Constitutive cannabinoid 1 and mu opioid receptor activity in the ventral tegmental area: occurrence, function and therapeutic relevance

Cannabinoid 1 receptors (CB1Rs) play a crucial role in regulating systems dedicated to processing rewards and emotions. It was known that in artificial systems, CB1Rs can exhibit activity that is independent of the typical agonist-driven form. However, it remained largely unclear whether this constitutive activity also occurred in native tissue (e.g. the brain), and if so, what role it plays in neurotransmission and behavior. In this thesis we have taken a multi-disciplinary approach to show that CB1R inverse agonism (which interferes with both constitutive and agonist-driven activation) and neutral antagonism (which only interferes with agonist-driven activation) has markedly different effects on neurotransmission in the dopamine reward system in brain slices, dopamine neuron activity in freely moving rats, and processing of rewards and emotions. The inverse agonist interfered with reward processing and induced anxiety, whereas the neutral antagonist did neither. Strikingly, the neutral antagonist and inverse agonist were equally able to reduce weight gain and food intake. This is particularly of interest since the inverse agonist (rimonabant) had been previously marketed as a weight loss compound, but was discontinued due to side effects in the realms of anxiety and depression. Our findings therefore suggest that CB1R neutral antagonists may provide a safer and efficacious alternative to the discontinued CB1R inverse agonists. In a similar vein, we show that the mu opioid receptor (MOR), which is closely akin to the CB1R, is also constitutively active in the dopamine reward system. We provide evidence that, during a state of morphine withdrawal, this MOR constitutive activity is enhanced and likely serves a protective function that limits withdrawal symptoms. MOR antagonists with inverse agonistic profiles are currently in use to treat narcotic overdose and drug addiction. Problematically, this can lead to severe side effects. Our findings provide evidence that MOR neutral antagonists may also prove to be better therapeutic alternatives with fewer side effects, as they leave the potentially protective MOR constitutive activity during opioid withdrawal intact

Cocaine withdrawal reduces GABA_B R transmission at entopeduncular nucleus - lateral habenula synapses.

Lateral habenula (LHb) hyperactivity plays a pivotal role in the emergence of negative emotional states, including those occurring during withdrawal from addictive drugs. We have previously implicated cocaine-driven adaptations at synapses from the entopeduncular nucleus (EPN) to the LHb in this process. Specifically, ionotropic GABA <sub>A</sub> receptor (R)-mediated neurotransmission at EPN-to-LHb synapses is reduced during cocaine withdrawal, due to impaired vesicle filling. Recent studies have shown that metabotropic GABA <sub>B</sub> R signaling also controls LHb activity, although its role at EPN-to-LHb synapses during drug withdrawal is unknown. Here, we predicted that cocaine treatment would reduce GABA <sub>B</sub> R-mediated neurotransmission at EPN-to-LHb synapses. We chronically treated mice with saline or cocaine, prepared brain slices after two days of withdrawal and performed voltage-clamp recordings from LHb neurons whilst optogenetically stimulating EPN terminals. Compared with controls, mice in cocaine withdrawal exhibited reduced GABA <sub>A</sub> R-mediated input to LHb neurons, and a reduced occurrence of GABA <sub>B</sub> R-signaling at EPN-to-LHb synapses. We then assessed the underlying mechanism of this decrease. Application of GABA <sub>B</sub> R agonist baclofen evoked similar postsynaptic responses in EPN-innervated LHb neurons in saline- and cocaine-treated mice. Release probability at EPN-to-LHb GABAergic synapses was also comparable between groups. However, incubating brain slices in glutamine to facilitate GABA vesicle filling, normalized GABA <sub>B</sub> R-currents at EPN-to-LHb synapses in cocaine-treated mice. Overall, we show that during cocaine withdrawal, together with reduced GABA <sub>A</sub> R transmission, also GABA <sub>B</sub> R-mediated inhibitory signaling is diminished at EPN-to-LHb synapses, likely via the same presynaptic deficit. In concert, these alterations are predicted to contribute to the emergence of drug withdrawal symptoms, facilitating drug relapse

Neural circuit adaptations during drug withdrawal - Spotlight on the lateral habenula.

Withdrawal after drug intake triggers a wealth of affective states including negative feelings reminiscent of depressive symptoms. This negative state can ultimately be crucial for relapse, a hallmark of addiction. Adaptations in a wide number of neuronal circuits underlie aspects of drug withdrawal, however causality between cellular modifications within these systems and precise behavioral phenotypes remains poorly described. Recent advances point to an instrumental role of the lateral habenula in driving depressive-like states during drug withdrawal. In this review we will discuss the general behavioral features of drug withdrawal, the importance of plasticity mechanisms in the mesolimbic systems, and the latest discoveries highlighting the implications of lateral habenula in drug addiction. We will further stress how specific interventions in the lateral habenula efficiently ameliorate depressive symptoms. Altogether, this work aims to provide a general knowledge on the cellular and circuit basis underlying drug withdrawal, ultimately speculating on potential treatment for precise aspects of addiction

Morphine withdrawal enhances constitutive μ-opioid receptor activity in the ventral tegmental area

μ-opioid receptors (MORs) in the ventral tegmental area (VTA) are pivotally involved in addictive behavior. While MORs are typically activated by opioids, they can also become constitutively active in the absence of any agonist. In the current study, we present evidence that MOR constitutive activity is highly relevant in the mouse VTA, as it regulates GABAergic input to dopamine neurons. Specifically, suppression of MOR constitutive activity with the inverse agonist KC-2-009 enhanced GABAergic neurotransmission onto VTA dopamine neurons. This inverse agonistic effect was fully blocked by the specific MOR neutral antagonist CTOP, which had no effect on GABAergic transmission itself. We next show that withdrawal from chronic morphine further increases the magnitude of inverse agonistic effects at the MOR, suggesting enhanced MOR constitutive activity. We demonstrate that this increase can be an adaptive response to the detrimental elevation in cAMP levels known to occur during morphine withdrawal. These findings offer important insights in the physiological occurrence and function of MOR constitutive activity, and have important implications for therapeutic strategies aimed at normalizing MOR signaling during addiction and opioid overdose

The continued need for animals to advance brain research

Policymakers aim to move toward animal-free alternatives for scientific research and have introduced very strict regulations for animal research. We argue that, for neuroscience research, until viable and translational alternatives become available and the value of these alternatives has been proven, the use of animals should not be compromised

The continued need for animals to advance brain research

Policymakers aim to move toward animal-free alternatives for scientific research and have introduced very strict regulations for animal research. We argue that, for neuroscience research, until viable and translational alternatives become available and the value of these alternatives has been proven, the use of animals should not be compromised