MOLECULAR BASES OF METHAMPHETAMINE-INDUCED NEURODEGENERATION

Abstract

Methamphetamine (METH) is a highly addictive psychostimulant drug, whose abuse has reached epidemic proportions worldwide. The addiction to METH is a major public concern because its chronic abuse is associated with serious health complications including deficits in attention, memory, and executive functions in humans. These neuropsychiatric complications might, in part, be related to drug-induced neurotoxic effects, which include damage to dopaminergic and serotonergic terminals, neuronal apoptosis, as well as activated astroglial and microglial cells in the brain. Thus, the purpose of the present paper is to review cellular and molecular mechanisms that might be responsible for METH neurotoxicity. These include oxidative stress, activation of transcription factors, DNA damage, excitotoxicity, blood–brain barrier breakdown, microglial activation, and various apoptotic pathways. Several approaches that allow protection against METH-induced neurotoxic effects are also discussed. Better understanding of the cellular and molecular mechanisms involved in METH toxicity should help to generate modern therapeutic approaches to prevent or attenuate the long-term consequences of psychostimulant use disorders in humans.

I. Epidemiology of Methamphetamine Abuse

Methamphetamine (METH) is an illegal psychostimulant that is abused by more than 25 million people in the world, which exceeds the amount of people who use heroin and cocaine (Rawson and Condon, 2007; Romanelli and Smith, 2006). METH is the most commonly synthesized illegal drug in the United States and has been cited by law enforcement officials as the leading crime problem in the country (Gettig et al., 2006). The inexpensive production of the drug, its low cost, and its long duration of action make it very desirable to abusers. Recently, METH abuse has reached epidemic proportions in the United States, with populations in the Western, Southern, and Midwest states being the most affected (Gettig et al., 2006; Rawson and Condon, 2007; Topolski, 2007). A 2003 survey found that 5.2% of the American adults have used METH at least once (Roehr, 2005). In addition, 5.3% and 6.2% of high school sophomores and seniors have tried METH (Gettig et al., 2006). METH-related emergency room admissions have also increased from 10 to 52 per 100,000 people between 1992 and 2002 (Roehr, 2005). Although METH abuse has been associated, traditionally, with blue-collar construction workers, truck drivers, and motorcycle gangs, the profile of the typical METH abusing individual has shifted due to the increased popularity among college students, women, and young professionals (Gettig et al., 2006; Romanelli and Smith, 2006). METH use has also highly increased in men who have sex with men resulting in a greater frequency of METH abuse in homosexual and bisexual men than in the general population (Shoptaw, 2006).

II. Clinical Toxicology of METH Abuse

METH abuse causes serious health complications in humans. METH users develop acute clinical symptoms that include agitation, anxiety, aggressive behaviors, paranoia, hypertension, hyperthermia, and psychosis (Albertson et al., 1999; Lynch and House, 1992; Murray, 1998). Ingestions of large doses of the drug can cause life-threatening hyperthermia above 41 °C, cardiac arrythmias, heart attacks, cerebrovascular hemorrhages, strokes, seizures, renal, and liver failure (Albertson et al., 1999; Perez et al., 1999).

Clinical studies have also shown the potential neuropathological consequences of METH abuse, which include deficits in attention, working memory, and executive functions in chronic psychostimulant abusers (Gonzalez et al., 2004; Rippeth et al., 2004; Salo et al., 2002; Sim et al., 2002; Simon et al., 2002, 2004; Woods et al., 2005). The drug can also cause neurodegenerative changes in the brains of human addicts. These include persistent loss of dopamine transporters (DAT) observed in the cortex and caudate-putamen (Sekine et al., 2003; Volkow et al., 2001), loss of serotonin transporters (5-HTT) in the cortex (Sekine et al., 2006), and decrease in the levels of dopamine (DA) and its metabolites in the caudate-putamen of METH abusers (Moszczynska et al., 2004; Wilson et al., 1996).

In agreement with clinical findings, a number of animal studies have shown that METH can cause long-term damage to presynaptic dopaminergic and serotonergic axons in rodents (Ricaurte et al., 1980; Wagner et al., 1980). More recently, it has also been shown that the drug can cause death of the neuronal bodies in the brain through apoptosis (Deng and Cadet, 1999, 2000; Eisch et al., 1998; O’Dell and Marshall, 2000; Zhu et al., 2006). In what follows, we discuss some of the mechanisms that might underlie these METH-induced neurodegenerative effects.

III. Role of Oxidative Stress in METH Toxicity

Toxic effects of METH are thought to depend on the similarity of its chemical structure to DA, which allows the drug to enter DA axons (Iversen, 2006), followed by DA release from synaptic vesicles into cytoplasm and by reverse transport into the synaptic cleft (Sulzer et al., 2005). METH neurotoxicity depends on the formation of DA quinones and superoxide radicals within nerve terminals (LaVoie and Hastings, 1999). DA metabolism by MAO can also increase hydrogen peroxide production, followed by its interactions with metal ions to form very toxic hydroxyl radicals (Cadet and Brannock, 1998). Evidence has accumulated to indicate that METH can also cause oxidative stress by switching the balance between ROS production and the capacity of antioxidant enzyme system to scavenge ROS (Gluck et al., 2001; Harold et al., 2000; Jayanthi et al., 1998). Excessive production of ROS that overwhelms this system can damage cellular components such as lipids, proteins, mitochondrial and nuclear DNA (Potashkin and Meredith, 2006).

A role for oxidative mechanisms in the drug toxicity is consistent with findings that pretreatment with N-acetyl-l-cysteine, ascorbic acid, or vitamin E allows protection against METH-induced depletion of monoaminergic axons (De Vito and Wagner, 1989; Fukami et al., 2004; Wagner et al., 1985). The role for superoxide radicals in the neurotoxic effects of METH on DA axons was tested by injecting METH to transgenic mice that overexpress the human CuZn superoxide dismutase (CuZnSOD) gene (Cadet et al., 1994; Hirata et al., 1996; Jayanthi et al., 1998). These mice have much higher CuZnSOD enzyme activity than control wild-type animals (Jayanthi et al., 1998, 1999) and were protected against METH toxicity. In addition, bromocriptine, which scavengers hydroxyl radicals, also attenuates METH-induced DA depletion in mice (Kondo et al., 1994). Together, these findings support the proposition that DA release caused by METH is accompanied by redox cycling of DA quinones and formation of superoxide radicals. The hypothesis that oxygen-based free radicals are involved in METH toxicity (Cadet and Brannock, 1998) is also supported by reports that the drug can reduce the levels of glutathione (Harold et al., 2000) and antioxidant enzymes (Jayanthi et al., 1998), increase lipid peroxidation (Gluck et al., 2001; Jayanthi et al., 1998), and cause the formation of protein carbonyls (Gluck et al., 2001).

Recent evidence also indicates that changes in nitric oxide (NO) metabolism can contribute to METH-induced oxidative stress and neurotoxicity (Itzhak and Ali, 2006). NO can react with superoxide radicals to form peroxynitrite, a strong oxidant and major neurotoxin (Pacher et al., 2007). Indeed, increased levels of a marker for peroxynitrite production 3-NT have been measured in vivo and in vitro in response to METH treatment (Imam et al., 2001a). Antioxidants selenium and melationin completely block the formation of 3-NT and striatal DA depletion (Imam et al., 2001a). In addition, free radical scavenger, edaravone, blocked METH-related increase in 3-NT immunoreactivity with subsequent attenuation of DA depletion and of reduction in TH immunoreactivity in the striatum (Kawasaki et al., 2006). Furthermore, the selective neuronal nitric oxide synthase (nNOS) inhibitor, 7-nitroindazole, protects against drug-induced formation of 3-NT, as well as DA and 5-HT depletion in the striatum (Ali and Itzhak, 1998; Di Monte et al., 1996; Itzhak and Ali, 1996). The participation of NO in METH toxicity is also proposed by findings that nNOS knockout mice were protected against the formation of 3-NT and DA terminal degeneration in the striatum (Imam et al., 2001b). Together, these data strongly support the hypothesis that NO and peroxynitrite, in particular, are involved in the mechanisms underlying METH-induced monoaminergic neurotoxicity (Imam et al., 2001b; Itzhak and Ali, 2006; Itzhak et al., 1998).

IV. Involvement of AP-1-Related Transcription Factors in METH-Induced Neurotoxicity

The accumulated data suggested that effects of METH might be mediated, in part, by activation of AP-1 transcription factors. These include upregulation of c-jun, c-fos, jun B, and jun D expression within 2 h after METH administration (Cadet et al., 2001). These changes, in turn, might be related to METH-induced generation of ROS. Specifically, hydroxyl and superoxide radicals can induce the expression of many genes via regulation of AP-1 transcription factors (Dalton et al., 1999). The role for c-fos in METH-induced neuropathological changes has been confirmed by using c-fos heterozygote mice that show increased degeneration of DA axons and increased cell death after psychostimulant treatment (Deng et al., 1999). These findings support a protective role for c-fos against METH damage. The factors that could be involved in this protection include cell adhesion receptors integrins because of decreased basal levels of integrin expression in c-fos heterozygote mice and the further reduction of these receptors in response to toxic doses of METH (Betts et al., 2002). This idea is also supported by the observations that integrins promote cell survival after injury and apoptotic insults via PI3K-Akt pathway which leads to phosphorylation of proapoptotic protein BAD, therefore, reducing its ability to block the antiapoptotic effects of Bcl-2 (Gilcrease, 2007). In contrast, inhibition of integrins increases apoptotic cell death (Gilcrease, 2007).

c-jun is another AP-1 transcription factor that might be involved in METH toxicity, because c-jun knockout mice show partial protection against damaging effects of the drug (Deng et al., 2002b). Moreover, because the c-jun knockout mice and their wild-type littermates show similar degree of dopaminergic toxicity after METH treatment, c-jun appears to only play role in the mediation of neuronal apoptosis in cells postsynaptic to DA axons.

V. Role of DNA Damage in METH-Induced Toxicity

As mentioned earlier, METH can cause neuronal apoptosis in several brain regions, including striatum, cortex, hippocampus, and olfactory bulb (Deng et al., 2001). Because apoptosis is associated with DNA damage, it was possible that treatment with METH might induce responses involved in the repair of the drug-related DNA damage. Data obtained using microarray analyses have shown that METH administration caused changes in the expression of a number of genes that participate in DNA repair, including APEX, PolB, and LIG1 (Cadet et al., 2002). These changes are probably related to METH-induced increase in the levels of free radicals because oxidative stress can cause single and double DNA strand breaks (Li and Trush, 1993). Thus, the upregulation of DNA repair genes following METH treatment suggests that these changes might be compensatory to counteract METH-related ROS-induced DNA damage. If the psychostimulant can cause similar DNA damages in humans, this might account for developmental deficits observed in children born of METH abusing mothers (Smith et al., 2006).

VI. METH Toxicity and Excitotoxicity

METH neurotoxicity might also occur via excitotoxic damage following glutamate release and activation of glutamate receptors. Glutamate toxicity is dependent, in part, on the production of NO (Chung et al., 2005). The hypothesis on the involvement of glutamate in METH toxicity is supported by findings that METH causes glutamate release in the brain (Abekawa et al., 1994; Baldwin et al., 1993; Mark et al., 2004; Marshall et al., 1993; Nash et al., 1988). In addition, some glutamate antagonists can attenuate METH-induced dopaminergic toxicity (Battaglia et al., 2002; Sonsalla et al., 1989). Glutamate-mediated NO formation might also be involved in METH toxicity because knockout mice deficient in either nNOS or iNOS (inducible nitric oxide synthase) are protected against psychostimulant-induced damage to monoaminergic axons (Itzhak et al., 1998). These data have provided strong support for the idea that the glutamate/NO pathway plays major role in METH neurotoxicity (Imam et al., 2001b; Itzhak and Ali, 2006; Itzhak et al., 1998). Finally, various nNOS inhibitors can also protect against depletion of monoaminergic axons caused by METH administration (Itzhak et al., 2000; Sanchez et al., 2003). In addition to their roles in the damage of monoaminergic axons, oxygen-based radicals and NO may be involved in METH-related neuronal death because CuZnSOD transgenic mice show partial protection against drug-induced apoptosis (Deng and Cadet, 2000).

VII. Role of Blood–Brain Barrier Dysfunction in METH Toxicity

Several recent papers have examined the effects of METH on the blood–brain barrier (BBB) and their potential relationships to METH toxicity (Bowyer and Ali, 2006; Bowyer et al., 2008; Kiyatkin et al., 2007; Sharma and Ali, 2006; Sharma et al., 2007). Using protein tracers and albumin immunohistochemistry, METH was shown to cause marked disruption of BBB at the levels of the cortex, hippocampus, thalamus, hypothalamus, cerebellum, and amygdala (Bowyer and Ali, 2006; Kiyatkin et al., 2007; Sharma et al., 2007). METH-induced BBB breakdown was evidenced by leakage of serum albumin into the brain tissue (Sharma et al., 2007). Doses of METH that cause BBB disturbances also induce neuronal damage, myelin degeneration, and reactive astrocytosis in the parietal and occipital cortices (Sharma et al., 2007). These doses also cause extensive degeneration of pyramidal cells and activation of microglia in amygdala and hippocampus of rats (Bowyer and Ali, 2006). These effects appear to depend on hyperthermia because the psychostimulant failed to induce BBB damage and neurodegeneration in the brains of animals that did not show increased temperature (Bowyer and Ali, 2006). Interestingly, mild BBB dysfunction found in the caudate-putamen after METH treatment was exacerbated by hyperthermia (Bowyer et al., 2008). It is interesting to point out that METH-induced leakage of serum albumin into brain tissue was attenuated by pretreatment with antioxidant, H-290/51, suggesting the involvement of free radicals in BBB damage (Sharma et al., 2007) and further supporting a role for oxidative stress in drug toxicity. Also of interest is the fact the antioxidant was able to attenuate METH-induced hyperthermia, neuronal damage, myelin degradation, and glial response (Sharma et al., 2007).

VIII. Involvement of Mitochondrial Death Pathway in METH-Induced Apoptosis

In addition to glutamate and NO, the Bcl-2 family of proteins may also be involved in the mechanisms underlying METH neurotoxicity (Cadet et al., 2001; Jayanthi et al., 2001; Stumm et al., 1999). In particular, METH induced increases in proapoptotic proteins, BAX and BID, and decreases in antiapoptotic proteins, Bcl-2 and Bcl-X_L. The upregulation of proapoptotic proteins is consistent with findings that METH treatment caused release of mitochondrial proteins cytochrome c and apoptosis inducing factor (AIF) into the cytosol (Deng et al., 2002a; Jayanthi et al., 2004). AIF and Smac/DIABLO released from mitochondria have also been shown to participate in METH-induced apoptosis (Jayanthi et al., 2004). Their release is followed by activation of caspases 9 and 3, and the breakdown of several structural cellular proteins (Jayanthi et al., 2004). Thus, these findings implicate the mitochondrial pathway in METH-related neuronal death in the brain (Cadet et al., 2005, 2007). This idea is consistent with the findings that overexpression of Bcl-2 can protect against drug-induced apoptosis (Cadet et al., 1997).

IX. Involvement of the Endoplasmic Reticulum-Dependent Death Pathway in METH-Induced Apoptosis

Oxidative stress can trigger cellular damage by causing dysfunctions of cellular organelles such as the endoplasmic reticulum (ER) (Gorlach et al., 2006). In addition to regulating synthesis, folding, and transport of proteins, ER also constitutes the main intracellular store for Ca²⁺, whose excess can contribute to cell death (Gorlach et al., 2006). At physiological levels, Ca²⁺ released from the ER is taken up by mitochondria to enhance metabolite flow on the outer mitochondrial membrane and to increase ATP production (Kroemer et al., 2007). However, sustained release of Ca²⁺ from the ER stores may initiate calcium-dependent apoptosis via the permeabilization of the outer mitochondrial membrane (Kroemer et al., 2007). ER stress and dysregulation of calcium homeostasis appear to participate in METH-induced cell death because the drug can induce activation of calpain (Jayanthi et al., 2004), a calcium-responsive cytosolic protease involved in ER-dependent apoptosis (Nakagawa and Yuan, 2000). METH has been shown to increase calpain-mediated protolysis of cytoskeletal protein spectrin and microtubule protein tau in the rat cortex, hippocampus (Warren et al., 2005), and striatum (Staszewski and Yamamoto, 2006). In contrast, the calpain inhibitors can attenuate psychostimulant-induced spectrin and tau proteolysis (Warren et al., 2007) as well as neuronal death (Samantaray et al., 2006), strongly implicating ER stress and calpain activation in the mechanisms of METH neuronal degeneration. A role for the ER in METH toxicity is further supported by the findings that apoptotic doses of the drug can increase the expression of proteins such as caspase-12, GRP78/BiP, and CHOP/GADD153 (Jayanthi et al., 2004) that participate in ER-induced apoptosis (Marciniak and Ron, 2006). This METH-related ER stress might be secondary to oxidative stress (Cadet and Brannock, 1998; Cadet et al., 1994; Jayanthi et al., 1998) and to increases in BAX/Bcl-2 ratios induced by this illicit drug (Jayanthi et al., 2001).

X. Microglial Reactions and METH Toxicity

Microglia are the resident immune cells within CNS that function to protect the brain against injury and chemical damage (Raivich, 2005). In the healthy mature brain, microglia typically exist in a resting state characterized by ramified morphology, and monitor the neuronal environment (Block et al., 2007; Raivich, 2005). However, in response to brain injury or damage, microglia are readily activated, undergoing a dramatic transformation from a resting ramified state into an amoeboid morphology. Although microglial activation is necessary for host defense and neuron survival, the overactivation of microglia results in deleterious and neurotoxic consequences. Specifically, microglia contributes to the progress of many neurodegenerative diseases, including Parkinson’s (Kim and Joh, 2006), Alzheimer’s (Xiang et al., 2006), and Huntington (Sapp et al., 2001) diseases, AIDS-related neuropathy (Gonzalez-Scarano and Martin-Garcia, 2005) as well as in the neurotoxic effects of MPTP (Gao et al., 2002) and kainic acid (Chen et al., 2005). Once activated, microglia become big, migrate to the site of the injury, and cause phagocytosis of dying and dead cells. In addition, microglia secrete a variety of cytokines, reactive oxygen and nitrogen species, and prostaglandins that are known to cause neuronal damage (Block et al., 2007; Perry et al., 2007).

Recently, emerging data have implicated microglial activation as an early event in the neurotoxic cascade that is initiated by METH treatment. Specifically, METH causes strong microglial response in the areas of the brain that show DA axonal degeneration (Thomas et al., 2004b). In contrast, attenuation of METH neurotoxicity by MK-801 and dextromethorphan inhibits microglial activation (Thomas and Kuhn, 2005). Moreover, anti-inflammatory drug ketoprofen allows some protection against METH toxicity and also reduces psychostimulant-induced microgliosis (Asanuma et al., 2003). However, attenuation of microglial activation itself is insufficient to protect against METH neurotoxicity (Sriram et al., 2006b). This microglial activation precedes METH-induced DA axonal degeneration in the striatum, suggesting that microglia might contribute to drug toxicity (LaVoie et al., 2004).

While neurotoxic amphetamines METH, MDMA, amphetamine and p-chloroamphetamine cause microglial activation (Thomas et al., 2004a) in addition to reactive astrocytosis (Deng and Cadet, 1999; Krasnova et al., 2005; O’Callaghan et al., 1995; Xu et al., 2005), nonneurotoxic drugs such as fenfluramine and DOI fail to activate microglia (Thomas et al., 2004a). These data establish a link between the neurotoxic amphetamines and microglial activation, suggesting that microglia might be a selective marker for neuronal axonal damage (Thomas et al., 2004a) in agreement with the ability for the same drugs to cause reactive astrocytosis, an established hallmark of neurotoxicity (O’Callaghan and Sriram, 2005).

Microglial cells might potentiate METH-related damage by releasing toxic substances such as superoxide radicals and NO which have already been implicated in drug neurotoxicity (see discussion above). In addition, METH causes increase in the levels of TNF-α and IL-β (Flora et al., 2002; Sriram et al., 2006a), proinflammatory cytokines that can also contribute to toxicity of the drug. Consistent with these findings, METH neurotoxicity and an increase in a marker for microglial activation PK11195 binding were attenuated in IL-6 null mice (Ladenheim et al., 2000). Together, these observations show that inflammatory reactions in microglia might participate in the molecular pathway underlying METH toxicity in the brain.

A schematic diagram showing molecular mechanisms that lead to METH-induced neuronal degeneration is presented in Fig. 1.

FIG. 1.

Mechanisms implicated in METH-induced neurotoxicity. The figure summarizes the pathways that are reviewed in this chapter.

XI. Neuroprotective Mechanisms and METH Toxicity

Several attempts have been made to identify ways to protect the brain against METH toxicity. DA uptake inhibitors and DA receptor antagonists have been shown to provide protection against METH-induced degeneration of striatal DA terminals (Angulo et al., 2004; Jayanthi et al., 2005; Marek et al., 1990; O’Dell et al., 1993; Schmidt and Gibb, 1985; Sonsalla et al., 1986). The DA D1 antagonist, SCH23390, also protects against METH-induced cell death in the striatum (Jayanthi et al., 2005). These neuroprotective effects may depend on changes in DA release because DA receptor antagonists, SCH23390 and eticlopride, were reported to partially block METH-related increases in DA release (O’Dell et al., 1993). DA D1-depending mechanisms are also mediated via Fas/FasL-related events since prior treatment with SCH23390 decreased translocation of NFATc3 and NFATc4 from the cytoplasm to the nucleus and reduced increases in calcineurin expression caused by METH administration (Jayanthi et al., 2005). Pretreatment with SCH23390 also resulted in significant inhibition of the METH-induced increases in the expression of FasL and caspase-3 in rat striatal cells (Jayanthi et al., 2005). Because the dose of SCH23390 used in that study completely blocks METH-induced decreases in DA levels while providing only partial protection against death of striatal neurons (Jayanthi et al., 2005), the possibility exists that METH-induced cell death might involved additional mechanisms independent of stimulation of DA D1 receptors.

In addition to DAT inhibitors and DA receptor antagonists, some trophic factors also provide protection against the toxic effects of METH in vitro (Chou et al., 2008; Mamounas et al., 1995; Matsuzaki et al., 2004; Zhou et al., 2004) and in vivo (Cass, 1996; Chou et al., 2008; Melega et al., 2000). Specifically, brain-derived neurotrophic factor (BDNF) was shown to prevent METH-induced caspase-3 activation and death in primary cultures of cortical neurons (Matsuzaki et al., 2004). The protective effect was blocked by phosphatydilinositol-3-kinase inhibitors and by the expression of kinase-deficient Akt, thus implicating the PI3K/Akt pathway in the effects of BDNF (Matsuzaki et al., 2004). Glial cell line-derived neurotrophic factor (GDNF) administration can also prevent METH-mediated reductions in DA levels in the rat striatum (Cass, 1996; Cass et al., 2006) and hasten the recovery of striatal DA functions in METH-treated rats (Cass et al., 2000). GDNF pretreatment could also partially prevent the loss of DAT binding caused by METH administration in the striatum of vervet monkeys (Melega et al., 2000). Bone morphogenetivc protein 7 (BMP7) can also attenuate METH-induced decrease in TH immunoreactivity and cell death in primary DA neurons and in the mouse striatum (Chou et al., 2008) while nerve growth factor was reported to protect R2 cells against drug-related DNA fragmentation and apoptosis (Zhou et al., 2004).

Several studies have demonstrated that METH neurotoxicity can be mediated, in part, via activation of neuroinflammatory responses (Kuhn et al., 2006; Ladenheim et al., 2000; Thomas et al., 2004b). In line with these findings, some cytokines have been shown to induce protection against METH toxicity. For example, METH-induced damage to DA axons was prevented by pretreatment with interferon γ, possibly, by suppression of its neuroinflammatory effects (Hozumi et al., 2008). In addition, interferon γ might cause neuroprotective effects by increasing GDNF production in astrocytes (Appel et al., 1997). Another cytokine, TNF-α, which can enhance autoimmunity and inflammation (Sriram and O’Callaghan, 2007), is protective against METH-induced toxicity (Nakajima et al., 2004). The neuroprotective effects of TNF-α may be mediated by inhibition of METH-related increases in extracellular DA levels in the striatum and by potentiating DA uptake into synaptosomes and synaptic vesicles (Nakajima et al., 2004). The neuroprotection against METH toxicity caused by TNF-α may also be mediated via its effects on temperature regulation, because TNF-α pretreatment prior to drug administration also induced hypothermia, whereas METH-induced hyperthermia was exacerbated in TNF-α knockout mice (Nakajima et al., 2004). In addition, TNF-α has been shown to cause upregulation of MnSOD expression through activation of NFκB in hippocampal neurons (Mattson et al., 1997) because MnSOD transgenic mice are protected against METH toxicity (Maragos et al., 2000).

Estrogen can also protect against METH-induced damage to nigrostriatal DA system (Dluzen and McDermott, 2006). Estrogen treatment caused neuroprotective effects against METH neurotoxicity in female, but not in male mice (D’Astous et al., 2004, 2005; Dluzen et al., 2002; Liu and Dluzen, 2006). Pretreatment with tamoxifen, an estrogen receptor ligand, attenuates METH-induced striatal DA depletion and the decrease in DAT binding via mechanisms that were independent of thermoregulation (Bourque et al., 2007; Dluzen et al., 2001). In addition to its effects on estrogen receptors, tamoxifen can also cause anti-inflammatory responses in microglial cells (Suuronen et al., 2005) and protect glial cells against glutamate toxicity (Shy et al., 2000). Because glutamate release and microgliosis contribute to METH toxicity (see above), the protective effects of tamoxifen probably occur via suppression of both glutamate- and microglia-dependent toxic mechanisms.