Exciting news about A2A receptors.
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Adenosine↗
Biomedical subjects
Per Svenningsson
Publications and source records attributed to Per Svenningsson.
Adenosine-dopamine interactions: development of a concept and some comments on therapeutic possibilities.
This brief review presents a personal perspective on the historical development of the current knowledge about the biologically important concept of functional antagonism between adenosine A2A and dopamine D2 receptors in caudate-putamen, accumbens, and tuberculum olfactorium. In the 1970s, studies of dopamine actions suggested an unexpected role of adenosine. Developments during the next decade substantiated this finding and demonstrated that a subform of adenosine A2 receptors was enriched in the basal ganglia. Cloning of adenosine receptors provided better tools for cellular localization and showed that A2A receptors are closely associated with D2 receptors. Distinct functional interactions at several levels were discovered, and there is now strong evidence that A2A receptors are tonically active and modified by dopamine acting at D2 receptors. Development of selective antagonists and knockout mice have highlighted the potential usefulness of A2A antagonists in decreasing symptoms and progression of Parkinson's disease-something that has also been vindicated by careful epidemiologic studies. There are issues of efficacy and potential side effects that need to be resolved, but the future looks bright.
Adenosine↗
Diverse psychotomimetics act through a common signaling pathway.
Three distinct classes of drugs: dopaminergic agonists (such as D-amphetamine), serotonergic agonists (such as LSD), and glutamatergic antagonists (such as PCP) all induce psychotomimetic states in experimental animals that closely resemble schizophrenia symptoms in humans. Here we implicate a common signaling pathway in mediating these effects. In this pathway, dopamine- and an adenosine 3',5'-monophosphate (cAMP)-regulated phospho-protein of 32 kilodaltons (DARPP-32) is phosphorylated or dephosphorylated at three sites, in a pattern predicted to cause a synergistic inhibition of protein phosphatase-1 and concomitant regulation of its downstream effector proteins glycogen synthesis kinase-3 (GSK-3), cAMP response element-binding protein (CREB), and c-Fos. In mice with a genetic deletion of DARPP-32 or with point mutations in phosphorylation sites of DARPP-32, the effects of D-amphetamine, LSD, and PCP on two behavioral parameters-sensorimotor gating and repetitive movements-were strongly attenuated.
1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
Effect of methylphenidate on dopamine/DARPP signalling in adult, but not young, mice.
Methylphenidate (MPH), a dopamine uptake inhibitor, is the most commonly prescribed drug for the treatment of attention-deficit/hyperactivity disorder (ADHD) in children. We examined the effect of MPH on dopamine- and cAMP-regulated phosphoprotein, Mr 32 kDa (DARPP-32) phosphorylation at Thr34 (PKA-site) and Thr75 (Cdk5-site) using neostriatal slices from young (14-15- and 21-22-day-old) and adult (6-8-week-old) mice. MPH increased DARPP-32 Thr34 phosphorylation and decreased Thr75 phosphorylation in slices from adult mice. The effect of MPH was blocked by a dopamine D1 antagonist, SCH23390. In slices from young mice, MPH did not affect DARPP-32 phosphorylation. As with MPH, cocaine stimulated DARPP-32 Thr34 phosphorylation in slices from adult, but not from young mice. In contrast, a dopamine D1 agonist, SKF81297, regulated DARPP-32 phosphorylation comparably in slices from young and adult mice, as did methamphetamine, a dopamine releaser. The results suggest that dopamine synthesis and the dopamine transporter are functional at dopaminergic terminals in young mice. In contrast, the lack of effect of MPH in young mice is likely attributable to immature development of the machinery that regulates vesicular dopamine release.
Aging↗
Pharmacology of adenosine A2A receptors and therapeutic applications.
Adenosine A(2A) receptors were cloned about ten years ago and are known to be well conserved among mammals. Rather selective agonists and antagonists are available. In addition, two different knock-out mice have been prepared and extensively characterized. A(2A) receptors are highly enriched in the basal ganglia and on cells involved in inflammatory reactions. At these sites they are likely to play physiologically important roles. Efforts to develop new therapies based on A(2A) receptors have focused on these topics. However, A(2A) receptors are found on many other cell types and on them as well agonists can exert effect.
Animals↗
Acute and repeated treatment with L-DOPA increase c-jun expression in the 6-hydroxydopamine-lesioned forebrain of rats and common marmosets.
L-DOPA was acutely or repeatedly administered to rats and common marmosets (Callithrix jacchus) with unilateral 6-hydroxydopamine (6-OHDA) denervation of the dopamine inputs to the forebrain. Using in situ hybridization it was found that L-DOPA-treated animals exhibited a pronounced induction in the gene expression of both c-jun and c-fos in striatum and cerebral cortex restricted to the dopamine-depleted hemisphere. In contrast, acute treatment with cocaine induced c-fos mRNA, but not c-jun mRNA, in the striatum of normal animals. These data suggest that dopamine denervation leads to neurochemical adaptations which enables L-DOPA to induce a sustained gene expression of c-jun. Such aberrant gene regulation may underlie the development of L-DOPA-induced movement disorders which are commonly found in patients with Parkinson's disease.
Animals↗
Involvement of DARPP-32 phosphorylation in the stimulant action of caffeine.
Caffeine has been imbibed since ancient times in tea and coffee, and more recently in colas. Caffeine owes its psychostimulant action to a blockade of adenosine A(2A) receptors, but little is known about its intracellular mechanism of action. Here we show that the stimulatory effect of caffeine on motor activity in mice was greatly reduced following genetic deletion of DARPP-32 (dopamine- and cyclic AMP-regulated phosphoprotein of relative molecular mass 32,000). Results virtually identical to those seen with caffeine were obtained with the selective A(2A) antagonist SCH 58261. The depressant effect of the A(2A) receptor agonist, CGS 21680, on motor activity was also greatly attenuated in DARPP-32 knockout mice. In support of a role for DARPP-32 in the action of caffeine, we found that, in striata of intact mice, caffeine increased the state of phosphorylation of DARPP-32 at Thr 75. Caffeine increased Thr 75 phosphorylation through inhibition of PP-2A-catalysed dephosphorylation, rather than through stimulation of cyclin-dependent kinase 5 (Cdk5)-catalysed phosphorylation, of this residue. Together, these studies demonstrate the involvement of DARPP-32 and its phosphorylation/dephosphorylation in the stimulant action of caffeine.
Adenosine↗
DARPP-32 mediates serotonergic neurotransmission in the forebrain.
Serotonin is implicated in the regulation of complex sensory, motor, affective, and cognitive functions. However, the biochemical mechanisms whereby this neurotransmitter exerts its actions remain largely unknown. DARPP-32 (dopamine- and cAMP-regulated phosphoprotein of molecular weight 32,000) is a phosphoprotein that has primarily been characterized in relation to dopaminergic neurotransmission. Here we report that serotonin regulates DARPP-32 phosphorylation both in vitro and in vivo. Stimulation of 5-hydroxy-tryptamine (5-HT4 and 5-HT6 receptors causes an increased phosphorylation state at Thr34-DARPP-32, the protein kinase A site, and a decreased phosphorylation state at Thr75-DARPP-32, the cyclin-dependent kinase 5 site. Furthermore, stimulation of 5-HT2 receptors increases the phosphorylation state of Ser137-DARPP-32, the casein kinase-1 site. Behavioral and gene transcriptional effects induced by compounds that selectively release serotonin were greatly reduced in DARPP-32 knockout mice. Our data indicate that DARPP-32 is essential not only for dopaminergic but also for serotonergic neurotransmission.
5-Hydroxytryptophan↗
Involvement of striatal and extrastriatal DARPP-32 in biochemical and behavioral effects of fluoxetine (Prozac).
Fluoxetine (Prozac) is the most widely prescribed medication for the treatment of depression. Nevertheless, little is known about the molecular basis of its clinical efficacy, apart from the fact that fluoxetine increases the synaptic availability of serotonin. Here we show that, in vivo, fluoxetine, given either acutely or chronically, regulates the phosphorylation state of dopamine- and cAMP-regulated phosphoprotein of M(r) 32,000 (DARPP-32) at multiple sites in prefrontal cortex, hippocampus, and striatum. Acute administration of fluoxetine increases phosphorylation of DARPP-32 at the protein kinase A site, Thr-34, and at the casein kinase-1 site, Ser-137, and decreases phosphorylation at the cyclin-dependent kinase 5 site, Thr-75. Each of these changes contributes, through distinct signaling pathways, to increased inhibition of protein phosphatase-1, a major serine/threonine protein phosphatase in the brain. Fluoxetine also increases phosphorylation of the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunit GluR1 at Ser-831 and Ser-845. Both the fluoxetine-mediated increase in AMPA receptor phosphorylation at Ser-845-GluR1 and the beneficial responsiveness to fluoxetine in an animal test of antidepressant efficacy were strongly reduced in DARPP-32 knockout mice, indicating a critical role for this phosphoprotein in the antidepressant actions of fluoxetine. Mice chronically treated with fluoxetine had increased levels of DARPP-32 mRNA and protein and a decreased ability to increase phospho-Ser-137-DARPP-32 and phospho-Ser-831-GluR1. These chronic changes may be relevant to the delayed onset of therapeutic efficacy of fluoxetine.
Animals↗
Dopamine D1/5 receptor stimulation induces c-fos expression in the subthalamic nucleus: possible involvement of local D5 receptors.
The activity of neurons in the subthalamic nucleus controls various aspects of movement. The present study examined the action of dopamine receptor agonists on c-fos gene expression in the subthalamic nucleus in normal rats. We found that systemic administration of the dopamine D1/5 receptor agonist, SKF 82958 (1 mg/kg), induces c-fos expression in the subthalamic nucleus. In contrast, systemic administration of the dopamine D2/3 receptor agonist, quinelorane (2 mg/kg) had no effect. When combined, SKF 82958 and quinelorane induced c-fos expression in subthalamic neurons that was similar to that found following administration of SKF 82958 alone. We also examined c-fos expression in the substantia nigra pars reticulata, the major projection area for subthalamic neurons, and found that SKF 82958, but not quinelorane, caused an induction of c-fos expression in this area. In order to clarify the mechanisms underlying the SKF 82958-mediated induction of c-fos expression in the subthalamic nucleus and substantia nigra pars reticulata, in situ hybridization for the dopamine D1, D2, D3 and D5 receptor mRNAs was performed. The only significant observation was that D5 receptor mRNA is expressed in subthalamic neurons. The present data show that dopamine, via D1/D5 receptors, upregulates c-fos expression in subthalamic neurons, and that the high expression of D5 receptors in this area might be involved. Taken together, these data suggest that dopamine D1/5 receptors are more important for the action of dopamine in the so-called indirect pathway of the basal ganglia circuitry than what is recognized in current models of basal ganglia organization.
Animals↗
Peptidomics-based discovery of novel neuropeptides.
Modern proteomic methodologies have significantly improved the possibilities of large-scale identification of proteins. However, these methodologies are limited by their inability to reliably detect endogenously expressed peptides. We describe a novel approach of combining sample preparation, comprising focused microwave irradiation and mass spectrometric peptide profiling that has enabled us to simultaneously detect more than 550 endogenous neuropeptides in 1 mg of hypothalamic extracts. Automatic switching tandem mass spectrometry and amino acid sequence determination of the peptides showed that they consist of both novel and previously described neuropeptides. The methodology includes virtual visualization of the peptides as two- and three-dimensional image maps. In addition, several novel and known post-translational modifications of the neuropeptides were identified. The peptidomic approach proved to be a powerful method for investigating endogenous peptides and their post-translational modifications in complex tissues such as the brain. It is anticipated that this approach will complement proteomic methods in the future.
Amino Acid Sequence↗
Increased levels of ubiquitin in the 6-OHDA-lesioned striatum of rats.
Multiple genetic deficits have linked impaired ubiquitin-conjugation pathways to various forms of familiar Parkinson's disease. We therefore examined the possible role of 6-hydroxydopamine, a dopaminergic neurotoxin used in Parkinson's disease experimental models, in causing protein degradation and its association with the ubiquitin proteasome system. Using unilaterally 6-hydroxydopamine-denervated rats and mass spectrometry profiling directly on brain tissue sections, we here report for the first time an increased level of unconjugated ubiquitin specifically in the dorsal striatum of the dopamine depleted hemisphere. No similar changes were found in the intact hemisphere or in the ventral striatum of the dopamine depleted hemisphere. The lesioning of the dopamine innervation to the striatum was confirmed by a strongly reduced dopamine transporter binding in the striatum, indicating an abundant loss of dopamine neurons. These results suggest that denervation of dopamine neurons per se is implicated in the regulation of ubiquitin pathways, at least in a classical animal model of Parkinson's disease. This study adds additional information regarding the involvement of the ubiquitin-proteasome system in Parkinson's disease.
Animals↗
Molecular profiling of experimental Parkinson's disease: direct analysis of peptides and proteins on brain tissue sections by MALDI mass spectrometry.
Direct molecular profiling of biological samples using matrix-assisted laser desorption ionization mass spectrometry is a powerful tool for identifying phenotypic markers. In this report, protein profiling was used for the first time to generate peptide and protein profiles of brain tissue sections obtained from experimental Parkinson's disease (unilaterally 6-hydroxydopamine treated rats). The mass spectrometer was used to map the peptide and protein expression directly on 12 microm tissue sections in mass-to-charge (m/z) values, providing the capability of mapping specific molecules of the original sample, that is, localization, intensity and m/z ratio. Several protein expression profile differences were found in the dopamine depleted side of the brain when compared to the corresponding intact side, for example, calmodulin, cytochrome c, and cytochrome c oxidase. An increased ratio of post-translational modifications such as acetylations were found in the striatum of proteins in the dopamine depleted side of the brain. These modifications were decreased after subchronic administration of L-Dopa. The present study shows that unique protein profiles can be obtained in specific brain regions (and subregions) directly on brain tissue sections and allows for the study of complex biochemical processes such as those occurring in experimental Parkinson's disease.
Adrenergic Agents↗