SPINE Volume 33, Number 26, pp 2863–2867 ©2008, Lippincott Williams & Wilkins Effects of Clotrimazole on Experimental Spinal Cord Ischemia/Reperfusion Injury in Rats Haydar Usul, MD,* Erhan Arslan, MD,* Tufan Cansever, MD,† Umit Cobanoglu, MD,‡ and Su ¨leyman Baykal, MD* Objective. The effect of clotrimazole was examined using a spinal cord ischemia/reperfusion model. Methods. Twenty albino Wistar rats weighing 234  12.3 g were used in this study. Rats were anesthetized intraperitoneally with 50 mg/kg ketamine HCl. All animals underwent laparotomy under aseptic conditions. Abdom- inal aortas of the animals in all but the sham group were exposed. After opening the retroperitoneum, the infrare- nal abdominal aorta was clipped for 45 minutes to pro- duce ischemia/reperfusion injury. Polyethylene glycol (PEG, 1 mL) was administrated to the vehicle group. PEG (1 mL) and clotrimazole (30 mg/kg) were administered intraperitoneally in the clotrimazole group. Total laminec- tomy of T8 –T12 was performed on all rats under a micro- scope. Spinal cords were excised for a length of 2-cm rostrally and 1-cm caudally to the injury site and deep frozen at -76°C for biochemical studies. The levels of malondialdehyde, glutathione-peroxidase, superoxide dismutase, and catalase were measured as an indicator of ischemia level. The most cranial part of the specimens was evaluated morphologically. Results. Treatment with clotrimazole significantly de- creased malondialdehyde, glutathione-peroxidase, su- peroxide dismutase, and catalase levels in comparison with other groups (P = 0.008). Morphologic evaluation revealed that clotrimazole protected the axons and their myelin sheaths from ischemic damage. Conclusion. This study showed the neuroprotective effects of clotrimazole on spinal cord ischemia/reperfu- sion injury. Key words: clotrimazole, ischemia/reperfusion, cata- lase, glutathione peroxidase, malondialdehyde, superox- ide dismutase.Spine 2008;33:2863–2867 Paraplegia is the most challenging complication in oper- ations involving the descending thoracic and thoracoab- dominal aorta. This challenge is encountered, despite re- cent improvements in operation techniques, with an incidence of 4% to 33% in all operations. 1 The etiology of paraplegia is thought to involve anoxia during the period that the aorta is cross-clamped along with dam- age to ascending and descending axons. Due to its high sensitivity to anoxia, the gray matter plays the most im- portant role in the pathophysiology of spinal cord isch- emic injury. Several methods, including hypothermia, cerebrospi- nal fluid drainage, and calcium channel blockers, have been tested to protect the spinal cord against ischemia. 2,3 Most studies have attempted to prevent excessive cal- cium (Ca ++ ) influx into the cells during hypoxia/ ischemia. Ca ++ overload can result in the activation of Ca ++ -dependent proteases, the destruction of mitochon- dria, the activation of nitric oxide synthase, and the gen- eration of free oxygen species. 4 One of the most impor- tant free radical scavengers is clotrimazole, which is also known as a potent antimycotic drug. Numerous studies have shown the radical scavenger effects of clotrimazole in various human cell types. 5 The neuroprotective effect of clotrimazole, through its modulation of Ca ++ trans- port and Ca ++ -dependent intracellular processes, was shown in primary cerebellar cultures and a spinal cord injury model. 5–7 In this study, we aimed to examine the effect of clotrimazole on a spinal cord ischemia/ reperfusion (I/R) model. Materials and Methods Animal Handling Twenty Wistar albino rats weighing 234  12.3 g were used in this study. The animals were kept under constant laboratory conditions of 18°C to 21°C with a 12-hour light-dark cycle and continuous access to food and tap water. All experiments were approved by the Institutional Review Board of Karadeniz Technical University, Faculty of Medicine, and were treated according to the research guidelines. Anesthesia and Surgical Procedure The rats were fasted for 24 hours with free access to water before the surgical procedure. Anesthesia was induced by in- tramuscular administration of 50 mg/kg ketamine hydrochlo- ride (Ketalar, Pfizer, Istanbul). The rats were numbered with ear tags. Their abdomens were shaved and cleaned with 10% polyvinylpyrrolidone–iodine. All animals underwent laparotomy under aseptic condi- tions. No further intervention was applied to the rats in the sham group. The surgical incisions of the rats were closed in layers. The abdominal aortas of the animals in the treatment groups were exposed after opening the retroperitoneum. An aneurysm clip with 50-g closing force (Yasargil FE 693, Aeus- culab, Germany) was applied to the abdominal aorta below the renal artery orifices for a period of 45 minutes. Rats were ran- domly allocated into 4 groups: a sham group of 5 rats in which From the *Department of Neurosurgery, Karadeniz Technical Univer- sity School of Medicine, Trabzon; †Department of Neurosurgery, Gul- hane Military Medical School, Ankara; and ‡Department of Pathol- ogy, Karadeniz Technical University School of Medicine, Trabzon, Turkey. Acknowledgment date: February 11, 2008. Revision date: June 16, 2008. Acceptance date: July 20, 2008. The manuscript submitted does not contain information about medical device(s)/drug(s). No funds were received in support of this work. No benefits in any form have been or will be received from a commercial party related directly or indirectly to the subject of this manuscript. All experiments were approved by the Institutional Review Board of Karadeniz Technical University, Faculty of Medicine, and were treated according to the research guidelines. Address correspondence and reprint requests to Erhan Arslan, MD, Department of Neurosurgery, KTU Tip Fakultesi, Norosirurji AD, 61080 Trabzon, Turkey; E-mail: arserhan@gmail.com 2863