence on the pancreatic hormone response. In contrast, opiate antagonism potentiates the hyperglycemic response to high intensity exercise at least in part by altering pancreatic hormone responses which may contribute to the hyperglycemia. / School of Physical Education