α2A adrenergic receptors highly expressed in mesoprefrontal dopamine neurons.

α2 adrenoreceptors (α2-ARs) play a key role in the control of noradrenaline and dopamine release in the medial prefrontal cortex (mPFC). Here, using UV-laser microdissection-based quantitative mRNA expression in individual neurons we show that in hTH-GFP rats, a transgenic line exhibiting intense and specific fluorescence in dopaminergic (DA) neurons, α2A adrenoreceptor (α2A-AR) mRNA is expressed at high and low levels in DA cells in the ventral tegmental area (VTA) and substantia nigra compacta (SNc), respectively. Confocal microscopy fluorescence immunohistochemistry revealed that α2A-AR immunoreactivity colocalized with tyrosine hydroxylase (TH) in nearly all DA cells in the VTA and SNc, both in hTH-GFP rats and their wild-type Sprague-Dawley (SD) counterparts. α2A-AR immunoreactivity was also found in DA axonal projections to the mPFC and dorsal caudate in the hTH-GFP and in the anterogradely labeled DA axonal projections from VTA to mPFC in SD rats. Importantly, the α2A-AR immunoreactivity localized in the DA cells of VTA and in their fibers in the mPFC was much higher than that in DA cells of SNc and their fibers in dorsal caudate, respectively. The finding that α2A-ARs are highly expressed in the cell bodies and axons of mesoprefrontal dopaminergic neurons provides a morphological basis to the vast functional evidence that somatodendritic and nerve-terminal α2A-AR receptors control dopaminergic activity and dopamine release in the prefrontal cortex. This finding raises the question whether α2A-ARs might function as autoreceptors in the mesoprefrontal dopaminergic neurons, replacing the lack of D2 autoreceptors.

Influence of caffeine on 3,4-methylenedioxymethamphetamine-induced dopaminergic neuron degeneration and neuroinflammation is age-dependent.

Previous studies have demonstrated that caffeine administration to adult mice potentiates glial activation induced by 3,4-methylenedioxymethamphetamine (MDMA). As neuroinflammatory response seems to correlate with neurodegeneration, and the young brain is particularly vulnerable to neurotoxicity, we evaluated dopamine neuron degeneration and glial activation in the caudate-putamen (CPu) and substantia nigra pars compacta (SNc) of adolescent and adult mice. Mice were treated with MDMA (4 × 20 mg/kg), alone or with caffeine (10 mg/kg). Interleukin (IL)-1ß, tumor necrosis factor (TNF)-α, neuronal nitric oxide synthase (nNOS) were evaluated in CPu, whereas tyrosine hydroxylase (TH), glial fibrillary acidic protein, and CD11b were evaluated in CPu and SNc by immunohistochemistry. MDMA decreased TH in SNc of both adolescent and adult mice, whereas TH-positive fibers in CPu were only decreased in adults. In CPu of adolescent mice, caffeine potentiated MDMA-induced glial fibrillary acidic protein without altering CD11b, whereas in SNc caffeine did not influence MDMA-induced glial activation. nNOS, IL-1ß, and TNF-α were increased by MDMA in CPu of adults, whereas in adolescents, levels were only elevated after combined MDMA plus caffeine. Caffeine alone modified only nNOS. Results suggest that the use of MDMA in association with caffeine during adolescence may exacerbate the neurotoxicity and neuroinflammation elicited by MDMA. Previous studies have demonstrated that caffeine potentiated glial activation induced by 3,4-methylenedioxymethamphetamine (MDMA) in adult mice. In this study, caffeine was shown to potentiate MDMA-induced dopamine neuron degeneration in substantia nigra pars compacta, astrogliosis, and TNF-α levels in caudate-putamen of adolescent mice. Results suggest that combined use of MDMA plus caffeine during adolescence may worsen the neurotoxicity and neuroinflammation elicited by MDMA.

Enhanced endocannabinoid-mediated modulation of rostromedial tegmental nucleus drive onto dopamine neurons in Sardinian alcohol-preferring rats.

The progressive predominance of rewarding effects of addictive drugs over their aversive properties likely contributes to the transition from drug use to drug dependence. By inhibiting the activity of DA neurons in the VTA, GABA projections from the rostromedial tegmental nucleus (RMTg) are well suited to shift the balance between drug-induced reward and aversion. Since cannabinoids suppress RMTg inputs to DA cells and CB1 receptors affect alcohol intake in rodents, we hypothesized that the endocannabinoid system, by modulating this pathway, might contribute to alcohol preference. Here we found that RMTg afferents onto VTA DA neurons express CB1 receptors and display a 2-arachidonoylglycerol (2-AG)-dependent form of short-term plasticity, that is, depolarization-induced suppression of inhibition (DSI). Next, we compared rodents with innate opposite alcohol preference, the Sardinian alcohol-preferring (sP) and alcohol-nonpreferring (sNP) rats. We found that DA cells from alcohol-naive sP rats displayed a decreased probability of GABA release and a larger DSI. This difference was due to the rate of 2-AG degradation. In vivo, we found a reduced RMTg-induced inhibition of putative DA neurons in sP rats that negatively correlated with an increased firing. Finally, alcohol failed to enhance RMTg spontaneous activity and to prolong RMTg-induced silencing of putative DA neurons in sP rats. Our results indicate functional modifications of RMTg projections to DA neurons that might impact the reward/aversion balance of alcohol attributes, which may contribute to the innate preference observed in sP rats and to their elevated alcohol intake.

Δ9-tetrahydrocannabinol prevents methamphetamine-induced neurotoxicity.

Methamphetamine (METH) is a potent psychostimulant with neurotoxic properties. Heavy use increases the activation of neuronal nitric oxide synthase (nNOS), production of peroxynitrites, microglia stimulation, and induces hyperthermia and anorectic effects. Most METH recreational users also consume cannabis. Preclinical studies have shown that natural (Δ9-tetrahydrocannabinol, Δ9-THC) and synthetic cannabinoid CB1 and CB2 receptor agonists exert neuroprotective effects on different models of cerebral damage. Here, we investigated the neuroprotective effect of Δ9-THC on METH-induced neurotoxicity by examining its ability to reduce astrocyte activation and nNOS overexpression in selected brain areas. Rats exposed to a METH neurotoxic regimen (4 × 10 mg/kg, 2 hours apart) were pre- or post-treated with Δ9-THC (1 or 3 mg/kg) and sacrificed 3 days after the last METH administration. Semi-quantitative immunohistochemistry was performed using antibodies against nNOS and Glial Fibrillary Acidic Protein (GFAP). Results showed that, as compared to corresponding controls (i) METH-induced nNOS overexpression in the caudate-putamen (CPu) was significantly attenuated by pre- and post-treatment with both doses of Δ9-THC (-19% and -28% for 1 mg/kg pre- and post-treated animals; -25% and -21% for 3 mg/kg pre- and post-treated animals); (ii) METH-induced GFAP-immunoreactivity (IR) was significantly reduced in the CPu by post-treatment with 1 mg/kg Δ9-THC1 (-50%) and by pre-treatment with 3 mg/kg Δ9-THC (-53%); (iii) METH-induced GFAP-IR was significantly decreased in the prefrontal cortex (PFC) by pre- and post-treatment with both doses of Δ9-THC (-34% and -47% for 1 mg/kg pre- and post-treated animals; -37% and -29% for 3 mg/kg pre- and post-treated animals). The cannabinoid CB1 receptor antagonist SR141716A attenuated METH-induced nNOS overexpression in the CPu, but failed to counteract the Δ9-THC-mediated reduction of METH-induced GFAP-IR both in the PFC and CPu. Our results indicate that Δ9-THC reduces METH-induced brain damage via inhibition of nNOS expression and astrocyte activation through CB1-dependent and independent mechanisms, respectively.

PPARα regulates cholinergic-driven activity of midbrain dopamine neurons via a novel mechanism involving α7 nicotinic acetylcholine receptors.

Ventral tegmental area dopamine neurons control reward-driven learning, and their dysregulation can lead to psychiatric disorders. Tonic and phasic activity of these dopaminergic neurons depends on cholinergic tone and activation of nicotinic acetylcholine receptors (nAChRs), particularly those containing the ß2 subunit (ß2*-nAChRs). Nuclear peroxisome proliferator-activated receptors type-α (PPARα) tonically regulate ß2*-nAChRs and thereby control dopamine neuron firing activity. However, it is unknown how and when PPARα endogenous ligands are synthesized by dopamine cells. Using ex vivo and in vivo electrophysiological techniques combined with biochemical and behavioral analysis, we show that activation of α7-nAChRs increases in the rat VTA both the tyrosine phosphorylation of the ß2 subunit of nAChRs and the levels of two PPARα endogenous ligands in a Ca(2+)-dependent manner. Accordingly, in vivo production of endogenous PPARα ligands, triggered by α7-nAChR activation, blocks in rats nicotine-induced increased firing activity of dopamine neurons and displays antidepressant-like properties. These data demonstrate that endogenous PPARα ligands are effectors of α7-nAChRs and that their neuromodulatory properties depend on phosphorylation of ß2*-nAChRs on VTA dopamine cells. This reveals an autoinhibitory mechanism aimed at reducing dopamine cell overexcitation engaged during hypercholinergic drive. Our results unveil important physiological functions of nAChR/PPARα signaling in dopamine neurons and how behavioral output can change after modifications of this signaling pathway. Overall, the present study suggests PPARα as new therapeutic targets for disorders associated with unbalanced dopamine-acetylcholine systems.