A Comparison of Daily Rhythm of Creatinine and Creatine Kinase in the Sedentary and Athlete Horse Giuseppe Piccione, Associate Professor, Claudia Giannetto, DVM, PhD, Francesco Fazio, DVM, PhD, Stefania Casella, PhD, and Giovanni Caola, Full Professor ABSTRACT The goal of this study was to investigate the existence of a daily rhythm of creatinine or creatine kinase (CK) and the influence of physical exercise on these rhythms. Blood samples from 20 Sella Italiana horses were col- lected every 4 hours for 48 consecutive hours via an in- travenous cannula inserted into the jugular vein. The horses were divided into two equal groups (Sedentary [S] and Athlete [A]). After 2 weeks of rest, athletes (AR) were subjected again to sample collection every 4 hours for 48 consecutive hours, and sera creatinine and CK were assessed. Statistical analysis of the data was performed by one-way repeated analysis of variance (ANOVA), paired Student’s t-test, unpaired Student’s t-test, and analysis of single cosinor method. Cosinor analysis identified the periodic parameters and their acrophases (expressed in hours) during the 2 days of monitoring. In all horses, we found a highly significant effect of time of the day on the studied parameters (P < .01). Statistically significant differences were observed for mean serum CK values between sedentary and ath- lete horses. The application of the periodic model showed a daily rhythm of the parameters studied in all experimental conditions. The results obtained during the experimental period indicated a daily rhythm of cre- atinine and CK in sedentary and athlete horses and also indicated that physical exercise was able to influence am- plitude, acrophase, and robustness of the creatinine and CK circadian rhythm. Keywords: Daily rhythm; Creatinine; Creatine kinase; Horse; Physical exercise INTRODUCTION The major aspect of chronobiology important to the ath- lete is the cyclic variation in many physiologic variables that contribute to an athlete’s performance. 1-3 Endoge- nous circadian changes in these resting parameters might mediate parallel changes in both performance and physio- logic response to exercise over a 24-hour period. Evening acrophase of serotonin and tryptophan blood levels are useful for the establishment of training programs when tak- ing into account the endogenous rhythm of these two pa- rameters widely involved in the onset of central fatigue. 3 The circadian oscillation of heart rate, blood pressure, and body temperature alters the absolute values of these variables both at rest and during exercise, raising the possi- bility that the safety margins against hyperthermia and hy- potension may decrease during evening exercise. 4 Many circadian rhythms in physiologic functions are evident un- der resting conditions; indeed, most of the body’s systems seem to exhibit circadian rhythmicity. Exercise imposes enormous perturbations on physiologic systems, particu- larly on metabolic, circulatory, hormonal, and thermoreg- ulatory mechanisms. 5-8 Increased serum enzyme activity occurs after exercise, and many factors determine the degree to which the serum activity of enzymes increases during and after exercise. 9 The serum level of the enzyme creatine kinase (CK) is fre- quently used for diagnostic purposes, and circadian varia- tion of this parameter has been observed under different stress conditions. 10 Information about the state of a muscle during exercise can be obtained by the study of CK. High levels of serum CK may be correlated with physical training status in appar- ently healthy subjects, although persistence of high levels at rest may reflect subclinical muscle disease. 11 CK isoenzymes catalyze the reversible phosphorylation of creatine to form creatine phosphate, making adenosine triphosphate avail- able for contraction by the phosporylation of adenosine di- phosphate from creatine phosphate. 12,13 Creatine conversion to creatinine is a nonenzymatic, irreversible pro- cess that occurs at rate of 1.6% to 2% daily in humans. 14,15 The quantity of creatinine formed each day depends on the body content of creatine, which in turn depends on di- etary intake, rate of synthesis of creatine, and muscle mass. ORIGINAL RESEARCH From the Department of Experimental Sciences and Applied Biotechnology, Section of Compared Physiology and Applied Ethology, Laboratory of Veterinary Chronophysiology, Polo Universitario dell’Annunziata, University of Messina, Messina, Italy. Reprint requests: Claudia Giannetto, Dipartimento di Scienze Sperimentali e Biotecnologie Applicate, Sezione di Fisiologia Comparata ed Etologia Applicata, Laboratorio di Cronofisiologia Veterinaria, Universita`degli Studi di Messina, Polo Universitario dell’Annunziata, 98168, Messina, Italy. 0737-0806/$ - see front matter Ó 2009 Elsevier Inc. All rights reserved. doi:10.1016/j.jevs.2009.05.013 Journal of Equine Veterinary Science  Vol 29, No 7 (2009) 575