Tong H. Lee

Methamphetamine neurotoxicity: necrotic and apoptotic mechanisms and relevance to human abuse and treatment

Research into methamphetamine-induced neurotoxicity has experienced a resurgence in recent years. This is due to (1) greater understanding of the mechanisms underlying methamphetamine neurotoxicity, (2) its usefulness as a model for Parkinsons disease and (3) an increased abuse of the substance, especially in the American Mid-West and Japan. It is suggested that the commonly used experimental one-day methamphetamine dosing regimen better models the acute overdose pathologies seen in humans, whereas chronic models are needed to accurately model human long-term abuse. Further, we suggest that these two dosing regimens will result in quite different neurochemical, neuropathological and behavioral outcomes. The relative importance of the dopamine transporter and vesicular monoamine transporter knockout is discussed and insights into oxidative mechanisms are described from observations of nNOS knockout and SOD overexpression. This review not only describes the neuropathologies associated with methamphetamine in rodents, non-human primates and human abusers, but also focuses on the more recent literature associated with reactive oxygen and nitrogen species and their contribution to neuronal death via necrosis and/or apoptosis. The effect of methamphetamine on the mitochondrial membrane potential and electron transport chain and subsequent apoptotic cascades are also emphasized. Finally, we describe potential treatments for methamphetamine abusers with reference to the time after withdrawal. We suggest that potential treatments can be divided into three categories; (1) the prevention of neurotoxicity if recidivism occurs, (2) amelioration of apoptotic cascades that may occur even in the withdrawal period and (3) treatment of the atypical depression associated with withdrawal.

Reversal of cocaine-induced behavioral sensitization and associated phosphorylation of the NR2B and GluR1 subunits of the NMDA and AMPA receptors.

Cocaine abusers remain vulnerable to drug craving and relapse for many years after abstinence is achieved. We have recently shown that ondansetron (a 5-HT3 receptor antagonist) given 3.5 h after each daily cocaine injection reverses previously established behavioral sensitization. The purpose of the present investigation was two-fold. First, as cocaine cannot be used as therapy, we examined whether pergolide (a D1/D2 receptor agonist with reduced abuse potential) and ondansetron could reverse behavioral sensitization. Second, we investigated whether these behavioral changes were associated with parallel alterations in expression levels and/or phosphorylation changes in the NR2B and GluR1 subunits of the respective NMDA and AMPA receptors. Rats were injected for 5 consecutive days with cocaine or saline followed by 9 days of withdrawal. Starting on withdrawal day 10, animals were given vehicle, pergolide/saline, or pergolide/ondansetron for 5 consecutive days. Following a second 9-day period of withdrawal, all animals were challenged with cocaine for assessment of behavioral sensitization and tissues were collected on the following day for Western blot. Sensitization was associated with increased NR2B expression in the accumbens (NAc) shell and decreased Tyr1472 phosphorylation in the NAc core, as well as increased Ser845 phosphorylation of the GluR1 subunit in prefrontal cortex, NAc core, and shell. Pergolide/ondansetron treatment, but not pergolide alone, consistently reversed both the behavioral sensitization and the associated changes in the NMDA and AMPA receptor subunits. To the extent that sensitization plays a role in chronic cocaine abuse, a combination of these clinically available drugs may be useful in treatment of the disorder.

Ondansetron Given in the Acute Withdrawal from a Repeated Cocaine Sensitization Dosing Regimen Reverses the Expression of Sensitization and Inhibits Self-administration

Male Sprague-Dawley rats were given two separate sensitizing regimens of cocaine (7 days on, 7 days off, 7 days on; 40 mg/kg/day s.c.) along with saline controls. Furthermore, animals also received the 5-HT3 antagonist ondansetron (0.2 mg/kg s.c.) either during the second dosing regimen (3.5 h after each cocaine/saline injection) or during the first five days of the second withdrawal period. Animals were then challenged, on day 10 of withdrawal, with cocaine (7.5 mg/kg i.p.) and assessed by a behavioral rating scale and locomotor activity monitoring. The cocaine regimen induced behavioral and locomotor sensitization on day 10 of withdrawal, further, ondansetron inhibited sensitization regardless of whether given after each second cocaine regimen dose or during the second withdrawal period, although treatment 3.5 h after each cocaine injection appeared more effective. Ondansetron did not inhibit behavior in control animals. In a second experiment animals were trained to self-administer cocaine via an indwelling jugular catheter. After stable fixed-ratio responding (FR1 then FR2) they were given a progressive ratio (PR) schedule until PR each day was stable. During the first five days of withdrawal they were given either ondansetron (0.2 mg/kg s.c.) or saline injections. On day 10 of withdrawal the cocaine PR schedule was reinstated. The ondansetron treated rats showed only a non-significant decrease in break point. After day 2 of the PR session rats were again injected with either ondansetron (0.2 mg/kg s.c.) or saline, 3.5 h after each PR session for five days. Ondansetron inhibited cocaine self-administration on each of the following days. Ondansetron may be a useful treatment for cocaine addicts who have undergone previous sensitization periods.

5-HT2 receptor antagonists given in the acute withdrawal from daily cocaine injections can reverse established sensitization.

Male Sprague-Dawley rats were given two separate sensitizing regimens of cocaine (7 days on, 7 days off, 7 days on at 40 mg/kg/day, s.c.) or saline injections. Half of the animals also received a drug with 5-hydroxytryptamine-2 (5-HT2) receptor antagonist properties (clozapine, 3 mg/kg; mianserin 6 mg/kg; ketanserin 1 mg/kg, all s.c.) or saline during the second cocaine dosing regimen in the acute withdrawal period, 3.5 h after each cocaine injection. On day 10 of withdrawal animals were challenged with cocaine (7.5 mg/kg, i.p.) and assessed by a behavioral rating scale and locomotor activity monitoring. The 5-HT2 receptor antagonists, but not saline, reversed behavioral sensitization and had little effect on behavior in the control animals. 5-HT2 receptor antagonists, therefore, may be a useful treatment for cocaine addicts that have undergone previous sensitization periods. The pharmacological profile of these antagonists suggests that the 5-HT2A receptor subtype may mediate this effect.

Acute and chronic continuous methamphetamine have different long-term behavioral and neurochemical consequences

We compared two different methamphetamine dosing regimens and found distinct long-term behavioral and neurochemical changes. Adult rats were treated with 1-day methamphetamine injection (3x5 mg/kg s.c., 3 h apart) or 7-day methamphetamine minipump (20 mg/kg/day s.c.). The minipump regimen models the sustained methamphetamine plasma levels in some human bingers whereas the 1-day regimen models a naive user overdose. On withdrawal days 7 and 28, rats were acutely challenged with cocaine to test for behavioral sensitization and subsequently sacrificed for caudate and accumbens dopamine tissue content. Other rats were analyzed on withdrawal days 3, 7 or 28 using voltammetry in caudate slices. On withdrawal days 7 and 28, the methamphetamine injection but not the minipump rats showed behavioral cross-sensitization to cocaine. There was no change in baseline dopamine release, reuptake or sensitivity to quinpirole in any treatment group on either withdrawal day. However, consistent with the behavioral sensitization, cocaine had a greater effect in potentiating dopamine release and in blocking dopamine reuptake in methamphetamine injection versus saline irrespective of withdrawal day. The minipump group showed tolerance to the dopamine releasing effect of cocaine on withdrawal day 28 and had lower dopamine tissue content in the caudate versus the methamphetamine injection group. Dopamine turnover as measured by the DOPAC/dopamine ratio tended to be higher in the minipump-treated rats. These data suggest that the behavioral cross-sensitization seen in the methamphetamine injection rats could be in part due to the increased potency of cocaine in blocking dopamine reuptake and in increasing dopamine release. The decreased potency of cocaine in the caudate slices from the minipump-treated group may be related to decreased dopamine tissue content.

Alterations in Baseline Activity and Quinpirole Sensitivity in Putative Dopamine Neurons in the Substantia Nigra and Ventral Tegmental Area after Withdrawal from Cocaine Pretreatment

Using in vivo single-unit recording, we compared in rats the effects of continuous infusion and once-a-day injections of cocaine on the activity of single putative dopamine neurons in the substantia nigra and ventral tegmental area. After a 7-day withdrawal, we determined: (1) the number of spontaneously active neurons and their bursting patterns and (2) sensitivity of these neurons to intravenous quinpirole. In the substantia nigra, continuous cocaine infusion reduced the number of neurons without affecting the bursting patterns; daily injections were without effects. In the ventral tegmental area, continuous infusion reduced the bursting activity without affecting the number of neurons, whereas infections increased number of neurons without changes in the bursting pattern. Acute sulpiride normalized all the changes in both cell body areas. The quinpirole sensitivity was selectively increased in the nigral neurons following withdrawal from continuous infusion. Possible role of D2/autoreceptor mechanisms in these changes is discussed.

Neural plasticity and addiction: integrin-linked kinase and cocaine behavioral sensitization.

Behavioral sensitization of psychostimulants was accompanied by alterations in a variety of biochemical molecules in different brain regions. However, which change is actually related to drug‐produced sensitization lacks of accurate clarification. In this study, we investigated the role of integrin‐linked kinase (ILK) in both the induction and expression of cocaine sensitization. Conditional inhibition of ILK expression was established in the nucleus accumbens (NAc) core by microinjecting recombinant adeno‐associated virus‐carrying, tetracycline‐on‐regulated small interfering RNA which reversed the chronic cocaine‐induced psychomotor sensitization, as well as the changes in protein kinase B Ser473 phosphorylation, dendritic density, and dendritic spine numbers locally. Importantly, the reversed psychomotor sensitization did not recover after cessation of the silencing for 8 days. We also demonstrated that inhibition of ILK expression pre‐ and during‐chronic cocaine treatments blocked the induction of cocaine psychomotor sensitization and abolished the stimulant effect of cocaine on ILK expression. In contrast, inhibition of ILK expression in the NAc core has no significant effect on cocaine‐induced stereotypical behaviors. This concludes that ILK is involved in cocaine sensitization with the earlier induction and later expression functioning as a kinase to regulate protein kinase B Ser473 phosphorylation and a scaffolding protein to regulate the reorganization of the NAc spine morphology.

Altered sensitivity of dopamine autoreceptors in rat accumbens 1 and 7 days after intermittent or continuous cocaine withdrawal.

Using slice preparations, we investigated the effects of chronic cocaine treatment on dopamine autoreceptor sensitivity in the nucleus accumbens core. Cocaine (40 mg/kg/day) was given for 14 days, either by continuous subcutaneous infusion (osmotic minipumps) or single daily injections. One or 7 days after cocaine withdrawal, we used fast scan cyclic voltammetry (10 Hz sampling rate) to measure inhibition of electrically evoked dopamine release by quinpirole (3-300 nM). Continuous cocaine infusion increased quinpirole sensitivity on day 1 of withdrawal, particularly at low concentrations of quinpirole, but this effect was no longer evident by day 7. Intermittent cocaine injections had no effect on day 1 of withdrawal but by day 7 there was a quinpirole subsensitivity. On either withdrawal day, the baseline peak dopamine release or uptake half-life exhibited no treatment group differences. It is suggested that these cocaine dosing regimes cause differential and dynamic changes in dopamine autoreceptor sensitivity during the early withdrawal phase.

Integrin-linked kinase is involved in cocaine sensitization by regulating PSD-95 and synapsin I expression and GluR1 Ser845 phosphorylation.

Our recent studies have demonstrated that integrin-linked kinase (ILK) is involved in the induction and maintenance of cocaine behavioral sensitization and chronic cocaine-induced neural plasticity in the nucleus accumbens (NAc) core. In the present study, we used ILK silencing to investigate how ILK may influence cocaine-induced neural plasticity. Adeno-associated virus carrying a small interfering RNA-ILK cassette under the control of an inducible Tet-On system was injected into the NAc core of Sprague–Dawley rats. Induced silencing was established during repeated cocaine injections (sensitization induction period) or between withdrawal days 9 and 22 (sensitization maintenance period). Under both paradigms, established cocaine sensitization under non-silenced conditions was associated with enhanced PSD-95 and synapsin I protein expression as well as enhanced Ser845 phosphorylation of the GluR1 subunit on withdrawal day. Silencing ILK expression under both paradigms prevented or reversed these changes. Importantly, ILK appears to form a complex with PSD-95 and synapsin I because it co-immunoprecipitated with each of these proteins. Together, these data suggest that ILK exerts pleiotropic actions by regulating pre- and postsynaptic neural plasticities within the NAc core in response to repeated cocaine exposure.

Behavioral sensitization is greater after repeated versus single chronic cocaine dosing regimens

Cocaine dosing regimens in animals are used to model behavioral and neurochemical changes in human cocaine abusers. Typically, rats are dosed for 5-14 days and assessed at some point during withdrawal. However, human cocaine bingers undergo multiple periods of several days of abuse. Here, we model the human binge pattern by giving rats two separate cocaine dosing regimens which results in greater behavioral sensitization than a single cocaine dosing regimen. This model also allows for the testing of drugs in reversal of a previously established sensitization. Multiple cocaine regimens may thus provide a better model for the human condition.