ROSIGLITAZONE DECREASES PEROXISOME PROLIFERATOR RECEPTOR-GAMMA LEVELS IN MICROGLIA AND INHIBITS TNF-ALPHA PRODUCTION: NEW EVIDENCES ON NEUROPROTECTION IN A PROGRESSIVE PARKINSON’S DISEASE MODEL A. R. CARTA, a * L. FRAU, a A. PISANU, a J. WARDAS, b S. SPIGA c AND E. CARBONI a a Department of Toxicology, University of Cagliari,and Centre ofEx- cellence for Studies on Neurobiology of Addiction,Via Ospedale 72, 09124 Cagliari, Italy b Department of Neuropsychopharmacology, Institute ofPharmacol- ogy, Polish Academy of Sciences, Smetna 12, 31–343 Krakow, Poland c Department of Life and Environment Science, University of Cagliari, Viale Poetto 1, 09126 Cagliari, Italy Abstract—Thiazolidinedione (TZD) class of peroxisome pro- liferator receptor gamma (PPAR- ␥ ) agonists display neuro- protective effects in experimental Parkinson’s disease (PD) models.Neurons and microglia express PPAR- ␥ , therefore both ofthem are potential targets for neuroprotection, al- though the role ofeach celltype is notclear.Moreover, receptor-dependent as well as receptor-independent mecha- nisms have been involved. This study further investigated mechanisms of TZD-mediated neuroprotection in PD. We in- vestigated the rosiglitazone effect in the progressive MPTP/ probenecid (MPTPp) model of PD.C57BL/6J mice received MPTP (25 mg/kg) plus probenecid (100 mg/kg) twice per week for 5 weeks. Rosiglitazone (10 mg/kg) was given daily until sacrifice, starting on the fourth week of MPTPp treatment, in presence of an ongoing neurodegeneration with microglio- sis. Changes in PPAR- ␥ levels were measured by immuno- fluorescence and confocal microscopy in tyrosine hydroxy- lase (TH)-positive neurons and CD11b-positive microglia of the substantia nigra pars compacta (SNc). Chronic MPTPp treatment induced a PPAR- ␥ overexpression in both TH-pos- itive neurons and microglia (139.9% and 121.7% over vehicle, respectively). Rosiglitazone administration to MPTPp-treated mice,reverted PPAR- ␥ overexpression in microglia without affecting TH-positive neurons. Thereafter, changes in CD11b and tumornecrosis factor ␣ (TNF- ␣ ) immunoreactivity in microglia were evaluated in the SNc. MPTPp progressively increased CD11b immunoreactivity, conferring to microglia a highly activated morphology. Moreover, TNF- ␣ levels were increased (457.38% over vehicle) after MPTPp. Rosiglitazone administration counteracted the increase in CD11b immuno- reactivity caused by MPTPp. Moreover, rosiglitazone reverted TNF- ␣ expression to control levels.Nigrostriatal degenera- tion was assessed by high pressure liquid chromatography (HPLC) measurement of striatal dopamine, and counting of TH-positive neurons in the SNc. MPTPp treatment caused a severe decline of striatal dopamine and a partial degenera- tion of the SNc. Rosiglitazone arrested the degenerative pro- cess in both areas. Results suggest that PPAR- ␥ expression in microglia and TNF- ␣ production by these cells are crucial changes by which rosiglitazone exerts neuroprotection in PD. © 2011 IBRO.Published by ElsevierLtd. All rights reserved. Key words: PPAR-gamma, neuroprotection, MPTP,neuroin- flammation, microglia, TNF-alpha. Parkinson’s disease (PD) is a neurodegenerative disorder characterized by the progressive death of dopaminergic neurons of the substantia nigra pars compacta (SNc), resulting in a progressive deficiency of nigrostriatal dopa- mine transmission. Clinical symptoms of PD generally ap- pear when striatal dopamine (DA) levels are largely re- duced and most nigral neurons are lost. Chronic neuroin- flammation plays a crucial role in PD neuropathology, being constantly associated with degenerating neurons (Hirsch and Hunot, 2009; McGeer and McGeer, 2008; Olanow, 2007). In the diseased SNc, microglia are en- gaged in a vicious cycle called reactive gliosis, in which microglia, once activated, may lose their ability to self- modulate. As a result, microglia perpetuate a neurotoxic phenotype of activation, sustaining neurodegeneration through the chronic release of toxic molecules, as the proinflammatory cytokine tumor necrosis factor ␣ (TNF- ␣ ) (Depino et al., 2003; Hirsch and Hunot, 2009). The control of reactive gliosis and the associated production of proin- flammatory factors is therefore a possible target of neuro- protective therapies aimed at stopping or slowing the pro- gression of neurodegeneration. In recent years, agonists of the peroxisome prolifera- tor-activated receptor gamma (PPAR- ␥ ) belonging to the thiazolidinedione (TZD) class have received increasing at- tention for its therapeutic potential in neurodegenerative disorders, including PD ( Bernardo and Minghetti, 2008; Carta et al., 2011; Chaturvedi and Beal, 2008; Heneka et al., 2007; Kiaei et al., 2005; Luo et al., 2006; Park et al., 2007; Zheng et al., 2010 ). We and others have shown that TZDs rosiglitazone and pioglitazone can prevent 1-methyl- 4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced do- paminergic degeneration and development of motor symp- toms, though mechanisms of neuroprotection are still elu- sive (Breidert et al., 2002; Dehmer et al., 2004; Schintu et al., 2009a). PPAR- ␥ are ligand-inducible transcription fac- tors belonging to the superclass of nuclear hormone re- ceptors. The highest level of expression is shown by adi- *Corresponding author. Tel: ⫹39-0706758662; fax: ⫹39-0706758665. E-mail address: acarta@unica.it (A. R. Carta). Abbreviations: DA, dopamine; DOPAC, 3,4-dihydroxyphenylacetic ac- id; TZD, thiazolidinedione; MPTPp, MPTP/probenecid; PD, Parkin- son’s disease; PPAR- ␥ , proliferator-activated receptor gamma; TH, tyrosine hydroxylase; SNc, substantia nigra pars compacta. Neuroscience 194 (2011) 250 –261 0306-4522/11 $ - see front matter © 2011 IBRO. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.neuroscience.2011.07.046 250