Dopamine D 1 receptor deletion strongly reduces neurotoxic effects of methamphetamine S. Ares-Santos a, b, 1 , N. Granado a, c, 1 , I. Oliva d , E. O'Shea c , E.D. Martin d , M.I. Colado c , R. Moratalla a, b, ⁎ a Instituto Cajal, Consejo Superior de Investigaciones Científicas, CSIC, 28002, Madrid, Spain b CIBERNED, Instituto de Salud Carlos III, Madrid, Spain c Departamento de Farmacología, Facultad de Medicina, Universidad Complutense de Madrid, 28040 Madrid, Spain d Laboratorio de Neurofisiología y Plasticidad Sináptica, Instituto de Investigación en Discapacidades Neurológicas (IDINE), Universidad de Castilla-La Mancha, Albacete, Spain abstract article info Article history: Received 5 September 2011 Revised 31 October 2011 Accepted 7 November 2011 Available online 13 November 2011 Keywords: Dopamine toxicity TH DAT Glia Amphetamine derivatives Striatum Substantia nigra Drug-addiction Parkinson's disease Dopamine overflow Methamphetamine (METH) is a potent, highly addictive psychostimulant consumed worldwide. In humans and experimental animals, repeated exposure to this drug induces persistent neurodegenerative changes. Damage occurs primarily to dopaminergic neurons, accompanied by gliosis. The toxic effects of METH involve excessive dopamine (DA) release, thus DA receptors are highly likely to play a role in this process. To define the role of D 1 receptors in the neurotoxic effects of METH we used D 1 receptor knock-out mice (D 1 R -/- ) and their WT lit- termates. Inactivation of D 1 R prevented METH-induced dopamine fibre loss and hyperthermia, and increases in gliosis and pro-inflammatory molecules such as iNOS in the striatum. In addition, D 1 R inactivation prevented METH-induced loss of dopaminergic neurons in the substantia nigra. To explore the relationship between hyper- thermia and neurotoxicity, METH was given at high ambient temperature (29 °C). In this condition, D 1 R -/- mice developed hyperthermia following drug delivery and the neuroprotection provided by D 1 R inactivation at 23 °C was no longer observed. However, reserpine, which empties vesicular dopamine stores, blocked hyperthermia and strongly potentiated dopamine toxicity in D 1 R -/- mice, suggesting that the protection afforded by D 1 R in- activation is due to both hypothermia and higher stored vesicular dopamine. Moreover, electrical stimulation evoked higher DA overflow in D 1 R -/- mice as demonstrated by fast scan cyclic voltammetry despite their lower basal DA content, suggesting higher vesicular DA content in D 1 R -/- than in WT mice. Altogether, these re- sults indicate that the D 1 R plays a significant role in METH-induced neurotoxicity by mediating drug-induced hy- perthermia and increasing the releasable cytosolic DA pool. © 2011 Published by Elsevier Inc. Introduction Methamphetamine (METH), a synthetic derivative of amphetamine, is a psychostimulant with high addictive potential used by between 13.7 and 52.9 million people world-wide (UNODC, World Drug Report, 2010). METH is a known neurotoxin, causing damage primarily to the dopaminergic system in all species studied: monkeys (Seiden et al., 1976), rodents (Granado et al., 2010, 2011a,b; Krasnova et al., 2011) and humans (McCann et al., 1998; Volkow et al., 2001). In mice, repeated exposure to METH causes persistent neurotoxicity to dopami- nergic terminals and neurons, evidenced by reduced tyrosine hydroxy- lase (TH) (Bowyer et al., 2008; Deng et al., 1999; O'Callaghan and Miller, 1994; Xu et al., 2005; Zhu et al., 2005) and dopamine transporter (DAT) levels (Achat-Mendes et al., 2005; Deng et al., 1999; Fumagalli et al., 1999) in the striatum. In addition, this drug induces neuronal death in the striatum, which occurs by a process resembling neuronal apoptosis (Cadet and Krasnova, 2009; Cadet et al., 2005, 2007). METH also causes cell body loss in the substantia nigra (Granado et al., 2011a,b; Sonsalla et al., 1996), affecting the same nigrostriatal dopaminergic neurons that undergo selective degeneration in Parkin- son's disease (Granado et al., 2010). We have recently shown that METH selectively damages the nigrostriatal pathway, sparing the mesolimbic dopaminergic pathway, and that the striosomes are more sensitive than the striatal matrix, as indicated by greater TH/ DAT-immunoreactivity loss (Granado et al., 2010). METH also causes reactive astrocytosis and microgliosis (Cadet and Krasnova, 2009; Fantegrossi et al., 2008; O'Callaghan and Miller, 1994; Thomas et al., 2004, 2008a), providing additional evidence for neuronal injury. Although the exact molecular mechanisms of METH-induced do- paminergic neurotoxicity are not established, dopamine itself appears to play a significant role (Albers and Sonsalla, 1995; Thomas et al., 2008b). Specifically, it has been suggested that newly synthesized DA in the cytoplasmic pool (Thomas et al., 2008b) can be metabolized via auto-oxidation to produce DA quinones, superoxide anions and hydrogen oxygen species, with subsequent generation of oxidative stress, mitochondrial dysfunction, and damage within dopaminergic Neurobiology of Disease 45 (2012) 810–820 ⁎ Corresponding author at: Instituto Cajal, CSIC, Avd. Dr. Arce 37, 28002, Madrid, Spain. Fax: + 34 91 585 4754. E-mail address: moratalla@cajal.csic.es (R. Moratalla). 1 Contributed equally in this work. Available online on ScienceDirect (www.sciencedirect.com). 0969-9961/$ – see front matter © 2011 Published by Elsevier Inc. doi:10.1016/j.nbd.2011.11.005 Contents lists available at SciVerse ScienceDirect Neurobiology of Disease journal homepage: www.elsevier.com/locate/ynbdi