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The Pharmacokinetics and Pharmacodynamics Of The pharmacokinetics and pharmacodynamics of intravenous magnesium sulfate in horses Dissertation Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Stephen A. Schumacher Graduate Program in Comparative and Veterinary Medicine The Ohio State University 2019 Dissertation Committee Alicia L. Bertone, DVM, PhD, Advisor Jeffrey Lakritz, DVM, PhD Phillip Lerche, BVSc, PhD Teresa A. Burns, DVM, PhD Copyrighted by Stephen A. Schumacher 2019 ii Abstract Intravenous administration of magnesium sulfate (MgSO4) is used to calm horses in equestrian competitions, which is contrary to the rules, and there are no current means to regulate its administration. Initial pilot data from magnesium sulfate (MgSO4) administrations identified increases in plasma ionized magnesium (Mg2+) concentrations and changes in the plasma ionized calcium (Ca2+) to Mg2+ concentration ratio. Three experiments were designed to evaluate these changes and to investigate the pharmacokinetics and pharmacodynamics of MgSO4 administration with a goal to identify regulatory methods for its control in horses. The first study investigated the effect the processing, handling, and storage procedures, of the United States Equestrian Federations Equine Drugs and Medications Program, have on Mg2+ and related electrolytes in a sample of 50 normal horses. It was concluded that these procedures could be used to establish regulatory thresholds through the use of electrolyte biomarkers for the administration of MgSO4. The second study detailed the pharmacokinetics of MgSO4 administration and included an investigation of the potential mechanism by which MgSO4 exerts its calming effect. This study examined pharmacodynamic related changes in cardiovascular and fractional excretion variables, as well as, changes in the endocrine profile and electrolyte levels in cerebrospinal fluid as a result of a 60 mg/kg intravenous administration of ii MgSO4. This study did not include a control group as it was not possible to maintain restraint on these intensively instrumented horses for 6 hours without the aid of MgSO4. It was concluded that the intravenous administration of MgSO4 decreased mean arterial blood pressures, increased left ventricular function, increased the fractional excretion of electrolytes, altered the endocrine profile for calcitonin and parathyroid hormone, but did not result in changes to concentrations of Mg2+ in the cerebrospinal fluid. Additionally, this work confirmed the changes in plasma Mg2+ concentrations and the Ca2+ to Mg2+ ratio as seen in the pilot data. In the third study, which was tightly controlled, a novel radiotelemetric approach was used to further elucidate the pharmacodynamics of intravenous MgSO4 administration without the need to for restraint of the horses. This technology was used to document changes in blood pressure, electrocardiograms, and locomotion in response to horses administered 60 mg/kg of MgSO4. We were able to demonstrate a decrease in blood pressure, changes in electrocardiogram conduction intervals, and a decrease in locomotion, all of which could be related to the calming effects of intravenous MgSO4 in horses. Conclusions drawn from these three studies include the identification of regulatory biomarkers capable of determining intravenous MgSO4, a decrease in blood pressure as the likely mechanism for calming, a decrease in activity as a result of administration, and the identification of potential endocrine biomarkers for regulatory control in the form of calcitonin and parathyroid hormone. Additional work is necessary iii and ongoing to further evaluate and develop thresholds for calcitonin and parathyroid hormone. iv Dedication This work is dedicated to my wife Erin and children, Nicholas and Elizabeth, my parents, Richard and Carol Schumacher, for all of their unquestionable love, support and understanding. v Acknowledgments Throughout this process I have been blessed with so many great individuals willing to share their time, talent and experience. I would like to first thank my advisor, Dr. Alicia Bertone for her support, guidance and enthusiasm, and for never giving up on me. I would also like to thank the members of my committee for their advice and support, Drs. Ramiro Toribio, Jeffrey Lakritz, Phillip Lerche and Teresa Burns. I truly appreciate all of the following who answered questions, collected data, motivated and were supportive through it all, and generally put up with me. Dr. Brian Scansen Judith Dutson Dr. Jonathon Yardley Janice Postlethwaite Dr. Robert Hamlin Diana Tartal Dr. William Muir Kari Hollback Dr. Kasia Dembek Sarah Kauffman Dr. Allison Kilbourne Jessica McKinney Dr. Nathalie Reisbig Carl O’Brian Dr. Hayam Hussein Cassy Schumacher Dr. Ahmed Kamr Lindsey Johnson Dr. Mari Kaido Taylor Myers Dr. Jake Swink Travis Brown Dr. Katy Timko Emily McKinley vi Vita Education The Citadel; Military College of South Carolina, Charleston, SC. B.S. Biology, 1993. The Ohio State University, Columbus, OH. DVM Veterinary Medicine, 2003. The Ohio State University, Columbus, OH. PhD-Candidate Graduate Student, 2012-Present Graduate Program in Comparative and Veterinary Medicine Professional Experience USA Equestrian, Lexington, KY Equine Drug Testing Veterinarian 2003-2006 Healthy Pets of Rome-Hilliard, Hilliard, OH Staff Veterinarian 2003-Present United States Equestrian Federation, Lexington, KY Equine Drugs and Medications Program Chief Administrator 2006-Present Publications Foreman, J. H., B. E. Bergstrom, K. S. Golden, J. J. Roark, D. S. Coren, C. R. Foreman, and S. A. Schumacher. "Dose Titration of the Clinical Efficacy of Intravenously Administered Flunixin Meglumine in a Reversible Model of Equine Foot Lameness." [In eng]. Equine Vet J Suppl, no. 43 (Dec 2012): 17-20. Allen, A. K., Schumacher, S. A. (2015). Impact of FEI Rules on Sport Horse Medications in N. E. Robinson (7th Ed.), Robinson's Current Therapy in Equine Medicine. St. Louis, MO: Elsevier. vii Schumacher, S. A., J. Yardley, and A. L. Bertone. "Ionized Magnesium and Calcium Concentration and Their Ratio in Equine Plasma Samples as Determined by a Regulatory Laboratory Compared to a Clinical Reference Laboratory." [In eng]. Drug Test Anal (Sep 25 2018). Fields of Study Major Field: Comparative and Veterinary Medicine viii Table of Contents Abstract ............................................................................................................................... ii Acknowledgments.............................................................................................................. vi Vita .................................................................................................................................... vii List of Tables ..................................................................................................................... xi List of Figures ................................................................................................................... xii Chapter 1. Introduction ....................................................................................................... 1 Chapter 2. Ionized magnesium and calcium concentration and their ratio in equine plasma samples as determined by a regulatory laboratory compared to a clinical reference laboratory ............................................................................................................................ 6 Abstract ........................................................................................................................... 6 Introduction ..................................................................................................................... 7 Materials and Methods .................................................................................................... 9 Statistical Analysis ........................................................................................................ 11 Results ........................................................................................................................... 12 Discussion ..................................................................................................................... 17 Conclusion .................................................................................................................... 22 Chapter 3. Pharmacokinetics and pharmacodynamics associated with experientally induced hypermagnesemia in horses ................................................................................ 24 Abstract ......................................................................................................................... 24 Introduction ................................................................................................................... 25 Materials and Methods .................................................................................................. 27 Statistical Analysis ........................................................................................................ 34 Results ........................................................................................................................... 34 Discussion ..................................................................................................................... 49 Conclusion .................................................................................................................... 55 ix Chapter 4. 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