Date of Award


Degree Type


Degree Name

Doctor of Philosophy

Field of Study

Integrative Physiology


Graduate School of Biomedical Sciences

First Advisor

Robert T. Mallet

Second Advisor

Steve W. Mifflin

Third Advisor

J. Thomas Cunningham


Normobaric intermittent hypoxia conditioning (IHC) induces a robust cardioprotected phenotype in dogs that is remarkably resistant to ischemia and reperfusion induced myocardial infarction and lethal arrhythmias. Previous studies demonstrated that IHC induced cardioprotection requires β1-adrenergic receptor activity. Cardiac opioid systems are stimulated by, and counteract the harmful effects of, excessive stressors such as sympathetic activity. Additional modes of hypoxic conditioning have been shown to induce synthesis of cardiac enkephalins and delta opioid receptors (DOR). The hypothesis that DOR mediates IHC cardioprotection was examined in two studies conducted in intermittent hypoxia conditioned and non-hypoxic sham dogs. For the first study dogs were assigned to groups subjected to non-hypoxic sham conditioning, IHC, IHC plus the aminothiol antioxidant N-acetylcysteine (NAC), and IHC plus the DOR antagonist naltrindole. After IHC or sham conditioning, the dogs were subjected to an left anterior descending coronary artery occlusion/reperfusion protocol and incidence of reperfusion arrhythmias and myocardial infarct size were measured and adjusted for coronary collateral flow. Naltrindole and NAC abolished the anti-infarct and anti-arrhythmia effects of IHC, in a manner independent of collateral blood flow. Intermittent hypoxia conditioning is thus dependent on DOR activity as well as formation of reactive oxygen species (ROS) during cylic hypoxia-reoxygenation. Whether ROS are generated upstream, downstream, or in parallel to DOR activation remains to be determined.

DORs are abundant on dog parasympathetic nerves and therefore are ideally positioned to stimulate cardioprotective cholinergic activity. However it is unknown in what direction IHC modulates bimodal DORs, i.e. modulation of synaptic inhibitory or excitatory activity. In the second study dogs were assigned to sham conditioned, IHC, and IHC plus naltrindole groups. IHC resulted in a profound enhancement of vagal bradycardia, in the absence and presence of increasingly vagolytic doses of the DOR agonist MEAP. This result demonstrated that IHC shifts DOR signaling in favor of the vagotonic DOR-1 receptor subtype. However, the fate of the vagolytic DOR-2 receptors was unknown. Immunolabeling of atrial tissue revealed that IHC increased content of the monosialoganglioside GM1 in autonomic nerve fibers associated with parasympathetic fibers, an effect which may shift DOR signaling in favor of the DOR-1 subtype. In addition, IHC increased the number of fibers containing the vesicular acetylcholine transporter within the sinoatrial node. However, DOR positive fibers in both the atria and SAN were decreased after IHC, perhaps reflecting redistribution or intracellular trafficking of DOR1 and/or DOR2 receptors. Immunoblotting revealed decreased content of adrenergic protein tyrosine hydroxylase in the left ventricle following IHC. Collectively, these results indicate IHC is dependent on opioidergic activity to induce cardioprotection by enhancing cholinergic signaling components at the expense of adrenergic proteins, suggesting IHC-induced shifting of autonomic balance in favor of parasympathetic control of the heart.


Estrada, Juan A., Intermittent hypoxia induced opioidergic protection of the heart. Doctor of Philosophy (Integrative Physiology), July 20, pp125, 4 tables, 16 figures. Available August 2016.