Jennifer L. Short

Combined antagonism of glutamate mGlu5 and adenosine A2A receptors interact to regulate alcohol-seeking in rats

Adenosine and glutamate have been implicated as mediators involved in the self-administration of alcohol. In the present study we sought to determine whether adenosine receptors could interact with metabotropic glutamate receptors to regulate operant responding for alcohol and also the integration of the salience of alcohol-paired cues. Alcohol-preferring (iP) rats were trained to self-administer alcohol under operant conditions. The availability of alcohol was paired with an olfactory cue plus a stimulus light. Rats were examined under fixed ratio responding and also following extinction under a cue-induced reinstatement paradigm. Administration of the selective adenosine A2A receptor antagonist, SCH 58261, reduced fixed ratio responding for alcohol in iP rats in a dose-related manner. Furthermore, the combination of a subthreshold dose of SCH 58261 with a subthreshold dose of the mGlu5 receptor antagonist MTEP also reduced alcohol self-administration and increased the latency to the first reinforced response, suggesting a pre-ingestive effect. Moreover, this combination of SCH 58261 and MTEP also prevented the conditioned reinstatement of alcohol-seeking elicited by the re-presentation of cues previously paired with alcohol availability. In contrast, combinations of the selective adenosine A1 receptor antagonist, DPCPX, with either SCH 58261 or MTEP had no effect on alcohol responding. Collectively, these data suggest a functional interaction between adenosine A2A and mGlu5 receptors in relation to alcohol-seeking and the integration of the drug-related cues.

Reduced startle habituation and prepulse inhibition in mice lacking the adenosine A2A receptor

Adenosine and dopamine receptors interact in the CNS to modulate behaviour, including sensorimotor gating. Prepulse inhibition (PPI) has been suggested to be an operational measure of sensorimotor gating. PPI and startle habituation are disrupted in patients with schizophrenia. In experimental animals, both parameters are modulated by dopaminergic and adenosine receptor agonists and antagonists. In the present study, we measured PPI and startle habituation in mice that lack functional adenosine A(2A) receptors. Startle amplitudes, startle habituation and PPI were significantly reduced in mice homozygous null for the adenosine A(2A) receptor (A(2A)(-/-)). In addition, differential effects of amphetamine and MK-801 on startle amplitude, startle habituation and PPI were observed between A(2A)(-/-) and wildtype controls. These data support the involvement of adenosine A(2A) receptors in regulation of PPI and startle habituation.

A differential role for the adenosine A2A receptor in opiate reinforcement vs opiate-seeking behavior.

The adenosine A2A receptor is specifically enriched in the medium spiny neurons that make up the ‘indirect’ output pathway from the ventral striatum, a structure known to have a crucial, integrative role in processes such as reward, motivation, and drug-seeking behavior. In the present study we investigated the impact of adenosine A2A receptor deletion on behavioral responses to morphine in a number of reward-related paradigms. The acute, rewarding effects of morphine were evaluated using the conditioned place preference paradigm. Operant self-administration of morphine on both fixed and progressive ratio schedules as well as cue-induced drug-seeking was assessed. In addition, the acute locomotor response to morphine as well as sensitization to morphine was evaluated. Decreased morphine self-administration and breakpoint in A2A knockout mice was observed. These data support a decrease in motivation to consume the drug, perhaps reflecting diminished rewarding effects of morphine in A2A knockout mice. In support of this finding, a place preference to morphine was not observed in A2A knockout mice but was present in wild-type mice. In contrast, robust cue-induced morphine-seeking behavior was exhibited by both A2A knockout and wild-type mice after a period of withdrawal. The acute locomotor response to morphine in the A2A knockout was similar to wild-type mice, yet A2A knockout mice did not display tolerance to chronic morphine under the present paradigm. Both genotypes display locomotor sensitization to morphine, implying a lack of a role for the A2A receptor in the drug-induced plasticity necessary for the development or expression of sensitization. Collectively, these data suggest a differential role for adenosine A2A receptors in opiate reinforcement compared to opiate-seeking.

Identification of brain nuclei implicated in cocaine-primed reinstatement of conditioned place preference: a behaviour dissociable from sensitization.

Relapse prevention represents the primary therapeutic challenge in the treatment of drug addiction. As with humans, drug-seeking behaviour can be precipitated in laboratory animals by exposure to a small dose of the drug (prime). The aim of this study was to identify brain nuclei implicated in the cocaine-primed reinstatement of a conditioned place preference (CPP). Thus, a group of mice were conditioned to cocaine, had this place preference extinguished and were then tested for primed reinstatement of the original place preference. There was no correlation between the extent of drug-seeking upon reinstatement and the extent of behavioural sensitization, the extent of original CPP or the extinction profile of mice, suggesting a dissociation of these components of addictive behaviour with a drug-primed reinstatement. Expression of the protein product of the neuronal activity marker c-fos was assessed in a number of brain regions of mice that exhibited reinstatement (R mice) versus those which did not (NR mice). Reinstatement generally conferred greater Fos expression in cortical and limbic structures previously implicated in drug-seeking behaviour, though a number of regions not typically associated with drug-seeking were also activated. In addition, positive correlations were found between neural activation of a number of brain regions and reinstatement behaviour. The most significant result was the activation of the lateral habenula and its positive correlation with reinstatement behaviour. The findings of this study question the relationship between primed reinstatement of a previously extinguished place preference for cocaine and behavioural sensitization. They also implicate activation patterns of discrete brain nuclei as differentiators between reinstating and non-reinstating mice.

Adenosine A2A receptors and their role in drug addiction

The specific events between initial presumably manageable drug intake and the development of a drug‐ addicted state are not yet known. Drugs of abuse have varying mechanisms of action that create a complex pattern of behaviour related to drug consumption, drug‐seeking, withdrawal and relapse. The neuromodulator adenosine has been shown to play a role in reward‐related behaviour, both as an independent mediator and via interactions of adenosine receptors with other receptors. Adenosine levels are elevated upon exposure to drugs of abuse and adenosine A2A receptors are enriched in brain nuclei known for their involvement in the processing of drug‐related reinforcement processing. A2A receptors are found in receptor clusters with dopamine and glutamate receptors. A2A receptors are thus ideally situated to influence the signalling of neurotransmitters relevant in the neuronal responses and plasticity that underlie the development of drug taking and drug‐seeking behaviour. In this review, we present evidence for the role of adenosine and A2A receptors in drug addiction, thereby providing support for current efforts aimed at developing drug therapies to combat substance abuse that target adenosine signalling via A2A receptors.

Receptor crosstalk: characterization of mice deficient in dopamine D1 and adenosine A2A receptors.

Here we report the development of D1A2A receptor knockout mice to investigate whether interactions between dopamine D1 and adenosine A2A receptors participate in reward-related behavior. The combined deletion of D1 and A2A receptors resulted in mice with decreased weight and appetitive processes, reduced rearing and exploratory behaviors, increased anxiety, and a significantly poorer performance on the rotarod, compared to wild-type littermates. D1A2A receptor knockout mice shared phenotypic similarities with mice deficient in D1 receptors, while also paralleling behavioral deficits seen in A2A receptor knockout mice, indicating individual components of the behavioral phenotype of the D1A2A receptor knockout attributable to the loss of both receptors. In contrast, ethanol and saccharin preference in D1A2A receptor knockout mice were distinctly different from that observed in derivative D1 or A2A receptor-deficient mice. Compared to wild types, preference and consumption of ethanol were decreased in D1A2A receptor knockout mice, the reduction in ethanol consumption greater even than that seen in D1 receptor-deficient mice. Preference and consumption of saccharin were also reduced in D1A2A receptor knockout mice, whereas saccharin preference was similar in wild-type, D1, and A2A receptor knockout mice. These data suggest an interaction of D1 and A2A receptors in the reinforcement processes underlying the intake of rewarding substances, whereby the A2A receptor seems involved in goal-directed behavior and the motor functions underlying the expression of such behaviors, and the D1 receptor is confirmed as essential in mediating motivational processes related to the repeated intake of novel substances and drugs.

The α1A‐adrenoceptor gene is required for the α1L‐adrenoceptor‐mediated response in isolated preparations of the mouse prostate

This study investigated whether deletion of the α1A‐adrenoceptor gene influences contractile responses of mouse prostate to noradrenaline. Responses of mouse prostate to noradrenaline are known to be mediated by α1L‐adrenoceptors, which are thought to be a functional phenotype of α1A‐adrenoceptor.

Cue-conditioned alcohol seeking in rats following abstinence: involvement of metabotropic glutamate 5 receptors

Background and purpose:  The current study was designed to: (i) examine whether functional interactions occur between receptors known to regulate alcohol self‐administration; and (ii) characterize relapse to alcohol seeking following abstinence.

Characterisation of central adenosine A1 receptors and adenosine transporters in mice lacking the adenosine A2a receptor

The present study was designed to assess whether adenosine A(2a) receptor knockout mice exhibit altered purine utilisation in brain nuclei. Specifically, the properties of adenosine transporters and adenosine A(1) receptors were characterised in brain membranes and on slide-mounted sections. The B(MAX) for [(3)H]nitrobenzylthioinosine ([(3)H]NBTI) binding (adenosine transporter density) was significantly reduced in brainstem membranes of homozygotes (560+/-52 fmol/mg protein, n=5, P<0.05, Kruskal-Wallis ANOVA) compared to wildtype (1239+/-213 fmol/mg protein) and heterozygous mice (1300+/-558 fmol/mg protein). Quantitative autoradiography data indicated that [(3)H]NBTI binding in the medulla oblongata of heterozygous mice was seen to decrease significantly (P<0.05) in the subpostremal nucleus tractus solitarius (NTS), medial NTS, inferior olive and area postrema (AP). On the other hand, in the homozygous mice a decrease was seen in the medial NTS and AP. In the pons, [(3)H]1, 3-dipropyl-8-cyclopentylxanthine ([(3)H]DPCPX) (adenosine A(1) receptor density) binding increased significantly (P<0.05, Kruskal-Wallis ANOVA) in the lateral parabrachial nucleus, caudal pontine reticular nucleus and locus coeruleus of homozygotes compared to wildtype. In higher brain centres, [(3)H]NBTI binding was reduced in the paraventricular thalamic nucleus of both heterozygous and homozygous mice, whereas [(3)H]DPCPX binding was reduced in the hippocampus and lateral hypothalamus of heterozygotes. In homozygotes, [(3)H]DPCPX binding in the hippocampus increased compared to wildtype mice. The present study indicates that deletion of the A(2a) receptor may have contributed to region-specific compensatory changes in purine utilisation in brain nuclei associated with autonomic, neuroendocrine and behavioural regulation.

Genetic interdependence of adenosine and dopamine receptors: Evidence from receptor knockout mice

Dopamine and adenosine receptors are known to share a considerable overlap in their regional distribution, being especially rich in the basal ganglia. Dopamine and adenosine receptors have been demonstrated to exhibit a parallel distribution on certain neuronal populations, and even when not directly co-localized, relationships (both antagonistic and synergistic) have been described. This study was designed to investigate dopaminergic and purinergic systems in mice with ablations of individual dopamine or adenosine receptors. In situ hybridization histochemistry and autoradiography was used to examine the level of mRNA and protein expression of specific receptors and transporters in dopaminergic pathways. Expression of the mRNA encoding the dopamine D2 receptor was elevated in the caudate putamen of D1, D3 and A2A receptor knockout mice; this was mirrored by an increase in D2 receptor protein in D1 and D3 receptor knockout mice, but not in A2A knockout mice. Dopamine D1 receptor binding was decreased in the caudate putamen, nucleus accumbens, olfactory tubercle and ventral pallidum of D2 receptor knockout mice. In substantia nigra pars compacta, dopamine transporter mRNA expression was dramatically decreased in D3 receptor knockout mice, but elevated in A2A receptor knockout mice. All dopamine receptor knockout mice examined exhibited increased A2A receptor binding in the caudate putamen, nucleus accumbens and olfactory tubercle. These data are consistent with the existence of functional interactions between dopaminergic and purinergic systems in these reward and motor-related brain regions.