Altered cardiac mitochondrial dynamics and biogenesis in rat after short-term cocaine administration

Abuse of the potent psychostimulant cocaine is widely established to have cardiovascular consequences. The cardiotoxicity of cocaine is mainly associated with oxidative stress and mitochondrial dysfunction. Mitochondrial dynamics and biogenesis, as well as the mitochondrial unfolded protein response (UPRmt), guarantee cardiac mitochondrial homeostasis. Collectively, these mechanisms act to protect against stress, injury, and the detrimental effects of chemicals on mitochondria. In this study, we examined the effects of cocaine on cardiac mitochondrial dynamics, biogenesis, and UPRmt in vivo. Rats administered cocaine via the tail vein at a dose of 20 mg/kg/day for 7 days showed no structural changes in the myocardium, but electron microscopy revealed a significant increase in the number of cardiac mitochondria. Correspondingly, the expressions of the mitochondrial fission gene and mitochondrial biogenesis were increased after cocaine administration. Significant increase in the expression and nuclear translocation of activating transcription factor 5, the major active regulator of UPRmt, were observed after cocaine administration. Accordingly, our findings show that before any structural changes are observable in the myocardium, cocaine alters mitochondrial dynamics, elevates mitochondrial biogenesis, and induces the activation of UPRmt. These alterations might reflect cardiac mitochondrial compensation to protect against the cardiotoxicity of cocaine.

Intravenous haloperidol and cocaine alter the distribution of T CD4+ and B lymphocytes and NKT cells in rats

Modulation of dopamine transmission evokes strong behavioral effects that can be achieved by psychoactive drugs such as haloperidol or cocaine. Cocaine non-specifically increases dopamine transmission by blocking dopamine active transporter (DAT) and evokes behavioral arousal, while haloperidol is a non-specific dopamine D2 receptor antagonist with sedative effects. Interestingly, dopamine has been found to affect immune cells in addition to its action in the central nervous system. Here we address the possible interactions between haloperidol and cocaine and their effects on both immune cells and behavior in freely moving rats. We use an intravenous model of haloperidol and binge cocaine administration to evaluate the drugs' impact on the distribution of lymphocyte subsets in both the peripheral blood and the spleen. We assess the drugs' behavioral effects by measuring locomotor activity. Cocaine evoked a pronounced locomotor response and stereotypic behaviors, both of which were completely blocked after pretreatment with haloperidol. The results suggest that blood lymphopenia which was induced by haloperidol and cocaine (except for NKT cells), is independent of dopaminergic activity and most likely results from the massive secretion of corticosterone. Haloperidol pretreatment prevented the cocaine-induced decrease in NKT cell numbers. On the other hand, the increased systemic dopaminergic activity after cocaine administration is a significant factor in retaining T CD4+ and B lymphocytes in the spleen.

An extended amygdala-midbrain circuit controlling cocaine withdrawal-induced anxiety and reinstatement

While midbrain dopamine (DA) neuronal circuits are central to motivated behaviors, much remains unknown about our knowledge of how these circuits are modified over time by experience to facilitate selective aspects of experience-dependent plasticity. Most studies of the DA system in drug addiction focus on the role of the mesolimbic DA pathway from the ventral tegmental area (VTA) to the nucleus accumbens (NAc) in facilitating drug-associated reward. In contrast, less is known about how midbrain DA cells and associated circuits contribute to negative affective states including anxiety that emerge during protracted withdrawal from drug administration. Here, we demonstrate the selective role of a midbrain DA projection to the amygdala (VTADA-Amygdala) for anxiety that develops during protracted withdrawal from cocaine administration but does not participate in cocaine reward or sensitization. Our rabies virus-mediated circuit mapping approach revealed a persistent elevation in spontaneous and task-related activity of GABAergic cells from the bed nucleus of the stria terminals (BNST) and downstream VTADA-Amygdala cells that could be detected even after a single cocaine exposure. Activity in BNSTGABA cells was related to cocaine-induced anxiety but not reward or sensitization, and silencing the projection from these cells to the midbrain was sufficient to prevent the development of anxiety during protracted withdrawal following cocaine administration. We observed that VTADA-Amygdala cells, but not other midbrain DA cells, were strongly activated after a challenge exposure to cocaine, and found that activity in these cells was necessary for the expression of reinstatement of cocaine place preference. Lastly, the importance of activity in VTADA-Amygdala cells extends beyond cocaine, as these cells mediate the development of anxiety states triggered by morphine and a predator odor. Our results provide an exemplar for how to identify key circuit substrates that contribute to behavioral adaptations and reveal a critical role for BNSTGABA-VTADA-Amygdala pathway in anxiety states induced by drugs of abuse or natural experiences as well as cocaine-primed reinstatement of conditioned place preference.

Role of hippocampal NF-κB and GluN2B in the memory acquisition impairment of experiences gathered prior to cocaine administration in rats

Cocaine can induce severe neurobehavioral changes, among others, the ones involved in learning and memory processes. It is known that during drug consumption, cocaine-associated memory and learning processes take place. However, much less is known about the effects of this drug upon the mechanisms involved in forgetting.The present report focuses on the mechanisms by which cocaine affects memory consolidation of experiences acquired prior to drug administration. We also study the involvement of hippocampus in these processes, with special interest on the role of Nuclear factor kappa B (NF-κB), N-methyl-D-aspartate glutamate receptor 2B (GluN2B), and their relationship with other proteins, such as cyclic AMP response element binding protein (CREB). For this purpose, we developed a rat experimental model of chronic cocaine administration in which spatial memory and the expression or activity of several proteins in the hippocampus were assessed after 36 days of drug administration. We report an impairment in memory acquisition of experiences gathered prior to cocaine administration, associated to an increase in GluN2B expression in the hippocampus. We also demonstrate a decrease in NF-κB activity, as well as in the expression of the active form of CREB, confirming the role of these transcription factors in the cocaine-induced memory impairment.

Evaluation of Cocaine Effect on Endogenous Metabolites of HepG2 Cells Using Targeted Metabolomics

Cocaine toxicity has been a subject of study because cocaine is one of the most common and potent drugs of abuse. In the current study the effect of cocaine on human liver cancer cell line (HepG2) was assessed. Cocaine toxicity (IC50) on HepG2 cells was experimentally calculated using an XTT assay at 2.428 mM. The metabolic profile of HepG2 cells was further evaluated to investigate the cytotoxic activity of cocaine at 2 mM at three different time points. Cell medium and intracellular material samples were analyzed with a validated HILIC-MS/MS method for targeted metabolomics on an ACQUITY Amide column in gradient mode with detection on a triple quadrupole mass spectrometer in multiple reaction monitoring. About 106 hydrophilic metabolites from different metabolic pathways were monitored. Multivariate analysis clearly separated the studied groups (cocaine-treated and control samples) and revealed potential biomarkers in the extracellular and intracellular samples. A predominant effect of cocaine administration on alanine, aspartate, and glutamate metabolic pathway was observed. Moreover, taurine and hypotaurine metabolism were found to be affected in cocaine-treated cells. Targeted metabolomics managed to reveal metabolic changes upon cocaine administration, however deciphering the exact cocaine cytotoxic mechanism is still challenging.

Cocaine-Induced Changes in Tonic Dopamine Concentrations Measured Using Multiple-Cyclic Square Wave Voltammetry in vivo

For over 40 years, in vivo microdialysis techniques have been at the forefront in measuring the effects of illicit substances on brain tonic extracellular levels of dopamine that underlie many aspects of drug addiction. However, the size of microdialysis probes and sampling rate may limit this technique’s ability to provide an accurate assessment of drug effects in microneural environments. A novel electrochemical method known as multiple-cyclic square wave voltammetry (M-CSWV), was recently developed to measure second-to-second changes in tonic dopamine levels at microelectrodes, providing spatiotemporal resolution superior to microdialysis. Here, we utilized M-CSWV and fast-scan cyclic voltammetry (FSCV) to measure changes in tonic or phasic dopamine release in the nucleus accumbens core (NAcc) after acute cocaine administration. Carbon-fiber microelectrodes (CFM) and stimulating electrodes were implanted into the NAcc and medial forebrain bundle (MFB) of urethane anesthetized (1.5 g/kg i.p.) Sprague-Dawley rats, respectively. Using FSCV, depths of each electrode were optimized by determining maximal MFB electrical stimulation-evoked phasic dopamine release. Changes in phasic responses were measured after a single dose of intravenous saline or cocaine hydrochloride (3 mg/kg; n = 4). In a separate group, changes in tonic dopamine levels were measured using M-CSWV after intravenous saline and after cocaine hydrochloride (3 mg/kg; n = 5). Both the phasic and tonic dopamine responses in the NAcc were augmented by the injection of cocaine compared to saline control. The phasic and tonic levels changed by approximately x2.4 and x1.9, respectively. These increases were largely consistent with previous studies using FSCV and microdialysis. However, the minimal disruption/disturbance of neuronal tissue by the CFM may explain why the baseline tonic dopamine values (134 ± 32 nM) measured by M-CSWV were found to be 10-fold higher when compared to conventional microdialysis. In this study, we demonstrated phasic dopamine dynamics in the NAcc with acute cocaine administration. M-CSWV was able to record rapid changes in tonic levels of dopamine, which cannot be achieved with other current voltammetric techniques. Taken together, M-CSWV has the potential to provide an unprecedented level of physiologic insight into dopamine signaling, both in vitro and in vivo, which will significantly enhance our understanding of neurochemical mechanisms underlying psychiatric conditions.

Sustained splenic contraction after daily cocaine administration in rats

The purpose of this study is to examine the effect of repeated cocaine administration on the whole body of rats. Rats (male, 6 weeks old, Sprague Dawley) were injected intraperitoneally with cocaine (50 mg/kg) once a day for 1, 3 or 7 days, and major organs (heart, liver, lung, brain, kidney, spleen) were excised from the sacrificed animals. During autopsy, we found a reduction in spleen size, but not other organs, in cocaine-administered rats as compared to control rats. This reduction became to be noticed at 3 day and easily perceived at 7 day. No marked changes were observed in other organs examined. H&E and EMG staining showed a tendency for a decrease in the number of red blood cells (RBCs) as well as an increase in collagen fibers in the spleens of rats treated repeatedly with cocaine. Transcriptome analysis indicated that repeated cocaine administration depletes RBCs from the spleen. Immunoblot analysis showed that cocaine increases the phosphorylation of myosin light chain (MYL) as well as the levels of transgelin, both of which are involved in the contraction of myofibrils. Collectively, these results show that repeated cocaine administration results in sustained contraction of the spleen, which leads to the release of RBCs from the spleen into circulation.