Medical Pharmacology Commons^™

Enzymatic Regulation Of Opioid Antinociception And Tolerance, Lynn Hull

Theses and Dissertations

ENZYMATIC REGULATION OF OPIOID ANTINOCICEPTION AND TOLERANCE By Lynn C. Hull, Ph.D. A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy at Virginia Commonwealth University. Virginia Commonwealth University, 2009 Director: William L. Dewey, Ph.D. Department of Pharmacology and Toxicology The involvement of kinases in opioid actions has long been established. The acute actions of opioids, through the Gi/Go G-proteins, cause the inhibition of adenylyl cyclase and therefore a decrease in protein kinase A (PKA) activation. Additionally, acute opioid administration may cause the G-protein to activate the phospholipase C (PLC)-mediated cascade leading to the …

Triptolide Is A Potential Therapeutic Agent For Alzheimer’S Disease, Matthew Allsbrook

Theses and Dissertations

Mounting evidence indicates an involvement of inflammation in the pathogenesis of Alzheimer’s disease. While there are other mechanisms involved, it is this role of inflammatory processes that we wish to investigate. Triptolide is the major constituent in the Chinese herb, Tripterygium wilfordii Hook F, and has been used for centuries as part of Chinese herbal medicine. The four ringed structure has close homology to drugs of the steroid class and it has been shown to be beneficial as an anti-inflammatory for rheumatoid arthritis and for treatment of certain cancers. The aim of this study was to evaluate the potential therapeutic …

Transcriptional, Epigenetic, And Signal Events In Antifolate Therapeutics, Alexandra Racanelli

Theses and Dissertations

A targeted approach to the development of antifolate therapies has been sought for many years. Central to the success of such development is an understanding of the molecular mechanisms dictating the sensitivity of cells to antifolates and the fundamental differences of these processes between normal and neoplastic phenotypes. This dissertation addressed transcriptional mechanisms and cell-signaling events responsible for the efficacy of antifolate therapies. Transcriptional processes and cell signaling pathways are often aberrant in neoplastic tissues, providing a potential point of distinction between a normal and neoplastic cellular state. Folylpolyglutamate synthetase (FPGS) catalyzes the formation of poly-γ-glutamate derivatives of folates and …

The Effect Of Chronic Constriction Injury On Cellular Systems Within Nociceptive Pathways In The Mouse, Michelle Hoot

Theses and Dissertations

Chronic neuropathic pain is often difficult to treat due to its resistance to therapeutic intervention. This is due in part to the poor understanding of the physiological mechanisms involved in the establishment and maintenance of neuropathic pain states. The neuropathic pain model, chronic constriction injury of the sciatic nerve, produced robust pain hypersensitivity in our mice. It also induced significant changes in the mitogen activated protein kinase family, and the cannabinoid and µ-opioid systems in three different brain areas involved in the modulation or regulation of pain states. CCI induced a 2.5 fold increase in mRNA of the kinase Raf-1 …

Therapeutic Drugs In Cancer, Teneille Walker

Theses and Dissertations

The first study examined the interaction between low doses of the multi-kinase inhibitor sorafenib and the histone deacetylase inhibitor vorinostat in colon cancer cells. Sorafenib and vorinostat synergized to kill HCT116 and SW480 cells. In SW480 cells, sorafenib+vorinostat toxicity correlated with CD95 activation and CD95-stimulated autophagy. Drug lethality in SW480 cells was blocked by knock down of CD95. In SW620 cells that are patient matched to SW480 cells, sorafenib+vorinostat toxicity was significantly reduced that correlated with a lack of CD95 activation and lower expression of ceramide synthase 6 (LASS6). Overexpression of LASS6 in SW620 cells enhanced drug-induced CD95 activation and …

Therapeutic Drugs In Cancer And Resistance., Aditi Pandya Martin

Theses and Dissertations

We investigated the mechanism of toxicity and resistance development of small molecule tyrosine kinase inhibitor lapatinib in HCT 116 colon cancer cells. Lapatinib mediated cell death in HCT 116 cells was caspase independent and involved cytosolic release of apoptosis inducing factor. Treatment of HCT 116 cells with 10µM Lapatinib lead to the outgrowth of lapatinib resistant HCT 116 cells. Our studies show that alterations in the expression and activation of Bcl-2 family proteins allow lapatinib resistant HCT 116 cells to resist cytotoxic effects of lapatinib as well as of other commonly used chemotherapeutic agents. In hepatoma and pancreatic cancer cells, …

Synchronization Of Pancreatic Islet Oscillations By Intrapancreatic Ganglia: A Modeling Study, B. Fendler, M. Zhang, L. Satin, R. Bertram

Pharmacology and Toxicology Publications

Abstract

Plasma insulin measurements from mice, rats, dogs, and humans indicate that insulin levels are oscillatory, reflecting pulsatile insulin secretion from individual islets. An unanswered question, however, is how the activity of a population of islets is coordinated to yield coherent oscillations in plasma insulin. Here, using mathematical modeling, we investigate the feasibility of a potential islet synchronization mechanism, cholinergic signaling. This hypothesis is based on well-established experimental evidence demonstrating intrapancreatic parasympathetic (cholinergic) ganglia and recent in vitro evidence that a brief application of a muscarinic agonist can transiently synchronize islets. We demonstrate using mathematical modeling that periodic pulses of …

Creation Of A Mouse With A Humanized Fpgs Gene Compatible With Normal Development, Linying Xie

Theses and Dissertations

Abstract: Folylpoly-γ-glutamate synthetase (FPGS) catalyzes the formation of polyglutamate forms of the reduced folates and antifolates such as methotrexate (MTX) and pemetrexed; this allows the retention of folates and antifolate cancer drugs inside the cell. The enzyme activity of FPGS is essential for cell proliferation and survival. The mouse fpgs gene contains two promoters spaced 10 kb apart which are activated in a tissue-specific manner. The upstream promoter (P1) and exons A1a and A1b are used in some differentiated tissues, mainly liver and kidney, whereas the downstream promoter (P2) and exon 1 are used in rapidly dividing cells. In contrast, …

Identification Of Pharmacological And Molecular Mechanisms Involved In Nicotine Withdrawal, Kia Jackson

Theses and Dissertations

Tobacco dependence is the leading cause of preventable death in the United States. Despite currently available smoking cessation therapies, there is a high rate of relapse in smoking among those attempting to quit. While the somatic signs of nicotine withdrawal (insomnia, increased appetite, weight gain) contribute to the continuation of smoking behavior, it has been hypothesized that the affective signs (depression, anxiety, craving, irritability) are greater motivators of relapse and continued tobacco use. There are few studies that assess the molecular and receptor-mediated mechanisms of nicotine withdrawal; therefore, our studies focus on identifying the nicotinic acetylcholine receptor (nAChR) subtypes and …

Promotion Of Tumor Cell Death Through The Induction Of, Tuyen Nguyen

Theses and Dissertations

Microtubule poisons have proven to be effective in the treatment of a variety of malignancies. Although taxol-based derivatives promote microtubule stabilization, there is continuing interest in compounds that, like colchicines, act as microtubule destabilizing agents. Previous work from this laboratory showed that the novel microtubule poison, JG- 03-14, was active against breast tumor cells, promoting autophagic cell death. In the current work, we studied the influence of JG-03-14 on p53 wild type HCT116 colon carcinoma cells. A crystal violet sensitivity assay indicated that JG-03-14 induced growth inhibition, with 75% suppression of growth evident at a concentration of 500 nM. Time …

Morphine Exacerbates Hiv-1 Tat-Induced Cytokine Production In Astrocytes Through Convergent Effects On [Ca2+]I, Nf-Κb Trafficking And Transcription, Nazira El-Hage, Annadora J. Bruce-Keller, Tatiana Yakovleva, Igor Bazov, Georgy Bakalkin, Pamela E. Knapp, Kurt F. Hauser

Pharmacology and Toxicology Publications

Astroglia are key cellular sites where opiate drug signals converge with the proinflammatory effects of HIV-1 Tat signals to exacerbate HIV encephalitis. Despite this understanding, the molecular sites of convergence driving opiate-accelerated neuropathogenesis have not been deciphered. We therefore explored potential points of interaction between the signaling pathways initiated by HIV-1 Tat and opioids in striatal astrocytes. Profiling studies screening 152 transcription factors indicated that the nuclear factor-kappa B (NF-κB) subunit, c-Rel, was a likely candidate for Tat or Tat plus opiate-induced increases in cytokine and chemokine production by astrocytes. Pretreatment with the NF-κB inhibitor parthenolide provided evidence that Tat±morphine-induced …

Long Lasting Synchronization Of Calcium Oscillations By Cholinergic Stimulation In Isolated Pancreatic Islets, Min Zhang, Bernard Fendler, Bradford Peercy, Pranay Goel, Richard Bertram, Arthur Sherman, Leslie Satin

Pharmacology and Toxicology Publications

Abstract

Individual mouse pancreatic islets exhibit oscillations in [Ca²⁺]_i and insulin secretion in response to glucose in vitro, but how the oscillations of a million islets are coordinated within the human pancreas in vivo is unclear. Islet to islet synchronization is necessary, however, for the pancreas to produce regular pulses of insulin. To determine whether neurohormone release within the pancreas might play a role in coordinating islet activity, [Ca²⁺]_i changes in 4–6 isolated mouse islets were simultaneously monitored before and after a transient pulse of a putative synchronizing agent. The degree of synchronicity was …

Gap Junction Coupling And Calcium Waves In The Pancreatic Islet, Richard K.P. Benninger, Min Zhang, W. Steven Head, Leslie S. Satin, David W. Piston

Pharmacology and Toxicology Publications

Abstract

The pancreatic islet is a highly coupled, multicellular system that exhibits complex spatiotemporal electrical activity in response to elevated glucose levels. The emergent properties of islets, which differ from those arising in isolated islet cells, are believed to arise in part by gap junctional coupling, but the mechanisms through which this coupling occurs are poorly understood. To uncover these mechanisms, we have used both high-speed imaging and theoretical modeling of the electrical activity in pancreatic islets under a reduction in the gap junction mediated electrical coupling. Utilizing islets from a gap junction protein connexin 36 knockout mouse model together …

Endocannabinoid Modulation Of Spatial Memory In Aversively And Appetitively Motivated Barnes Maze Tasks, John Pinckney Harloe

Theses and Dissertations

Genetic deletion or pharmacological blockade of the CB1 receptor has been reported to impair extinction learning in aversive conditioning (i.e., conditioned fear and Morris water maze) paradigms, but not in operant procedures in which food reinforcement is earned. It is difficult to discern whether the differential effects caused by CB1 receptor disruption on extinction result from the hedonics (i.e., aversive vs. appetitive) or is related to the required responses associated with these disparate tasks. In order to evaluate whether the hedonics is the determining factor, we used either aversive (i.e., escape from bright lights and air turbulence) or appetitive (i.e., …

Examining The Behavioral And Molecular Aspects Of Adolescent Nicotine Dependence: Implications For Vulnerability To Drugs Of Abuse, Dena Heath Kota

Theses and Dissertations

Approximately 200 million men and 100 million women smoke worldwide. In the United States, an estimated 25.9 million men (23.9 percent) and 20.7 million women (18.1 percent) are smokers. The commencement of smoking at a young age is thought to increase addiction liability, decrease the probability of successful cessation, and correlate with a higher number of cigarettes smoked per day. Studies from the World Health Organization indicate that between 80,000 and 100,000 children start smoking every day worldwide. These statistics suggest that adolescence is a critical phase for developing nicotine dependence. The work in this dissertation contributes to the further …

Interaction Of Glycolysis And Mitochondrial Respiration In Metabolic Oscillations Of Pancreatic Islets, Richard Bertram, Leslie S. Satin, Morten Gram Pedersen, Dan S. Luciani, Arthur Sherman

Pharmacology and Toxicology Publications

Abstract

Insulin secretion from pancreatic β-cells is oscillatory, with a typical period of 2–7 min, reflecting oscillations in membrane potential and the cytosolic Ca²⁺ concentration. Our central hypothesis is that the slow 2–7 min oscillations are due to glycolytic oscillations, whereas faster oscillations that are superimposed are due to Ca²⁺ feedback onto metabolism or ion channels. We extend a previous mathematical model based on this hypothesis to include a more detailed description of mitochondrial metabolism. We demonstrate that this model can account for typical oscillatory patterns of membrane potential and Ca²⁺ concentration in islets. It also …

A Comparision Of Delta-9-Tetrahydrocannabinol Dependence In C57bl/6j Mice And Fatty Acid Amide Hydrolase Knock Out Mice, Brittany Leigh Alice Carlson

Theses and Dissertations

The idea that humans and laboratory animals can become physically dependent on marijuana or its primary psychoactive constituent, delta-9-tetrahydrocannabinol (THC), is gaining acceptance. However, there are no currently approved pharmacotherapies to treat cannabinoid withdrawal. The objective of this thesis was to evaluate whether elevating endogenous anandamide levels using mice lacking fatty acid amide hydrolase (FAAH), the enzyme responsible for anandamide metabolism, would ameliorate THC dependence. Mice were treated subchronically with a low or high THC dosing regimen and challenged with the CB1 receptor antagonist, rimonabant, to precipitate withdrawal. Following subchronic THC treatment, rimonabant precipitated a significant increase in paw flutters …

The Substituted Pyrrole Jb-03-14 Induces Autophagic Cell Death And Growth Arrest In Breast Tumor Cells, Christopher Ryan Arthur

Theses and Dissertations

The use of chemotherapy in the treatment of cancer has stimulated the demand for better chemotherapeutic agents that are more potent at destroying tumor cell populations and more selective for the specific tumor versus normal host tissues. This project is directed at discovering new anti-tumor agents that are effective against breast cancer based on structures derived from marine organisms, specifically brominated pyrroles. We utilized an in vitro breast cancer model to study the effects of pyrroles on tumor proliferation and survival, as well as growth arrest and cell death. Our findings indicate that the substituted pyrrole JG-03-14 induces time dependent …

Single Nucleotide Polymorphisms In The Folypoly-Gamma-Glutamate Synthetase Gene, Nadine Thompson

Theses and Dissertations

Folic acid is an essential vitamin utilized in the one-carbon metabolism pathway for the synthesis of purine and thymidine nucleotides, which are necessary for cell growth and proliferation. As a result, the enzymes that participate in the metabolism of folic acid have been good targets for cancer chemotherapy. Folylpoly-γ-glutamate synthetase (FPGS) is an enzyme in the folate metabolism pathway that catalyzes the addition of glutamic acid to the naturally occurring folates, thereby allowing the retention of folate cofactors in cells. Similarly, in the case of cancer chemotherapy, antifolates, such as Lometrexol and Tomudex are retained in cells through the activity …

The Influence Of Sustained Cb1 Blockade During Adolescence On Breakpoints In A Progressive-Ratio Paradigm, Mayo Jerry Wright Jr.

Theses and Dissertations

The developmental psychopharmacology of cannabinoids is poorly understood and little is known about the developmental consequences of repeated exposure to cannabinoid antagonists. In these experiments, male Long-Evans rats were treated with SR141716A, a cannabinoid antagonist, throughout adolescence and allowed unrestricted access to food. Control groups were treated with vehicle during the same developmental period and allowed either unrestricted access to food or were pair-fed with a member of the SR-treated group. Motivation to work for food was measured in progressive-ratio sessions at varying levels of food deprivation. For rats that consumed fewer calories throughout adolescence, whether because of pharmacological intervention …

Action Of Tyrosyl Dna Phosphodiesterase On 3'-Phosphoglycolate Terminated Dna Strand Breaks, Haritha Tatavarthi

Theses and Dissertations

Free radical-mediated DNA double strand breaks (DSBs) are induced either directly by ionizing radiation or by certain chemicals like bleomycin. These breaks are terminated by 3'-PG (PO4CH2COOˉ) or 3'-phosphate groups formed as a result of fragmentation of deoxyribose. To study the nature of repair of these 3'-blocked breaks, we constructed substrates mimicking free-radical induced DSBs. Human and yeast tyrosyl DNA-phosphodiesterase (Tdpl) efficiently processed substrates with 3'-PGs, in either the presence or absence of magnesium, to give a 3'-phosphate. Gel filtration chromatography and western blotting codmed that the putative enzyme in human extracts that efficiently processed PG was indeed tyrosyl DNA-phosphodiesterase. …

Relationship Between Cb1 And S1p Receptors In The Central Nervous System, Lauren Michele Collier

Theses and Dissertations

There is significant sequence homology and anatomical co-distribution between cannabinoid (CB1) and sphingosine-1-phosphate (S1P) receptors in the CNS, but potential functional relationships between these lysolipid receptors have not been examined. Therefore, to investigate possible relationships between these two systems at the level of G-protein activation, agonist-stimulated [35S]GTPγS binding and autoradiography were conducted. Autoradiographic studies were first performed to localize receptor-mediated G-protein activation in mouse brain. Coronal brain slices were processed for stimulation of [35S]GTPγS binding using the synthetic cannabinoid agonist WIN 55,212-2 (WIN) or SIP. High levels of WIN- and S1P-stimulated [35S]GTPγS binding were observed in the caudate putamen, hippocampus, …

Immunotoxicological Evaluation Of Critical Windows Of Development Following Exposure To 1,2:5,6 Dibenzanthracene In B6c3f1 Mice, Denise Marie Hernandez

Theses and Dissertations

Numerous findings have suggested that the increased prevalence of childhood illnesses such as cancer, infection, and allergy may be due to environmental exposures. One such cause may be maternal smoking or passive smoke exposure. Known immunotoxicants in cigarette smoke and environmental pollution include polycyclic aromatic hydrocarbons such as 1,2:5,6 Dibenzanthracene (DBA). The objective of these studies was to evaluate the immunosuppressive effects of DBA on various stages of immune system development. Adult mice were administered DBA daily in corn oil at dose levels of 158, 500, 1580, and 5000 µglkg S.C. for 28 days. Immunosuppression was not observed at levels …

Glutamate Excitotoxicty Activates A Novel Calcium Permeable Ion Channel In Cultured Hippocampal Neurons, Laxmikant Sudhir Deshpande

Theses and Dissertations

Glutamate excitotoxicity is the predominant mechanism implicated in neuronal cell death associated with neurological disorders such as stroke, epilepsy, traumatic brain injury and ALS. Excessive stimulation of NMDA subtypes of glutamate receptors leads to protracted intracellular calcium elevations triggering calcium mediated neurotoxic mechanisms culminating in delayed neuronal cell death. In addition, glutamate excitotoxicity induces a NMDA dependent extended neuronal depolarization mediated by continuous calcium influx that correlates with delayed neuronal death. Attempts to prevent neuronal death by blocking calcium entry into the neurons using calcium channel blockers or NMDA receptor antagonists have failed to provide any beneficial effects in clinical …

Influence Of The Vitamin D3 Analog Eb 1089 On Senescence And Cell Death Pathways In The Response Of Breast Tumor Cells To Ionizing Radiation, Gerald Alan Demasters

Theses and Dissertations

A senescence-like growth arrest succeeded by rapid recovery of proliferative capacity is observed in MCF-7 breast tumor cells exposed to fractionated radiation (5 x 2Gy) alone. Exposure to the vitamin D3 analog EB 1089 (100nM) prior to irradiation converts the initial growth arrest response to cell death in part through the inhibition of radiation-induced senescence and promotion of both apoptotic and autophagic cell death. More importantly, EB 1089 was shown to profoundly reduce the rate of recovery following fractionated irradiation. The effect of EB 1089 on the temporal response to radiation is also observed in MCF-7 cells expressing caspase 3, …

Pathological Upregulation Of A Calcium-Stimulated Phosphatase, Calcineurin, In Two Models Of Neuronal Injury, Jonathan Elledge Kurz

Theses and Dissertations

Excitotoxic calcium influx and activation of calcium-regulated systems is a common event in several types of neuronal injury. This mechanism has been the focus of intense research, with the hope that a more complete understanding of how neuronal injury affects calcium-regulated systems will provide effective treatment options. This study examines one such calcium-stimulated enzyme, calcineurin, in the context of two common neurological pathologies, status epilepticus and traumatic brain injury.Status epilepticus was induced by pilocarpine injection. NMDA-dependent increases in calcineurin activity were observed in cortical and hippocampal homogenates. Upon closer examination, the most profound increases in activity were found to be …

The Role Of Gonadal Hormones On Opioid Receptor Protein Density In Arthritic Rats, Matthew Kren

Theses and Dissertations

The majority of research on the endogenous opioid system has focused on various pain assays and the efficacy of different opioid agonists. However, minimal attention has been focused on the effects of gonadal hormones and their impact on the opioid peptide system. The present study was designed to determine the effects of modulation of gonadal hormones on the opioid receptor protein levels in Complete Freund's Adjuvant (CFA)-treated arthritic and non-arthritic male and female Lewis rats. Midbrain and spinal cord tissues were collected for comparison of the μ, δ, and κ receptor protein levels in arthritic and non-arthritic animals. Male gonadectomy …

The Effects Of Footshock On The Reinforcing Efficacy Of Cocaine In Male Long-Evans Rats, Elizabeth S. Hendrick

Theses and Dissertations

Many links exist between cocaine abuse and stress. The literature and laboratory studies in rats suggest that this could be because stress increases the reinforcing efficacy of cocaine. Using male Long-Evans rats, experiments in this thesis tested effects of footshock on the reinforcing efficacy of cocaine using a progressive ratio schedule of reinforcement. They also examined effects of footshock on the reinforcing efficacy of a half-maximal dose of cocaine. Finally, they tested the effects of footshock on cocaine self-administration in rats initially resistant to acquisition of cocaine self-administration. Footshock did not increase reinforcing efficacy of cocaine on a PR schedule …

Post-Juvenile Brain Development Modulates Seizure Characteristics And Diazepam Efficacy In The Rat Pilocarpine-Se Model, William H. Holbert Ii

Theses and Dissertations

These studies were completed to examine how status epilpeticus seizure characteristics are modulated during post-juvenile brain development. This may determine if postnatal age in rats is a better identifier of stages of post-juvenile brain development. The first study fully detailed the acute discrete seizure phase of the rat pilocarpine-SE model. Results for this study showed that Racine behavioral severity score, spike frequency, and seizure severity during the acute discrete seizure phase change in relation to post-juvenile brain developmental stages. The second study fully detailed early and late patterns of status epilepticus. Results for this study displayed modulation of time in …

The Study Of The Effect Of Drugs Of Abuse On Protein Kinase A Activity In Mouse Brain And Spinal Cord, George D. Dalton

Theses and Dissertations

Morphine and Δ9-THC are drugs that produce analgesia and rewarding effects. However, chronic treatment with morphine and Δ9-THC produces problematic side-effects including tolerance and physical dependence. The cellular mechanisms underlying opioid and cannabinoid antinociceptive tolerance have been studied for years. Research has demonstrated that the expression of morphine and Δ9-THC antinociceptive tolerance may be mediated through intracellular signaling pathways, such as the adenylyl cyclase /Protein Kinase A (PKA) cascade. The present study investigated the role of PKA in the expression of morphine and Δ9-THC antinociceptive tolerance. Male Swiss Webster mice were treated chronically with morphine or Δ9-THC and the warm-water …