Tehila Beiser, Rami Yaka*
Institute for Drug Research (IDR), School of Pharmacy, Faculty of Medicine, The Hebrew University of Jerusalem, Jerusalem, 91120, Israel
Cocaine is a powerfully addictive psychostimulant that elevates dopamine (DA) levels in the mesolimbic system and causes a feeling of wellbeing. At the same time, cocaine leads to toxic effects in many essential organs, including the brain. The harmful effects of cocaine on the brain are the basis for the development of compulsive and irrational behaviors, an integral part of cocaine addiction. Over the last two decades, it has been suggested that the damage and reinforcing properties of cocaine are associated with increased reactive oxygen species (ROS) production. This increase impairs the endogenous defense antioxidant system, either directly by cocaine metabolites, or indirectly via increased DA metabolites, resulting in oxidative stress (OS). It was thus plausible to seek an exogenous, stable and non-toxic antioxidant, which can penetrate the blood brain barrier and counteract the oxidative damage in the brain caused by drugs of abuse such as cocaine. In this mini-review we describe studies that explore the role of antioxidants in reducing the OS state in the brain reward system and consequently reversing negative behavioral outcomes induced by cocaine.
Cocaine addiction has taken a severe toll not only on drug users themselves but on society as well. The strong craving and impaired judgment associated with cocaine addiction leads to increased and regular drug use despite its harmful consequences. According to the 2018 world drug report, in 2016 there were over 18 million cocaine users worldwide aged 15-641, and the use and demand for cocaine is still growing. Cocaine dependence and craving, representing physical and neuronal damage in the brain, might last years after withdrawal, creating a lifetime risk for relapse2. Thus, researchers strive to understand the mechanism of cocaine neuronal damage in the hope of finding a suitable treatment.
Cocaine has been known to be very toxic to many body organs including the liver, the cardiovascular system, and the central nervous system3–5. However, the mechanisms for cocaine`s toxicity is still poorly understood. Numerous studies suggested that cocaine`s toxicity results in part from oxidative stress (OS)6. First, cocaine metabolites, such as norcocaine and its derivatives nitroxide and nitrosonium were found to be even more toxic than cocaine itself in increased reactive oxygen species (ROS)7 and in reducing activity of the endogenous protecting antioxidant system, including its main components superoxide dismutase (SOD), glutathione (GSH)8 and catalase (CAT)9. Second, cocaine blocks re-uptake of dopamine, and as a result elevates dopamine levels. These findings suggest that dopamine and its oxidative metabolites, mainly dopamine quinone, accumulate in the brain and increase ROS production10.
Evidence relating cocaine administration to OS is increasing over the years. One of the first indications for the damage of cocaine to the endogenous antioxidative system was found by Lipton et al8 who reported that even a single prenatal cocaine intake reduces levels of GSH. Another indication was found by Baker et al11 who showed that down-regulation of cystine-glutamate exchanger is involved in cocaine relapse during withdrawal from cocaine self-administration (Box 1). A later study reported that while in the hippocampus of rats the activity of neuronal nitric oxide synthase (nNOS) was increased in response to cocaine, GSH concentration and glutathione peroxidase activity was decreased12. Uys et al.13 found redox homeostasis changes in glutathione that were associated with OS and plasticity induced behavioral changes in psychomotor sensitization (Box 1) and conditioned reward (Box 1). Taken together, these studies suggest that while ROS are elevated, the glutathione system, as part of the antioxidant defense system, is critically interrupted by acute and chronic exposure to cocaine and may be related to cocaine induced behaviors.
Another aspect of the endogenous antioxidant system is CAT that converts hydrogen peroxide to water. It was reported that the activity of catalase is weakened in response to cocaine in the striatum and pre frontal cortex (PFC) of mice9. Since hydrogen peroxide is reported to inhibit dopamine transporters, catalase inactivation results in accumulated toxic effects. On the same line, it was found that the levels of hydrogen peroxide increased in response to a single or repeated cocaine injections, emphasizing the involvement of cocaine in reducing catalase activity14. Walker et al.15 were the first to evaluate the deficiency of the endogenous antioxidant defense system including CAT, SOD, and GSH in the blood of methamphetamine and cocaine dependent humans. More reports found that exposure to cocaine in utero led to an increase in NO and lipid peroxidation in the hippocampus and cortex16 and that cocaine-induced OS provoked cell apoptosis in human neuronal networks17. In our previous studies, we found that cocaine induced OS in the nucleus accumbens (NAc) and PFC, two key structures of the reward system, both in slices and following administration of a single or repeated cocaine injections18. Similarly, we showed that cocaine induced cell death in PC-12 cell-line through OS19. Together, these studies suggest that antioxidants can be suitable candidates to reduce cocaine-induced OS and cocaine-induced behaviors.
Tempol (4-hydroxy-2,2,6,6 tetramethylpiperidine-N-oxyl), is a stable nitroxide radical20,21 that mimics SOD in a catalytic mode and interacts with radicals to prevent radical chain reactions21. Tempol has more advantages that make it a good candidate to protect against cocaine-induced OS and subsequent neuronal damage; it easily permeates cell membranes22,23, it penetrates and protects the blood brain barrier (BBB)24,25 and more importantly, it is a stable radical20,21. The biological activity of Tempol in treating OS damage has been studied in diverse areas: it was found to be effective in protecting from radiation26 and traumatic head injury27; it was reported to prevent taxol-induced neuronal death28 and cisplatin-induced ototoxicity in vitro29; it prevented cognition impairments in a simulation to Alzheimer’s disease30, inflammatory pain31, short and long-term PTSD-induced memory impairments32 and β-cell death33 just to name a few; in addition to other reports on medical applications of nitroxide radicals34,35. Here we review only studies presenting the antioxidant effects of Tempol against cocaine-induced OS.
The first evidence for the neuroprotective activity of Tempol against cocaine-induced oxidative stress and the attenuation of cocaine-induced behaviors were obtained in our lab. We found that in sympathetic-like PC12 cells, Tempol significantly reduced cocaine-induced cell death19. In addition, we found that two other members of the nitroxide family, Tempo and Tempamine, exhibited a similar effect. Moreover, we confirmed that cocaine induced cell death through the production of ROS by measuring mitochondrial superoxide radical formation and peroxide levels. Indeed, both ROS markers were elevated following cocaine exposure, and decreased when treated with Tempol, indicating that Tempol is able to reverse cocaine-induced cell death. Furthermore, we examined the effect of Tempol in attenuating cocaine-induced OS in animals. We found that Tempol decreased lipid peroxidation and the levels of NO radicals in both the NAc and PFC of rats following a single cocaine injection18. Pretreatment with Tempol in PFC and NAc containing brain slices decreased malondialdehyde (MDA); a lipid peroxidation marker; and nitrites levels and increased total antioxidant capacity (TAC), indicating that Tempol may contribute to the endogenous antioxidant defense system. We next evaluated the effect of Tempol on cocaine-induced behaviors. Characteristically, locomotor activity increased in response to repeated injections of the same dose of cocaine and other drugs of abuse. Psychomotor sensitization (PS) reflects the enduring rise in neuronal processes that lie behind addictive behaviors and is also known to be correlated indirectly with drug seeking (see Box 1). We found that in animals treated with Tempol prior to cocaine within the PS paradigm, acute locomotor activity was evident but the development and expression (locomotion activity measured following challenge cocaine injection) of PS was completely abolished. Moreover, we found that in Tempol treated animals, OS was returned to base line levels following the behavioral tests suggesting that Tempol attenuated cocaine PS via the reduction in oxidative stress18. Recently, we studied the effect of Tempol on cocaine conditioned reward using the conditioned place preference (CPP) paradigm (Box 1). We demonstrated that injection of Tempol together with cocaine during acquisition of CPP attenuated the expression of CPP measured one day following conditioning36. We found that OS was significantly elevated following the establishment of CPP, and that cocaine-induced OS was significantly diminished by pretreatment with Tempol during conditioning. Furthermore, we found that repeated, but not a single, administration of Tempol for seven days during withdrawal from CPP resulted in a significant attenuation in the expression of CPP, without affecting the expression of food reward36. Together these results suggest that antioxidants such as Tempol, when administered before or during withdrawal, can interfere with reward processing of drugs of abuse without affecting the response to natural rewards.
Jang et al37 studied the effects of Tempol on cocaine self-administration (SA), a behavioral paradigm that model drug seeking and craving (Box 1). Injecting Tempol before or after the initiation of SA sessions resulted in a significantly reduced number of the active lever presses. These results support the hypothesis that ROS has a critical role in cocaine craving. They further found that OS occurs mainly in neurons and not in astrocytes, microglia or oligodendrocytes in the NAc of the cocaine SA animals. Moreover, in order to study the involvement of ROS on cocaine reinforcement specifically in the NAc, they microinjected Tempol bilaterally into the NAc prior to cocaine SA and found that the active lever presses were significantly reduced in the Tempol injected group, compared to the saline-injected control group. Systemic injections of Tempol before food SA demonstrated that Tempol did not have any effect on food reinforcement, suggesting that Tempol suppresses cocaine SA without affecting food SA or inactive lever presses, similar to our finding using CPP paradigm36.
Table 1. Evidence for cocaine-induced oxidative stress.
|Brain area||Cocaine effect on oxidative stress||Paradigm||Reference|
|Fetus brain||Single cocaine injection decreased endogenous antioxidant glutathione and exogenous antioxidant alpha Tocopherol but increased their oxidized form||Prenatal effect of single cocaine injection||8|
|NAc, PFC and striatum||Down-regulation of cystine-glutamate exchanger in cocaine relapse||Withdrawal from self- administration in rats||11|
|Hippocampus and frontal cortex||Increased nNOS, GSH concentration and glutathione peroxidase activity is decreased. Memory retrieval is impaired||Morris water maze in rats||12|
|NAc||OS and plasticity is affected by redox homeostasis changes in glutathione following acute cocaine||Psychomotor sensitization, conditioned place preference in rats||13|
|Striatum and PFC||Decreased catalase activity||Single injection of cocaine in mice||9|
|Striatum and frontal cortex||Cocaine increased hydrogen peroxide and induced glutathione peroxidase and SOD activity||Single and repeated injections of cocaine in rats||14|
|Blood||CAT, SOD, and GSH deficiency and reduction of total antioxidant capacity||Human blood analysis in cocaine and methamphetamine dependent users||15|
|Hippocampus and cortex||increase in NO and lipid peroxidation||In utero in mice||16|
|NAc and PFC||Increased lipid peroxidation and nitrites||Acute and repeated cocaine injections in rat||18|
|Human neuronal progenitor cells||OS leads to cell apoptosis||Acute cocaine treatment||17|
|Neuronal-like PC12 cells||Increased OS and cell death||Acute cocaine treatment||19|
The role of Tempol in attenuating drug reinforcement was studied using other drugs of abuse. Qi et al38 explored the involvement of mGluR5 in morphine CPP and found that microinfusion of MTEP (mGluR5 antagonist) into the NAc shell (the outer substructure of the NAc), attenuated morphine CPP, and blocked the increase of membrane mGluR5 that was observed following morphine CPP in rats. They found that microinjection of Tempol into the NAc shell prevented morphine CPP expression in a similar way to MTEP, but the evidence that mGluR5 and ROS signaling is involved in retrieval of morphine contextual memory was found when microinjection of t-BOOH, a ROS donor, reversed the effect of mGluR5 antagonist MTEP in attenuating morphine CPP. Jang et al39 found that methamphetamine (METH) induced dopamine release and consequently ROS formation in the NAc. METH-induced ROS were found only in neurons but not in glia, astrocytes or oligodendrocytes. They further showed that systemic administration of Tempol decreased not only dopamine release in the NAc, but it also attenuated METH-induced locomotor activity and the reinforcing effects of METH self-administration in rats, without affecting food intake, similar to what was found with cocaine36,37.
In this article, we reviewed the studies that demonstrated the harmful properties of cocaine at the molecular, cellular and behavioral levels which is evidently promoted by ROS formation and become persistent by ROS-induced signaling cascades. In the reviewed studies it is suggested that the harmful consequences of cocaine use could be reversed by employing the antioxidant Tempol as pretreatment and, in some cases, as a treatment following exposure to cocaine. These studies demonstrate a variety of behavioral models: psychomotor sensitization, CPP and self-administration, which together represent a different aspect of addictive behaviors. Thus, these studies suggest that Tempol could be a potential pharmacological treatment not only for cocaine addiction but also to other drugs of abuse.
Supported by the Israel Anti-Drug Authority (IADA) and by the Israel Science Foundation (ISF). R. Yaka is affiliated with the David R. Bloom Center for Pharmacy and the Brettler Center for Research in Molecular Pharmacology and Therapeutics, School of Pharmacy, The Hebrew University of Jerusalem.