Dopaminergic Cells Of Ventral Tegmental Area Research Articles

Intermittent Cocaine Self‐Administration but not Passive Cocaine Infusions Reduces H‐Current in Putative Dopaminergic Neurons of the Ventral Tegmental Area

Intermittent Access (IntA) cocaine self‐administration is a protocol suggested to better simulate human drug use patterns due to its temporal dynamics of drug administration. IntA is also known to produce incentive salience and psychomotor sensitization. Dopaminergic (DA) neurons display a prominent mixed cation current conductance known as the hyperpolarization‐activated cyclic nucleotide current, or Ih, which contributes to neural processes such as resting membrane potential, firing frequency modulation, and synaptic integration. Previous results from our laboratory demonstrated that Ih amplitude is reduced significantly after cocaine sensitization. This Ih reduction resulted in an increased temporal summation, mean depolarization, and excitatory postsynaptic potential (EPSP) amplitude, all factors related to an enhanced excitability state. Since the cocaine sensitization model involves noncontingent drug injections administered by the experimenter, it is crucial to determine if electrophysiological changes in ventral tegmental area (VTA) DA cells are present when drugs are self‐administered (contingent). In the present study, we explored if DA neurons present an alteration in Ih after exposure to IntA. We hypothesize that VTA DA cells’ Ih modulation is dependent on the associative learning processes acquired during operant conditioning. Using the whole‐cell patch‐clamp technique in brain slices, we investigated the effects of cocaine IntA, and passive cocaine infusions (yoked controls) on Ihamplitude and rebound excitability. Our results demonstrate that an IntA protocol, but not passive cocaine infusions, produces a significant Ih amplitude reduction (P<0.05). No differences in rebound action potentials (APs) were observed. A depolarizing current protocol showed a significant increase in the number of APs (P<0.05). These results suggest that Ih modulation and intrinsic activity regulation are dependent on associative learning to drug cues.

Superior Colliculus GABAergic Neurons Are Essential for Acute Dark Induction of Wakefulness in Mice

Sleep is regulated by homeostatic process and circadian clock. Light indirectly modulates sleep by entraining the circadian clock to the solar day. Light can also influence sleep independent of photo-entrainment [1]. An acute light exposure could induce sleep, and an acute dark pulse could increase wakefulness in nocturnal animals [1, 2]. The photoreceptors and cell types in the retina that mediate light and dark effects on sleep are well characterized [1-4]. A few studies have explored the brain region involved in acute light induction of sleep. Fos expression and nonspecific lesions suggest that the superior colliculus (SC) may play a role in acute light induction of sleep [2, 5]. In contrast, the brain area and neural circuits mediating acute dark induction of wakefulness are unknown. Here, we demonstrated that retina ganglion cells (RGCs) had direct innervations on the GABAergic neurons in the mouse SC, and the activities of these cells were inhibited by an acute dark pulse, but not influenced by a light pulse. Moreover, ablating SC GABAergic neurons abolished the acute dark induction of wakefulness, but not light induction of sleep. Based on optogenetic and electrophysiological experiments, we found that SC GABAergic neurons formed monosynaptic functional connections with dopaminergic neurons in the ventral tegmental area (VTA). Selective lesions of VTA dopaminergic cells totally abolished acute dark induction of wakefulness without affecting the lightinduction of sleep. Collectively, our findings uncover a fundamental role for a retinal-SC GABAergic-VTA dopaminergic circuit in acute dark induction of wakefulness and indicate that the dark and light signals affect sleep-wake behaviors through distinct pathways.

A Hypothalamic Midbrain Pathway Essential for Driving Maternal Behaviors

Maternal behaviors are essential for the survival of the young. Previous studies implicated the medial preoptic area (MPOA) as an important region for maternal behaviors, but details of the maternal circuit remain incompletely understood. Here we identify estrogen receptor alpha (Esr1)-expressing cells inthe MPOA as key mediators of pup approach andretrieval. Reversible inactivation of MPOAEsr1+ cells impairs those behaviors, whereas optogenetic activation induces immediate pup retrieval. Invivo recordings demonstrate preferential activation of MPOAEsr1+ cells during maternal behaviors and changes in MPOA cell responses across reproductive states. Furthermore, channelrhodopsin-assisted circuit mapping reveals a strong inhibitory projection from MPOAEsr1+ cells to ventral tegmental area (VTA) non-dopaminergic cells. Pathway-specific manipulations reveal that this projection is essential for driving pup approach and retrieval and that VTA dopaminergic cells are reliably activated during those behaviors. Altogether, this study provides new insight into the neural circuit that generates maternal behaviors.

Synaptic function and plasticity in identified inhibitory inputs onto VTA dopamine neurons.

Ventral tegmental area (VTA) dopaminergic neurons are key components of the reward pathway, and their activity is powerfully controlled by a diverse array of inhibitory GABAergic inputs. Two major sources of GABAergic nerve terminals within the VTA are local VTA interneurons and neurons in the rostromedial tegmental nucleus (RMTg). Here, using optogenetics, we compared synaptic properties of GABAergic synapses on VTA dopamine neurons using selective activation of afferents that originate from these two cell populations. We found little evidence of co-release of glutamate from either input, but RMTg-originating synaptic currents were reduced by strychnine, suggesting co-release of glycine and GABA. VTA-originating synapses displayed a lower initial release probability, and at higher frequency stimulation, short-term depression was more marked in VTA- but not RMTg-originating synapses. We previously reported that nitric oxide (NO)-induced potentiation of GABAergic synapses on VTA dopaminergic cells is lost after exposure to drugs of abuse or acute stress; in these experiments, multiple GABAergic afferents were simultaneously activated by electrical stimulation. Here we found that optogenetically-activated VTA-originating synapses on presumptive dopamine neurons also exhibited NO-induced potentiation, whereas RMTg-originating synapses did not. Despite providing a robust inhibitory input to the VTA, RMTg GABAergic synapses are most likely not those previously shown by our work to be persistently altered by addictive drugs and stress. Our work emphasises the idea that dopamine neuron excitability is controlled by diverse inhibitory inputs expected to exert varying degrees of inhibition and to participate differently in a range of behaviours.

GABAergic Neurons as Putative Neurochemical Substrate Mediating Aversive Effects of Nicotine.

Nicotine, the main addictive component of tobacco smoke, has both rewarding and aversive properties. Recent studies have suggested that GABAergic neurons, one of the main neurochemical components of the reward-addiction circuitry, may also play a role in the aversive responses to nicotine. In the present study of transgenic mice expressing Green Fluorescent Protein (GFP) in Glutamate Decarboxylase 67 (GAD67) neurons, we hypothesized that a subpopulation of GABAergic neurons in the Ventral Tegmental Area (VTA) are the targets of aversive doses of nicotine in the CNS. We tested this hypothesis using c-Fos immunohistochemical techniques to identify GAD67-GFP positive cells within the VTA, that are activated by a single intraperitoneal (i.p.) injection of a low (40 ug/kg) or a high (2 mg/kg) dose of nicotine. We also assessed the anatomical location of GAD67-GFP positive cells with respect to tyrosine hydroxylase (TH) Immunoreactive (IR) dopaminergic cells in VTA. Consistent with our previous studies low- and high-dose nicotine both induced c-Fos activation of various intensities at multiple sites in VTA. Double labeling of c-Fos activated cells with GAD67-GFP positive cells identified a subpopulation of GABAergic neurons in Substantia Nigra Compact part Medial tier (SNCM) that were activated by high- but not by low-dose nicotine. Of 217 GABAergic cells counted at this site, 48.9% exhibited nicotine induced c-fos immunoreactivity. GAD67-GFP positive cells in other regions of VTA were not activated by the nicotine doses tested. Double labeling of GAD67-GFP positive cells with TH IR cells showed that the GABAergic neurons that were activated by high-dose nicotine were located in close proximity to the dopaminergic neurons of substantia nigra compact part and VTA. Dose-dependent activation of GAD67-GFP positive neurons in SNCM, by a nicotine dose known to produce aversive responses, implies that GABAergic neurons at these sites may be an important component of the nicotine aversive circuitry.

Circadian Rhythms and Substance Abuse: Chronobiological Considerations for the Treatment of Addiction.

Reward-related learning, including that associated with drugs of abuse, is largely mediated by the dopaminergic mesolimbic pathway. Mesolimbic neurophysiology and motivated behavior, in turn, are modulated by the circadian timing system which generates ∼24-h rhythms in cellular activity. Both drug taking and seeking and mesolimbic dopaminergic neurotransmission can vary widely over the day. Moreover, circadian clock genes are expressed in ventral tegmental area dopaminergic cells and in mesolimbic target regions where they can directly modulate reward-related neurophysiology and behavior. There also exists a reciprocal influence between drug taking and circadian timing as the administration of drugs of abuse can alter behavioral rhythms and circadian clock gene expression in mesocorticolimbic structures. These interactions suggest that manipulations of the circadian timing system may have some utility in the treatment of substance abuse disorders. Here, the literature on bidirectional interactions between the circadian timing system and drug taking is briefly reviewed, and potential chronotherapeutic considerations for the treatment of addiction are discussed.

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.

Persistent Adaptations in Afferents to Ventral Tegmental Dopamine Neurons after Opiate Withdrawal.

Dopaminergic (DA) cells of the ventral tegmental area (VTA) are the origin of a brain reward system and are critically involved in drug abuse. Morphine has long been known to affect VTA DA cells via GABAergic interneurons. Recently, GABAergic neurons caudal to the VTA were discovered and named the tail of VTA (tVTA). Here, we show that tVTA GABA neurons lose their capacity to disinhibit, but not to inhibit, VTA DA cells after chronic opiate exposure. The failure of disinhibition was associated with a loss of glutamatergic input to DA neurons after chronic morphine. These findings reveal mechanisms by which the tVTA may play a key role in long-term negative affective states during opiate withdrawal.

Glutamatergic regulation of ghrelin-induced activation of the mesolimbic dopamine system

Recently, we demonstrated that the central ghrelin signalling system, involving the ghrelin receptor (GHS-R1A), is important for alcohol reinforcement. Ghrelin targets a key mesolimbic circuit involved in natural as well as drug-induced reinforcement, that includes a dopamine projection from the ventral tegmental area (VTA) to the nucleus accumbens. The aim of the present study was to determine whether it is possible to suppress ghrelin's effects on this mesolimbic dopaminergic pathway can be suppressed, by interrupting afferent inputs to the VTA dopaminergic cells, as shown previously for cholinergic afferents. Thus, the effects of pharmacological suppression of glutamatergic, orexin A and opioid neurotransmitter systems on ghrelin-induced activation of the mesolimbic dopamine system were investigated. We found in the present study that ghrelin-induced locomotor stimulation was attenuated by VTA administration of the N-methyl-D-aspartic acid receptor antagonist (AP5) but not by VTA administration of an orexin A receptor antagonist (SB334867) or by peripheral administration of an opioid receptor antagonist (naltrexone). Intra-VTA administration of AP5 also suppressed the ghrelin-induced dopamine release in the nucleus accumbens. Finally the effects of peripheral ghrelin on locomotor stimulation and accumbal dopamine release were blocked by intra-VTA administration of a GHS-R1A antagonist (BIM28163), indicating that GHS-R1A signalling within the VTA is required for the ghrelin-induced activation of the mesolimbic dopamine system. Given the clinical knowledge that hyperghrelinemia is associated with addictive behaviours (such as compulsive overeating and alcohol use disorder) our finding highlights a potential therapeutic strategy involving glutamatergic control of ghrelin action at the level of the mesolimbic dopamine system.

Cocaine-mediated synaptic potentiation is absent in VTA neurons from mGlu5-deficient mice

Drugs of abuse have the ability to instantiate plastic adaptations within the central nervous system, and this property may relate to the development and persistence of addiction. In this context, a single exposure to cocaine in rodents may induce synaptic plasticity by increasing the AMPA/NMDA receptor excitatory post-synaptic current (EPSC) amplitude ratio in dopaminergic cells of the ventral tegmental area (VTA). Here, we examine the role of the metabotropic glutamate 5 (mGlu5) receptor in this regard using a genetic mouse model. The control AMPA/NMDA EPSC ratio is reduced in mGlu5-deficient mice compared to wild-types. Moreover, cocaine-induced enhancement of this EPSC ratio is also absent in mutant mice, which suggests that mGlu5 receptors are required for single-dose cocaine-induced plasticity onto VTA cells. While the temporal profile of hyperactivity to acute cocaine is altered in mGlu5-deficient mice; these mice still develop and express sensitized psychomotor responses to cocaine. These data suggest that the mGlu5 receptor is required for cocaine-induced plasticity in VTA dopaminergic cells. In contrast, the mGlu5 receptor may not be essential for psychostimulant behavioural sensitization; although it probably impacts other aspects drug addiction, such as motivation to self-administer.

Peripheral electrical stimulation reversed the cell size reduction and increased BDNF level in the ventral tegmental area in chronic morphine-treated rats

Chronic morphine administration induces functional and morphological alterations in the mesolimbic dopamine system (MLDS), which is believed to be the neurobiological substrate of opiate addiction. Our previous studies have demonstrated that peripheral electrical stimulation (PES) can suppress morphine withdrawal syndrome and morphine-induced conditioned place preference (CPP) in rats. The present study was designed to investigate if PES could reverse the cell size reduction induced by chronic morphine treatment in the ventral tegmental area (VTA), which is an important area of the MLDS. Immunohistochemical observations showed that the cell size of dopaminergic neurons in the VTA reduced significantly in the chronic morphine-treated rats with a concomitant decrease in the number of BDNF-positive cells compared to the saline-treated rats. A much milder morphological change, accompanying with an increased number of BDNF-positive cells, was observed in dopaminergic neurons in the rats that received repeated 100 Hz PES after morphine withdrawal. In another experiment, enzyme-linked immunosorbent assay (ELISA) reconfirmed a significant up-regulation of BDNF protein level in the VTA in the rats received 100 Hz PES after morphine abstinence. These results indicate that PES could facilitate the morphological recovery of the VTA dopaminergic cells damaged by chronic morphine treatment and up-regulate the BDNF protein level in the VTA. Activation of endogenous BDNF by PES may play a role in the recovery of the injured dopaminergic neurons in the morphine addictive rats.

Striatal expression of GDNF and differential vulnerability of midbrain dopaminergic cells

Glial cell line-derived neurotrophic factor (GDNF) is a member of the transforming growth factor-beta superfamily that when exogenously administrated exerts a potent trophic action on dopaminergic (DA) cells. Although we know a lot about its signalling mechanisms and pharmacological effects, physiological actions of GDNF on the adult brain remain unclear. Here, we have used morphological and molecular techniques, and an experimental model of Parkinson's disease in rats, to investigate whether GDNF constitutively expressed in the adult mesostriatal system plays a neuroprotective role on midbrain DA cells. We found that although all midbrain DA cells express both receptor components of GDNF (GFRalpha1 and Ret), those in the ventral tegmental area (VTA) and rostromedial substantia nigra (SNrm) also contain GDNF but not GDNFmRNA. The levels of GDNFmRNA are significantly higher in the ventral striatum (vSt), the target region of VTA and SNrm cells, than in the dorsal striatum (dSt), the target region of DA cells in the caudoventral substantia nigra (SNcv). After fluoro-gold injection in striatum, VTA and SNrm DA cells show triple labelling for tyrosine hydroxylase, GDNF and fluoro-gold, and after colchicine injection in the lateral ventricle, they become GDNF-immunonegative, suggesting that GDNF in DA somata comes from their striatal target. As DA cells in VTA and SNrm are more resistant than those in SNcv to intracerebroventricular injection of 6-OHDA, as occurs in Parkinson's disease, we can suggest that the fact that they project to vSt, where GDNF expression is significantly higher than in the dSt, is a neuroprotective factor involved in the differential vulnerability of midbrain DA neurons.

Selective serotonin reuptake inhibitors reduce the spontaneous activity of dopaminergic neurons in the ventral tegmental area

Electrophysiological techniques were used to study the effects of paroxetine, sertraline, and fluvoxamine on the basal activity of dopaminergic neurons in the ventral tegmental area (VTA) of rats. Acute i.v. administrations of paroxetine (20–1280 μg/kg), sertraline (20–1280 μg/kg), and fluvoxamine (20–1280 μg/kg) caused a slight but significant reduction in the firing rate of the VTA dopaminergic cells studied. Paroxetine produced a maximal inhibitory effect of 10 ± 11% at the cumulative dose of 160 μg/kg. Sertraline induced a dose-related inhibition of VTA dopaminergic neurons, which reached its maximum (10 ± 7%) at the cumulative dose of 1280 μg/kg. The effect of fluvoxamine on the basal firing rate of VTA dopaminergic neurons was more pronounced as compared to that of paroxetine and sertraline, in that it produced a maximal inhibition of 17 ± 12% at the cumulative dose of 1280 μg/kg. Acute i.v. injections of paroxetine (20–1280 μg/kg), sertraline (20–1280 μg/kg), and fluvoxamine (20–5120 μg/kg) caused a dose-dependent decrease in the basal firing rate of serotonergic neurons in the dorsal raphé nucleus (DRN). Paroxetine and sertraline stopped the spontaneous firing of serotonergic neurons at the cumulative dose of 1280 μg/kg, whereas fluvoxamine reached the same effect only at the cumulative dose of 5120 μg/kg. Pretreatment with the 5-HT 1A receptor antagonist tertatolol (1 mg/kg, i.v.) reduced the inhibitory effects of paroxetine, fluvoxamine, and sertraline on the basal activity of serotonergic neurons in the DRN. Administration of tertatolol induced a 15-fold increase in the ED 50 for fluvoxamine. The antagonistic effect of tertatolol was much less evident in blocking the inhibitory action exerted by paroxetine and sertraline on the activity of serotonergic neurons. Pretreatment with tertatolol (1 mg/kg, i.v.) potentiated the inhibitory effect of fluvoxamine on the basal activity of VTA dopaminergic neurons. Tertatolol did not affect the inhibitory action exerted by paroxetine and sertraline on these neurons. It is concluded that inhibition of the basal firing rate of dopaminergic neurons in the VTA is a common characteristic of selective serotonin reuptake inhibitors (SSRIs). The effects of SSRIs on VTA dopaminergic cell activity might be relevant for their therapeutic action and may explain the origin of the reported cases of akathisia.

Differential effects of acute and chronic fluoxetine administration on the spontaneous activity of dopaminergic neurones in the ventral tegmental area.

1. Electrophysiological techniques were used to study the effects of fluoxetine and citalopram on the basal activity of dopaminergic neurones in the ventral tegmental area (VTA) and substantia nigra, pars compacta (SNc) of rats. 2. Acute i.v. injection of fluoxetine (20-1280 micrograms kg-1) caused a dose-dependent inhibition of the firing rate of VTA dopaminergic neurones, but did not affect the activity of dopaminergic cells in the SNc. Citalopram (20-1280 micrograms kg-1, i.v.) inhibited the firing rate of dopaminergic neurones in the VTA, but its effect (maximal inhibition: 14 +/- 7%) was less pronounced than that of fluoxetine (maximal inhibition: 34 +/- 7%). 3. Pretreatment with mesulergine (80 micrograms kg-1, i.v.), a 5-hydroxytryptamine2C/2B (5-HT2C/2B) receptor antagonist, blocked the inhibitory effect of fluoxetine on VTA dopaminergic cells. Selective lesions of 5-hydroxytryptaminergic neurones by the neurotoxin, 5,7-dihydroxytryptamine (5,7-DHT), abolished the fluoxetine-induced reduction of VTA dopaminergic activity. 4. In a series of experiments, fluoxetine (10 mg kg-1, i.p.) was administered once daily for 21 consecutive days. Acute i.v. administration of fluoxetine (20-1280 micrograms kg-1, 72 h after the last i.p. injection) did not cause any change in the basal firing rate of VTA dopaminergic neurones in treated rats, whereas it induced the typical inhibitory effect in control animals. A group of rats chronically treated with fluoxetine, received i.v. m-chlorophenylpiperazine (mCPP; 10-320 micrograms kg-1), a 5-HT2C/2B receptor agonist. This drug significantly inhibited VTA dopaminergic function in control rats, but did not modify the basal activity of dopaminergic cells in animals given chronic fluoxetine. 5 It is concluded that fluoxetine inhibits dopaminergic function in the VTA by enhancing the synaptic levels of 5-HT, which possibly acts through the 5-HT2C/2B receptor subtype. Repeated treatment with fluoxetine induces tolerance to its inhibitory effect on dopaminergic activity, possibly as a consequence of down-regulation of 5-HT2C/2B receptors. The effects of fluoxetine on VTA dopaminergic cell activity might be relevant for its therapeutic actions and may explain the origin of the reported cases of akathisia.