Neuroprotective effects of TEMPOL in central and peripheral nervous system models of Parkinson’s disease Qinghua Liang a , Amanda D. Smith b , Stephen Pan a , Vladimir A. Tyurin c , Valerian E. Kagan c , Teresa G. Hastings b , Nina Felice Schor a,b, * a Department of Pediatrics, University of Pittsburgh, Pittsburgh, PA, USA b Department of Neurology, University of Pittsburgh, Pittsburgh, PA, USA c Department of Environmental and Occupational Health, University of Pittsburgh, Pittsburgh, PA, USA Received 30 December 2004; accepted 14 April 2005 Abstract TEMPOL (4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl) is a stable nitroxyl antioxidant. Previous studies have suggested that TEMPOL is protective in acute disorders thought to involve reactive oxygen species (ROS), such as ischemic stroke and cardiac reperfusion injury. Oxidized TEMPOL can be recycled to its redox-active reducing form by co-administration with polynitroxylated albumin, making it a candidate as a pharmacological ‘‘reservoir’’ for reducing potential of use in chronic disorders involving ROS. The present studies examine the efficacy of TEMPOL in cell culture and animal models of the central and peripheral dysfunction associated with Parkinson’s disease, a disorder in the pathogenesis of which ROS generated from dopamine have been implicated. Antioxidants have been proposed as both preventive and symptomatic therapy for Parkinson’s disease. TEMPOL protects MN9D dopaminergic mesencephalic cells in culture from 6-hydroxydopamine (6-OHDA)-induced apoptosis. Translocation of the p65 component of NF- kB to the nucleus accompanies protection by TEMPOL. In vivo, intraperitoneal TEMPOL protects mice from intrastriatal 6-OHDA- induced cell and dopamine metabolite loss in the striatum. TEMPOL also protects mice against the 6-OHDA-induced rotational behavior elicited by intrastriatal administration of D-amphetamine. In addition, TEMPOL protects mice from the ptosis, activity level decrement, and mortality induced by intraperitoneal administration of 6-OHDA, a model of autonomic dysfunction in Parkinson’s disease. Adjunctive use of polynitroxylated albumin enhances the in vitro and in vivo effects of TEMPOL. # 2005 Elsevier Inc. All rights reserved. Keywords: Parkinson’s disease; Reactive oxygen species; Antioxidants; Neuroprotection; Dopamine; Autonomic nervous system 1. Introduction Parkinson’s disease is a neurodegenerative disorder that primarily affects dopaminergic neurons in the central nervous system. Peripheral neurons, particularly those in the autonomic nervous system, can be affected as well [1,2]. The tendency of dopamine to oxidize forming reac- tive oxygen species (ROS) has been proposed to be a component of the pathogenesis of Parkinson’s disease. Under normal circumstances, endogenous antioxidant mechanisms serve to detoxify those ROS formed along with neuromelanin in dopaminergic neurons. When these mechanisms fail, it has been hypothesized and demon- strated in cell culture and animal model systems, these dopaminergic neurons are injured and die, often by apop- tosis [3–8]. Both abortive and preventive approaches to Parkinson’s disease have included the use of antioxidant drugs. Oxygen radical scavengers and reducing agents, like Vitamins C and E, have not, however, shown unequivocal promise in this regard ([9,10]; reviewed in [11]). It has been suggested that this is because, although they rapidly reduce ROS, themselves becoming oxidized, their redox potential is such that under physiological conditions, they are not readily re-reduced to their active scavenging form [12–14]. That is, www.elsevier.com/locate/biochempharm Biochemical Pharmacology 70 (2005) 1371–1381 Abbreviations: TEMPOL, 4-hydroxy-2,2,6,6-tetramethylpiperidine-N- oxyl; PNA, polynitroxylated albumin; DOPAC, dihydroxyphenylacetic acid; HVA, homovanilic acid; TH, tyrosine hydroxylase; MTS, 3-(4,5- dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H- tetrazolium; i.p., intraperitoneal; i.v., intravenous * Corresponding author at: Children’s Hospital of Pittsburgh, Pediatric Center for Neuroscience, 3460 Fifth Avenue, Pittsburgh, PA 15213, USA. Tel.: +1 412 692 6182; fax: +1 412 692 6787. E-mail address: nfschor@pitt.edu (N.F. Schor). 0006-2952/$ – see front matter # 2005 Elsevier Inc. All rights reserved. doi:10.1016/j.bcp.2005.04.011