Matlab 2007 Plp Crack Cocaine
Johna Delehanty
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Heroin, nicotine, cocaine, and MDMA are abused by billions of people. They are believed to target midbrain dopamine neurons and/or serotonin neurons, but their effects on the dynamic neuronal activity remain unclear in behaving states. By combining cell-type-specific fiber photometry of Ca2+ signals and intravenous drug infusion, here we show that these four drugs of abuse profoundly modulate the activity of mouse midbrain dopamine neurons and serotonin neurons with distinct potency and kinetics. Heroin strongly activates dopamine neurons, and only excites serotonin neurons at higher doses. Nicotine activates dopamine neurons in merely a few seconds, but produces minimal effects on serotonin neurons. Cocaine and MDMA cause long-lasting suppression of both dopamine neurons and serotonin neurons, although MDMA inhibits serotonin neurons more profoundly. Moreover, these inhibitory effects are mediated through the activity of dopamine and serotonin autoreceptors. These results suggest that the activity of dopamine neurons and that of serotonin neurons are more closely associated with the drug's reinforcing property and the drug's euphorigenic property, respectively. This study also shows that our methodology may facilitate further in-vivo interrogation of neural dynamics using animal models of drug addiction.
Drug abuse causes serious health and social problems. The United Nations World Drug Report estimates that a quarter of a billion world population uses at least one illegal drug in a year1. Opiates and cocaine represent two of the most addictive and devastating drugs of abuse2. MDMA is a popular recreational drug that often causes hallucination, cognitive defects, and post-drug depression3. Nicotine, though legal and widely consumed in most societies, is linked to a multitude of malignant diseases including lung cancer4.
Given the difficulty in vivo recording in a cell-type-specific manner, we still lack systematic data about how various drugs of abuse directly affect the activity of dopamine neurons and serotonin neurons in freely behaving animals. Although microdialysis studies have detected the release of dopamine and serotonin within minutes during drug administration15,16, this technique does not precisely monitor neuronal activity at sub-second temporal resolution. Fast-scan cyclic voltammetry (FSCV) has been developed to evaluate the sub-second volume transmission of dopamine17,18,19,20. However, FSCV in freely behaving animals faces forbiddingly demanding technical challenges. Similarly, recordings from slice preparations fail to reveal in vivo kinetics of neuronal activity challenged with different doses of drugs of abuse21,22. Again, electrophysiological recordings in vivo are difficult and often lack cell-type specificity and require anesthetic treatment23,24.
Here we studied how acute exposure of cocaine, MDMA, heroin, or nicotine modulates the neuronal activity of VTA dopamine neurons and DRN serotonin neurons by combining fiber photometry with intravenous drug infusion. Fiber photometry, together with the genetically-encoded Ca2+ indicator GCaMP6, has permitted the tracking of the activity dynamics of cell-type-specific neurons with sub-second temporal resolution25,26,27; these ease-of-use techniques are well adapted for freely moving transgenic mice during drug infusion. Our study revealed several interesting features of how four commonly abused drugs dynamically and differentially modulate the activity of dopamine neurons and serotonin neurons in freely behaving states.
We combined an intravenous infusion system with fiber photometry to monitor real-time changes in neuronal activity from freely behaving mice challenged with drugs of abuse (Fig. 1a). Specifically, we targeted GCaMP6m to VTA dopamine neurons and DRN serotonin neurons by stereotaxically injecting Cre-dependent AAVs into the VTA of DAT-Cre mice (DAT-VTA-GCaMP6 mice for simplicity) or into the DRN of Sert-Cre (SERT-DRN-GCaMP6) mice (Supplementary Fig. S1a). During the surgical procedure, optical fiber tips were implanted at the injection sites (Supplementary Fig. S1b). Consistent with our previous studies9,10,28, GCaMP6m was selectively expressed within VTA dopamine neurons and within DRN serotonin neurons (Supplementary Fig. S1c).
In contrast to dopamine neurons, the overall effect of nicotine on serotonin neurons were very modest at all three dose (Fig. 2b, c). Furthermore, serotonin neurons showed decreases in the amplitude and frequency of Ca2+ transients during nicotine exposure (Supplementary Fig. S6b, S6d and S6f). Our results suggest that nicotine produces a blunting inhibition of serotonin neurons in freely behaving animals. Because nicotine lacked a clear effect on the Ca2+ signal of serotonin neurons, no correlation was observed between the change of locomotor activity with Ca2+ signals of serotonin neurons (Supplementary Fig. S5c and S5d).
Heroin, nicotine, cocaine, and MDMA are commonly abused drugs that are believed to target neurons involved in reward processing. Considering that rapid change in the activity of dopamine and serotonin neurons could have profound effect on animal behaviors and the state of anesthesia fundamentally influences the activity of these neurons40,41,42, it is important to study how drugs of abuse affect the activity of these two neuron populations in freely behaving animals. Using fiber photometry of Ca2+ signals, we demonstrated that these drugs potently and differentially modulate the activity of VTA dopamine neurons and DRN serotonin neurons in freely behaving mice. Our results have several functional implications on how drugs may produce different behavioral effects by distinctly affecting the activity of dopamine neurons and serotonin neurons. Moreover, the method of combining fiber photometry and intravenous drug delivery may well complement current approaches, including microdialysis, FSCV and electrophysiology, for unraveling rapid impacts of the wide-ranging drugs.
We found that heroin strongly activates VTA dopamine neurons in behaving mice. We also provide the important information regarding the dose-kinetic relationship. Intensive research has hypothesized that opiate activates -opioid receptors on local and VTA-projecting GABA neurons and then disinhibits dopamine neurons22,24. However, the finding of sustained activation of dopamine neurons in vivo has not been observed by classic electrophysiology22,24. It is an interesting question whether this long-term activation of dopamine neurons is correlated with self-administration performance. Very few studies have studied the in vivo responses of DRN serotonin neurons to opiate exposure. We show that heroin at higher doses also significantly activates serotonin neurons. This heroin-induced activation may be achieved by disinhibiting serotonin neurons through reducing local GABAergic activity43,44. Given that some DRN serotonin neurons may corelease glutamate8,45, some heroin-associate behaviors might be mediated by serotonin and the potentially coreleased glutamate.
This study provides the first demonstration that cocaine causes strong and sustained inhibition of both dopamine neurons and serotonin neurons in behaving state. This result substantiates early observations using single-unit recording in anesthetic animals that psychostimulants decrease the firing rate of dopamine neurons35,38. We further revealed that both dopamine neurons and serotonin neurons undergo the long-lasting inhibition from MDMA exposure, with serotonin neurons exhibiting a much greater extent of inhibition. This is consistent with the concept that cocaine and MDMA enhances extracellular levels of dopamine and serotonin by binding to DAT and SERT, thus blocking their reuptake. MDMA binds more weakly to DAT than to SERT33, which explains a milder effect of MDMA on dopamine neurons. The increased levels of dopamine and serotonin then in turn acts on inhibitory autoreceptors to inhibit dopamine neurons and serotonin neurons through the opening of GIRK-type potassium channels34,55.
The strong inhibitory effects of cocaine and MDMA on the activity of dopamine neurons and serotonin neurons have several functional implications. First, previous studies focus on the effect of cocaine on dopamine levels59 and that of MDMA on serotonin levels60. Our data indicate that we should also consider the role of serotonin in cocaine-induced behaviors and that of dopamine in MDMA-induced behaviors. At least, some animal studies suggest a role of serotonin transporters in cocaine-related reward behaviors11,12,13. Second, VTA dopamine neurons and DRN serotonin neurons respond phasically to rewards and reward predicting cues9,10. It is interesting to test whether cocaine or MDMA would prevent these neurons from rapidly responding to external reward-related signals. Thus far this issue has not been fully resolved. Acute exposure of cocaine would enhance the dopamine release within NAc shell during a cue-cocaine Pavlovian training61. However, chronic use of cocaine would dramatically dampen the outflow of dopamine within NAc responding to drug-delivered cues, which then promotes the escalation of cocaine intake62. Third, glutamate is coreleased from subsets of dopamine neurons63,64 and serotonin neurons8,45. Cocaine could thus enhance extracellular dopamine and serotonin, and simultaneously suppress glutamate corelease owing to the auto-inhibition, which may underpin certain drug-associated behaviors.
Current views of drug abuse propose a unitary theory of addiction based on the midbrain dopamine system. However, more balanced researches are in need with careful characterization of similarities and differences in neurophysiological changes induced by different classes of drugs69. Here we revealed that different drugs dramatically and differentially regulate the activity of both VTA dopamine neurons and DRN serotonin neurons, suggesting that drugs of abuse could have distinct behavioral impacts in addition to their common effects on dopamine release. Especially, the opposing effects of heroin and cocaine over the activity of VTA dopamine neurons may help understand the opposite effects on the morphology of dopamine neurons70 and the differential engagement of direct and indirect pathways69 across opiates and psychostimulants. In future studies, monitoring the response of VTA dopamine neurons and DRN serotonin neurons within the context of drug self-administration may reveal similarities and differences across different drugs to interact with internal state and drug-taking environment to induce addictive-like behaviors71.
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