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Rodrigo A Cunha

Publications and source records attributed to Rodrigo A Cunha.

At least 19 recordsLinked to original sources
PubMed2007

Increase of cannabinoid CB1 receptor density in the hippocampus of streptozotocin-induced diabetic rats.

In the hippocampus, impaired neurophysiology, compromised neurogenesis, and eventually apoptosis accompany cognitive deficits in insulinopenic (type-1) diabetes (T1D). The underlying pathological mechanisms remain to be defined. The hippocampus has a high density of the cannabinoid type 1 receptor (CB(1)R), which has been shown to control several brain functions affected by diabetes, such as synaptic plasticity, glucose utilisation, memory consolidation and cognition, and maturation and survival of hippocampal neurons. However, the role of this receptor has not been investigated yet in diabetic encephalopathy. We report now that in the streptozotocin animal model of T1D, the hippocampal densities of CB(1)R protein and of specific CB(1)R binding sites are significantly increased both in the nerve terminals and in total membranes (changes between 13% and 38%), whereas CB(1)R mRNA expression is decreased by 25%, suggesting that CB(1)Rs might play a role in diabetic encephalopathy.

Animals↗
PubMed2006

Caffeine and adenosine A(2a) receptor antagonists prevent beta-amyloid (25-35)-induced cognitive deficits in mice.

Consumption of caffeine, an adenosine receptor antagonist, was found to be inversely associated with the incidence of Alzheimer's disease. Moreover, caffeine protects cultured neurons against beta-amyloid-induced toxicity, an effect mimicked by adenosine A(2A) but not A(1) receptor antagonists. We now tested if caffeine administration would prevent beta-amyloid-induced cognitive impairment in mice and if this was mimicked by A(2A) receptor blockade. One week after icv administration of the 25-35 fragment of beta-amyloid (Abeta, 3 nmol), mice displayed impaired performance in both inhibitory avoidance and spontaneous alternation tests. Prolonged treatment with caffeine (1 mg/ml) had no effect alone but prevented the Abeta-induced cognitive impairment in both tasks when associated with acute caffeine (30 mg/kg) 30 min treatment before Abeta administration. The same protective effect was observed after subchronic (4 days) treatment with daily injections of either caffeine (30 mg/kg) or the selective adenosine A(2A) receptor antagonist SCH58261 (0.5 mg/kg). This provides the first direct in vivo evidence that caffeine and A(2A) receptor antagonists afford a protection against Abeta-induced amnesia, which prompts their interest for managing Alzheimer's disease.

Adenosine A2 Receptor Antagonists↗
PubMed2006

CB1 receptor antagonism increases hippocampal acetylcholine release: site and mechanism of action.

Evidence indicates that blockade of cannabinoid receptors increases acetylcholine (ACh) release in brain cortical regions. Although it is assumed that this type of effect is mediated through CB1 receptor (CB1R) antagonism, several in vitro functional studies recently have suggested non-CB1R involvement. In addition, neither the precise neuroanatomical site nor the exact mechanisms underlying this effect are known. We thoroughly examined these issues using a combination of systemic and local administration of CB1R antagonists, different methods of in vivo microdialysis, CB1R knockout (KO) mice, tissue measurements of ACh, and immunochemistry. First, we showed that systemic injections of the CB1R antagonists N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboximide hydrochloride (SR-141716A) and N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2, 4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (AM251) dose-dependently increased hippocampal ACh efflux. Likewise, local hippocampal, but not septal, infusions of SR141716A or AM251 increased hippocampal ACh release. It is noteworthy that the stimulatory effects of systemically administered CB1R antagonists on hippocampal ACh release were completely abolished in CB1R KO mice. CB1R KO mice had similar basal but higher stress-enhanced hippocampal ACh levels compared with wild-type controls. It is interesting that dopamine D1 receptor antagonism counteracted the stimulatory effect of CB1R blockade on hippocampal ACh levels. Finally, immunohistochemical methods revealed that a high proportion of CB1R-positive nerve terminals were found in hippocampus and confirmed the colocalization of CB1 receptors with cholinergic and dopaminergic nerve terminals. In conclusion, hippocampal ACh release may specifically be controlled through CB1Rs located on both cholinergic and dopaminergic neuronal projections, and CB1R antagonism increases hippocampal ACh release, probably through both a direct disinhibition of ACh release and an indirect increase in dopaminergic neurotransmission at the D1 receptors.

Acetylcholine↗
PubMed2006

Lack of evidence for functional TRPV1 vanilloid receptors in rat hippocampal nerve terminals.

Although TRPV(1) vanilloid receptors (TRPV(1)Rs) have been assumed to be present in the brain, their role is not well-defined. Here, we tested the widely used TRPV(1)R agonists (E)-capsaicin (0.1-100 microM) and resiniferatoxin (RTX, 0.1 microM) on resting and K(+)-evoked Ca(2+) entry and radiolabelled GABA release in rat hippocampal nerve terminals. (E)-capsaicin and RTX failed to evoke Ca(2+) transients or to trigger [(3)H]GABA outflow. Both (E)-capsaicin (EC(50), 40.4 microM) and its enantiomer (Z)-capsaicin (EC(50), 22.9 microM), which is inactive at the TRPV(1)R, inhibited the K(+)-evoked Ca(2+) entry, and to similar extent, the Ca(2+)-dependent K(+)-evoked [(3)H]GABA release. The TRPV(1)R enhancer/partial agonist 2-aminoethoxydiphenyl borate (1-300 microM) induced rapid Ca(2+) entry. None of the above-mentioned findings proved to be sensitive to the TRPV(1)R antagonists iodoresiniferatoxin (I-RTX; 3 microM) and SB366791 (3 microM). The CB(1) cannabinoid receptor antagonist AM251 (EC(50), 1.1 microM) and I-RTX (EC(50), 4.6 microM) also diminished the K(+)-evoked Ca(2+) entry per se. We observed competitive antagonism between I-RTX and AM251, indicating that the two molecules may act at the same site. In conclusion, there is a need to examine the discrepancy between ex vivo and in vitro data to understand the neurochemical and physiological functions of brain TRPV(1)Rs.

Animals↗
PubMed2006

Presynaptic control of striatal glutamatergic neurotransmission by adenosine A1-A2A receptor heteromers.

The functional role of heteromers of G-protein-coupled receptors is a matter of debate. In the present study, we demonstrate that heteromerization of adenosine A1 receptors (A1Rs) and A2A receptors (A2ARs) allows adenosine to exert a fine-tuning modulation of glutamatergic neurotransmission. By means of coimmunoprecipitation, bioluminescence and time-resolved fluorescence resonance energy transfer techniques, we showed the existence of A1R-A2AR heteromers in the cell surface of cotransfected cells. Immunogold detection and coimmunoprecipitation experiments indicated that A1R and A2AR are colocalized in the same striatal glutamatergic nerve terminals. Radioligand-binding experiments in cotransfected cells and rat striatum showed that a main biochemical characteristic of the A1R-A2AR heteromer is the ability of A2AR activation to reduce the affinity of the A1R for agonists. This provides a switch mechanism by which low and high concentrations of adenosine inhibit and stimulate, respectively, glutamate release. Furthermore, it is also shown that A1R-A2AR heteromers constitute a unique target for caffeine and that chronic caffeine treatment leads to modifications in the function of the A1R-A2AR heteromer that could underlie the strong tolerance to the psychomotor effects of caffeine.

Animals↗
PubMed2006

Trkb receptors modulation of glutamate release is limited to a subset of nerve terminals in the adult rat hippocampus.

Brain-derived neurotrophic factor (BDNF) modulates glutamatergic excitatory transmission in hippocampal primary cultures by acting at a presynaptic locus. Although it has been suggested that BDNF also modulates adult hippocampus glutamatergic transmission, this remains a matter of controversy. To clarify a putative role for this neurotrophin in the modulation of glutamate release we applied exogenous BDNF to isolated adult rat hippocampal nerve terminals. BDNF, at 100 ng/ml, potentiated by 25% the K(+)-evoked release of [(3)H]glutamate from hippocampal synaptosomes. The small effect of BDNF on [(3)H]glutamate release correlated with a modest increase in phospholipase Cgamma (PLCgamma) phosphorylation, and with the lack of effect of BDNF on extracellular-signal regulated kinase (ERK) and Akt phosphorylation. Immunocytochemistry studies demonstrated that only about one-third of glutamatergic synaptosomes were positive for TrkB immunoreactivity. Furthermore, biotinylation and subsynaptic fractionation studies showed that only one-fourth of total full-length TrkB was present at the plasma membrane, evenly distributed between the presynaptic active zone and the postsynaptic density. These results indicate that BDNF modulates synaptic transmission presynaptically in a small subset of hippocampal glutamatergic synapses that contain TrkB and that express the receptor on the plasma membrane.

Animals↗
PubMed2006

Interaction between P2X and nicotinic acetylcholine receptors in glutamate nerve terminals of the rat hippocampus.

Nicotinic acetylcholine receptors (nAChRs [constituted by pentameric association of alpha2-10 and beta2-4 subunits]) and P2X receptors (P2XRs [activated by ATP and constituted by multimeric association of P2X1-7 subunits]) are both ionotropic receptors permeable to cations, which have in common the disparity between the wealth of data showing their presence in the brain and little evidence of their participation in mediating synaptic transmission. This has led to the proposal that both nAChRs and P2XRs might primarily modulate rather than directly mediate synaptic transmission, which is in accordance with the predominant presynaptic localization of both receptor subtypes (Role and Berg, 1996; Cunha and Ribeiro, 2000). Interestingly, both functional neurochemical (Allgaier et al., 1995; Salgado et al., 2000; Diáz-Hernández et al., 2002) and electrophysiological studies (Barajas-Lopez et al., 1998; Searl et al., 1998; Zhou and Calligan, 1998; Khakh et al., 2000) indicated a close interaction between nAChRs and P2XRs, which is paralleled by a co-release of ATPand ACh from central terminals (e.g., Richardson and Brown, 1987). Because glutamate release in the hippocampus is controlled by both nAChRs (e.g., McGehee et al., 1995) and P2XRs (Khakh et al., 2003; Rodrigues et al., 2005), we investigated if there was a functional interaction between these two presynaptic ionotropic receptors in the control of glutamate release in the rat hippocampus.

Animals↗
PubMed2005

Modification of adenosine A1 and A2A receptor density in the hippocampus of streptozotocin-induced diabetic rats.

Adenosine A(1) and A(2A) receptors are neuromodulatory systems that can control mnemonic behavior, which is modified by diabetes. Since the density of these adenosine receptors can change upon chronic noxious brain conditions, we now tested if the density of A(1) and A(2A) receptors was modified in the hippocampus of streptozotocin-induced diabetic rats. The binding density of the selective A(1) receptor antagonist, (3)H-DPCPX, was decreased by 36% in total hippocampal membranes 7 days after induction of diabetes and this down-regulation was maintained after 30 and 90 days, which was also confirmed by Western blot analysis of A(1) receptor immunoreactivity. In contrast, the binding density of the selective A(2A) receptor antagonist, (3)H-SCH 58261, was enhanced by 83% in total hippocampal membranes 7 days after induction of diabetes and this up-regulation persisted after 30 and 90 days. These results show that the balance between inhibitory A(1) and facilitatory A(2A) adenosine receptors is modified in the hippocampus of streptozotocin-induced diabetic rats. Thus, the most abundant A(1) receptors are down-regulated and there is an up-regulation of A(2A) receptors, suggesting a gain of function of hippocampal A(2A) receptors compared to A(1) receptors in diabetes, as has been observed in other chronic noxious brain conditions where A(2A) receptor blockade affords robust neuroprotection.

Animals↗
PubMed2005

Dual presynaptic control by ATP of glutamate release via facilitatory P2X1, P2X2/3, and P2X3 and inhibitory P2Y1, P2Y2, and/or P2Y4 receptors in the rat hippocampus.

ATP is released in a vesicular manner from nerve terminals mainly at higher stimulation frequencies. There is a robust expression of ATP (P2) receptors in the brain, but their role is primarily unknown. We report that ATP analogs biphasically modulate the evoked release of glutamate from purified nerve terminals of the rat hippocampus, the facilitation being mediated by P2X1, P2X2/3, and P2X3 [antagonized by 8-(benzamido)naphthalene-1,3,5-trisulfonate and 2',3'-O-(2,4,6-trinitrophenyl)-ATP] and the inhibition by P2Y1, P2Y2, and/or P2Y4 [antagonized by reactive blue 2 and 2'deoxy-N6-methyladenosine-3',5'-bisphosphate and mimicked by P1-(urinine 5'-),P4-(inosine 5'-) tetraphosphate and 2-methylthio-ADP] receptors. The combination of single-cell PCR analysis of rat hippocampal pyramidal neurons, Western blot analysis of purified presynaptic active zone fraction, and immunocytochemical analysis of hippocampal glutamatergic terminals revealed that the P2 receptors expressed in glutamatergic neurons, located in the active zone and in glutamatergic terminals, were precisely P2X1, P2X2, and P2X3 subunits and P2Y1, P2Y2, and P2Y4 receptors. This provides coincident functional and molecular evidence that P2 receptors are present and act presynaptically as a modulatory system controlling hippocampal glutamate release.

Adenosine Diphosphate↗
PubMed2005

Involvement of cannabinoid receptors in the regulation of neurotransmitter release in the rodent striatum: a combined immunochemical and pharmacological analysis.

Despite the profound effect of cannabinoids on motor function, and their therapeutic potential in Parkinson's and Huntington's diseases, the cellular and subcellular distributions of striatal CB1 receptors are not well defined. Here, we show that CB1 receptors are primarily located on GABAergic (vesicular GABA transporter-positive) and glutamatergic [vesicular glutamate transporter-1 (VGLUT-1)- and VGLUT-2-positive] striatal nerve terminals and are present in the presynaptic active zone, in the postsynaptic density, as well as in the extrasynaptic membrane. Both the nonselective agonist WIN552122 [(R)-(+)-[2,3-dihydro-5-methyl-3[(4-morpholinyl)methyl] pyrrolo[1,2,3-de]-1,4-benzoxazinyl]-(1-naphthalenyl)methanone mesylate salt] (EC50, 32 nM) and the CB1-selective agonist ACEA [N-(2-chloroethyl)-5Z,8Z,11Z,14Z-eicosatetraenamide] inhibited [3H]GABA release from rat striatal slices. The effect of these agonists was prevented by the CB1-selective antagonists SR141716A [N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide] (1 microM) and AM251 [1-(2,4-dichlorophenyl)-5-(4-iodophenyl)-4-methyl-N-1-piperidinyl-1H-pyrazole-3-carboxamide trifluoroacetate salt] (1 microM), indicating that cannabinoids inhibit the release of GABA via activation of presynaptic CB1 receptors. Cannabinoids modulated glutamate release via both CB1 and non-CB1 mechanisms. Cannabinoid agonists and antagonists inhibited 25 mM K+-evoked [3H]glutamate release and sodium-dependent [3H]glutamate uptake. Partial involvement of CB1 receptors is suggested because low concentrations of SR141716A partly and AM251 fully prevented the effect of WIN552122 and CP55940 [5-(1,1-dimethylheptyl)-2-[5-hydroxy-2-(3-hydroxypropyl)cyclohexyl]phenol]. However, the effect of CB1 agonists and antagonists persisted in CB1 knock-out mice, indicating the involvement of non-CB1,CB1-like receptors. In contrast, cannabinoids did not modulate [3H]dopamine release or [3H]dopamine and [3H]GABA uptake. Our results indicate distinct modulation of striatal GABAergic and glutamatergic transmission by cannabinoids and will facilitate the understanding of the role and importance of the cannabinoid system in normal and pathological motor function.

6-Cyano-7-nitroquinoxaline-2,3-dione↗
PubMed2005

Different roles of adenosine A1, A2A and A3 receptors in controlling kainate-induced toxicity in cortical cultured neurons.

Adenosine is a neuromodulator that can control brain damage through activation of A(1), A(2A) and A(3) receptors, which are located in both neurons and other brain cells. We took advantage of cultured neurons to investigate the role of neuronal adenosine receptors in the control of neurotoxicity caused by kainate and cyclothiazide. Both A(1), A(2A) and A(3) receptors were immunocytochemically identified in cortical neurons. Activation of A(1) receptors with 100 nM CPA did not modify the extent of neuronal death whereas the A(1) receptor antagonist, DPCPX (50 nM), attenuated neurotoxicity by 28 +/- 5%, and effect similar to that resulting from the removal of endogenous adenosine with 2U/ml of adenosine deaminase (27 +/- 3% attenuation of neurotoxicity). In the presence of adenosine deaminase, DPCPX had no further effect and CPA now exacerbated neurotoxicity by 42 +/- 4%. Activation of A(2A) receptor with 30 nM CGS21680 attenuated neurotoxicity by 40 +/- 8%, an effect prevented by the A(2A) receptor antagonists, SCH58261 (50 nM) or ZM241385 (50 nM), which by themselves were devoid of effect. Finally, neither A(3) receptor activation with Cl-IB-MECA (100-500 nM) nor blockade with MRS1191 (5 microM) modified neurotoxicity. These results show that A(1) receptor activation enhances and A(2A) receptor activation attenuates neurotoxicity in cultured cortical neurons, indicating that these two neuronal adenosine receptors directly control neurodegeneration. Interestingly, the control by adenosine of neurotoxicity in cultured neurons is similar to that observed in vivo in newborn animals and is the opposite of what is observed in adult brain preparations where A(1) receptor activation and A(2A) receptor blockade are neuroprotective.

Adenosine↗
PubMed2005

Co-localization and functional interaction between adenosine A(2A) and metabotropic group 5 receptors in glutamatergic nerve terminals of the rat striatum.

The anti-Parkinsonian effect of glutamate metabotropic group 5 (mGluR5) and adenosine A(2A) receptor antagonists is believed to result from their ability to postsynaptically control the responsiveness of the indirect pathway that is hyperfunctioning in Parkinson's disease. mGluR5 and A(2A) antagonists are also neuroprotective in brain injury models involving glutamate excitotoxicity. Thus, we hypothesized that the anti-Parkinsonian and neuroprotective effects of A(2A) and mGluR5 receptors might be related to their control of striatal glutamate release that actually triggers the indirect pathway. The A(2A) agonist, CGS21680 (1-30 nM) facilitated glutamate release from striatal nerve terminals up to 57%, an effect prevented by the A(2A) antagonist, SCH58261 (50 nM). The mGluR5 agonist, CHPG (300-600 mum) also facilitated glutamate release up to 29%, an effect prevented by the mGluR5 antagonist, MPEP (10 microm). Both mGluR5 and A(2A) receptors were located in the active zone and 57 +/- 6% of striatal glutamatergic nerve terminals possessed both A(2A) and mGluR5 receptors, suggesting a presynaptic functional interaction. Indeed, submaximal concentrations of CGS21680 (1 nM) and CHPG (100 microm) synergistically facilitated glutamate release and the facilitation of glutamate release by 10 nM CGS21680 was prevented by 10 microm MPEP, whereas facilitation by 300 microm CHPG was prevented by 10 nM SCH58261. These results provide the first direct evidence that A(2A) and mGluR5 receptors are co-located in more than half of the striatal glutamatergic terminals where they facilitate glutamate release in a synergistic manner. This emphasizes the role of the modulation of glutamate release as a likely mechanism of action of these receptors both in striatal neuroprotection and in Parkinson's disease.

Adenosine↗
PubMed2005

Adenosine A2A receptors control the extracellular levels of adenosine through modulation of nucleoside transporters activity in the rat hippocampus.

Adenosine, a neuromodulator of the CNS, activates inhibitory-A1 receptors and facilitatory-A2A receptors; its synaptic levels are controlled by the activity of bi-directional equilibrative nucleoside transporters. To study the relationship between the extracellular formation/inactivation of adenosine and the activation of adenosine receptors, we investigated how A1 and A2A receptor activation modifies adenosine transport in hippocampal synaptosomes. The A2A receptor agonist, CGS 21680 (30 nm), facilitated adenosine uptake through a PKC-dependent mechanism, but A1 receptor activation had no effect. CGS 21680 (30 nm) also increased depolarization-induced release of adenosine. Both effects were prevented by A2A receptor blockade. A2A receptor-mediated enhancement of adenosine transport system is important for formatting adenosine neuromodulation according to the stimulation frequency, as: (1) A1 receptor antagonist, DPCPX (250 nm), facilitated the evoked release of [(3)H]acetylcholine under low-frequency stimulation (2 Hz) from CA3 hippocampal slices, but had no effect under high-frequency stimulation (50 Hz); (2) either nucleoside transporter or A2A receptor blockade revealed the facilitatory effect of DPCPX (250 nm) on [3H]acetylcholine evoked-release triggered by high-frequency stimulation. These results indicate that A2A receptor activation facilitates the activity of nucleoside transporters, which have a preponderant role in modulating the extracellular adenosine levels available to activate A1 receptors.

Adenosine↗
PubMed2005

Long-term effect of convulsive behavior on the density of adenosine A1 and A 2A receptors in the rat cerebral cortex.

PURPOSE: Adenosine is a neuromodulator that has been proposed to act as an anticonvulsant mainly via inhibitory A1 receptors, but recent data show that genetic deletion of facilitatory A 2A receptors might also attenuate convulsions. Since both A1 and A 2A receptors are prone to down- and upregulation in different stressful situations, we investigated if convulsive behavior leads to a long-term change in A1 and A 2A receptor density in the rat cerebral cortex. METHODS: Stage 4-5 convulsions (Racine's scale) were induced in adult Wistar rats either through amygdala stimulation (kindling) or by intraperitoneal injection of kainate (10 mg/ml). Rats were killed after 4 weeks to evaluate adenosine A1 and A 2A receptor density in the cerebral cortex using both Western blot and membrane binding assays. RESULTS: The binding density of the A1 antagonist, 3H-DPCPX, decreased by 40. +/- 4.4% and by 20.7 +/- 0.5% after kindling or kainate injection. Likewise, A1 receptor immunoreactivity in cortical membranes from kindled or kainate-injected rats decreased by 19.1 +/- 3.3% and 12.7 +/- 5.7%, respectively. In contrast, the binding density of the A 2A receptor antagonist 3H-SCH 58261 increased by 293 +/- 34% and by 159 +/- 32% in cortical membranes from kindled or kainate-injected rats, and A 2A receptor immunoreactivity also increased by 151 +/- 12% and 79.6 +/- 7.0%. CONCLUSIONS: This indicates that after convulsive behavior there is a long-term decrease of A1 receptors accompanied by an increased density of A 2A receptors, suggesting that A 2A antagonists rather than A1 agonists may be more promising anticonvulsive drugs.

Amygdala↗
PubMed2004

Binding of adenosine receptor ligands to brain of adenosine receptor knock-out mice: evidence that CGS 21680 binds to A1 receptors in hippocampus.

The adenosine receptor agonist 2-[ p-(2-carboxyethyl)phenylethylamino]-5'- N-ethylcarboxamidoadenosine (CGS 21680) is generally considered to be a selective adenosine A(2A) receptor ligand. However, the compound has previously been shown to exhibit binding characteristics that are not compatible with adenosine A(2A) receptor binding, at least in brain regions other than the striatum. We have examined binding of [(3)H]CGS 21680 and of antagonist radioligands with high selectivity for adenosine A(1) or A(2A) receptors to hippocampus and striatum of mice lacking either adenosine A(1) (A1R((-/-))) or A(2A) (A2AR((-/-))) receptors. Both receptor autoradiography and membrane binding techniques were used for this purpose and gave similar results. There were no significant changes in the binding of the A(1) receptor antagonist [(3)H]DPCPX in mice lacking A(2A) receptors, or in the binding of the A(2A) receptor antagonists [(3)H]SCH 58261 and [(3)H]ZM 241385 in mice lacking A(1) receptors. Furthermore, [(3)H]CGS 21680 binding in striatum was abolished in the A2AR((-/-)), and essentially unaffected in striatum from mice lacking A(1) receptors. In hippocampus, however, binding of [(3)H]CGS 21680 remained in the A2AR((-/-)), whereas binding was virtually abolished in the A1R((-/-)). There were no adaptive alterations in A(2A) receptor expression in this region in A1R((-/-)) mice. Thus, most of the [(3)H]CGS 21680 binding in hippocampus is dependent on the presence of adenosine A(1) receptors, but not on A(2A) receptors, indicating a novel binding site or novel binding mode.

Adenosine↗
PubMed2004

Binding of the prototypical adenosine A(2A) receptor agonist CGS 21680 to the cerebral cortex of adenosine A(1) and A(2A) receptor knockout mice.

1. 2-p-(2-carboxyethylphenethylamino-5'-ethylcarboxamidoadenosine) (CGS 21680) is considered the reference compound to study adenosine A(2A) receptors. However, CGS 21680 binding in the cerebral cortex, where adenosine A(1) receptors are predominant, displays a mixed A(2A)/A(1) receptor pharmacology. We now use adenosine A(1) and A(2A) receptor knockout mice to investigate the characteristics of cortical [(3)H]CGS 21680 binding. 2. [(3)H]CGS 21680 binding to the cerebral cortex was strongly reduced in adenosine A(1) receptor knockout mice, but only slightly reduced in A(2A) receptor knockout mice compared with the corresponding wild-type littermates. 3. Another selective A(2A) receptor ligand, [(3)H]-5-amino-7-(2-phenylethyl)-2-(2-furyl)-pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine ([(3)H]SCH 58261), displayed a saturable binding to mouse cortical membranes, albeit with a binding density 20 times lower than that of striatal membranes, and this [(3)H]SCH58261 binding was abolished in both striatal and cortical membranes of A(2A) receptor knockout mice and unchanged in A(1) receptor knockout mice. 4. The presence of A(2A) receptors in cortical neurons was further confirmed by Western blot in mouse cortical nerve terminal membranes. 5. It is concluded that, although A(2A) receptors are present in the cerebral cortex, the purportedly selective A(2A) receptor agonist [(3)H]CGS 21680 binds in the cerebral cortex to an entity that requires the presence of adenosine A(1) receptors. Thus, CGS 21680 should be used with care in all preparations where adenosine A(1) receptors out-number A(2A) receptors.

Adenosine↗
PubMed2004

Effect of free radicals on adenosine A(2A) and dopamine D2 receptors in the striatum of young adult and aged rats.

Parkinson's disease (PD) is a prevalent age-related motor dysfunction resulting from the hyperactivity of the indirect striatal pathway, which is controlled in an antagonistic manner by inhibitory dopamine D2 and facilitatory adenosine A(2A) receptors. Thus, dopamine precursors like l-DOPA are the standard therapy and A(2A) antagonists are now tested as anti-parkinsonians. Increased free radicals levels occur on aging and are proposed to be a contributing factor for PD. We now tested if free radicals affected A(2A) and D2 receptors in striatal membranes of young adult (2 months) and old (24 months) rats. The A(2A) receptor antagonist [3H]SCH 58261 bound to striatal membranes with a KD of 0.9 nM and a Bmax of 953 fmol/mg protein in young rats and with a KD of 0.8 nM and a Bmax of 725 fmol/mg protein in aged rats (24% decrease). The D2 receptor antagonist [3H]raclopride bound to striatal membranes with a KD of 4.0 nM and a Bmax of 598 fmol/mg protein in young rats and with a KD of 4.3 nM and a Bmax of 368 fmol/mg protein in aged rats (38% decrease). Exposure of striatal membranes to a free radical generation system (2 mM FeSO4 and 4 mM ascorbate) caused a similar decrease of [3H]SCH 58261 (35%) and [3H]raclopride (37%) binding in young adult rats but caused a greater decrease of [3H]SCH 58261 (49%) than of [3H]raclopride (20%) binding in aged rats. Thus, in aged rats, there is an unbalance of A(2A)/D2 receptor density favouring A(2A) receptors, which is restored on exposure to free radicals. This supports the hypothesis that the effectiveness of A(2A) receptor antagonists as anti-parkinsonians, demonstrated in young adult animals, may not be affected by a modified A(2A)/D2 receptor density in aged individuals suffering from exposure to increased free radical levels, as occurs in PD.

Aging↗