Postsynaptic dopamine/adenosine interaction: I. Adenosine analogues inhibit dopamine D2-mediated behaviour in short-term reserpinized mice.

Mice pretreated with reserpine 5 mg/kg (4 h prior to the start of motor activity recording) showed locomotor activation after the administration of the D-2 agonist bromocriptine (5 mg/kg). This bromocriptine-induced locomotor activity was dose dependently inhibited by the co-administration of a D-2 antagonist (sulpiride) and dose dependently potentiated by a D-1 agonist (CY 208-243). The potentiating effect of the D-1 agonist could be inhibited by either a D-1 or a D-2 antagonist (SCH 23390 1 mg/kg or sulpiride 100 mg/kg, respectively). The bromocriptine-induced locomotor activity was not altered by either blockade of D-1 dopaminergic receptors (SCH 23390 1 mg/kg) or by co-administration of a greater dose of reserpine (10 mg/kg) plus the dopamine synthesis inhibitor, alpha-methyl-p-tyrosine (200 mg/kg). The adenosine agonists, L-PIA (a preferentially A-1 adenosine agonist) and NECA (an A-1 and A-2 adenosine agonist with above 10-fold greater affinity for A-2 than L-PIA) inhibited in a dose-dependent manner the effect of bromocriptine, NECA being above ten times more potent than L-PIA. The findings show that bromocriptine stimulates postsynaptic D-2 receptors in dopamine-depleted mice and that this effect can be inhibited by adenosine stimulation. The existence of a postsynaptic D-2/A-2 interaction is suggested, the stimulation of A-2 receptors causing an inhibition of responses elicited by postsynaptic D-2 stimulation.     

Psychomotor stimulant effects of methylxanthines in squirrel monkeys: relation to adenosine antagonism

The behavioral effects of six methylxanthines were studied in squirrel monkeys responding under a fixed-interval (FI) schedule of stimulus-shock termination. Dose-response curves were determined for each drug by administering cumulative doses IV during timeout periods that preceded sequential components of the FI schedule. Low to intermediate doses of caffeine, theophylline, 8-phenyltheophylline (8-PT), 8-cyclopentyltheophylline (CPT), and 3-isobutyl-1-methylxanthine (IBMX) produced dose-related increases in response rate, whereas higher doses increased response rate less or decreased it. Enprofylline did not increase response rate at any dose. Pretreatment with a high dose of the adenosine agonist 5-N-ethylcarboxamide adenosine (NECA) suppressed responding throughout the experimental session. Caffeine, theophylline, 8-PT, CPT, and IBMX, but not enprofylline, antagonized the suppressant effects of NECA in a dose-related manner. The potencies of the methylxanthines for increasing response rate under the FI schedule were positively correlated with their potencies for antagonizing the suppressant effects of NECA, suggesting that the psychomotor stimulant effects of methylxanthines are linked to their antagonistic actions at adenosine recognition sites. CPT, which in vitro has much higher affinity (>10³-fold) than caffeine for adenosine A₁, but not for A₂, recognition sites, was only 3–6 times more potent than caffeine in increasing response rate or in antagonizing the effects of NECA. The psychomotor stimulant effects of methylxanthines therefore appear to be more closely associated with antagonism at adenosine A₂ than adenosine A₁ recognition sites.Animals used in this study were maintained in accordance with the guidelines of the Committee on Animals of the Harvard Medical School and of the Guide for Care and Use of Laboratory Animals of the Institute of Laboratory Animal Resources, National Research Council, Department of Health, Education and Welfare, Publication No. (NIH)85-23, revised 1985     

Effects on tracheal smooth muscle of adenosine and methylxanthines,and their interaction

The relaxant effects of xanthine and 15 different methylxanthines were studied in a guinea-pig isolated tracheal ring preparation. Substitution in the 3- and 1-positions of the xanthine molecule were found to be of greatest importance for improving tracheal relaxant potency. The unsubstituted xanthine and all 9-methylated xanthines were weakly active. Adenosine occasionally contracted (slightly) the tracheae but a concentration-dependent relaxation was always recorded. Endogenous adenosine did not seem to contribute to the tracheal tone. Xanthines methylated in the 1-position consistently antagonized the relaxant effect of adenosine and produced graded rightward shifts of the concentration-response line to adenosine (competitive antagonism). Xanthines with an unsubstituted 1-position or with comethylation in the 9-position were generally without antagonism. The results support our view that bronchodilating xanthine derivatives lacking universal adenosine antagonism can be produced.     

L-dopa esters as potential prodrugs: effect on brain concentration of dopamine metabolites in reserpinized mice

The intraperitoneal administration of L-dopa and a series of ester prodrugs of L-dopa to reserpinized mice produced elevations of striatal and tuberculum olfactorium homovanillic acid (HVA) and 3,4-dihydroxyphenylacetic acid (DOPAC) levels. Differences in the pattern of change produced by individual drugs, compared with L-dopa, were observed. Only the phenoxyethyl ester caused elevations of both striatal and tuberculum olfactorium HVA and DOPAC, greater than those measured following L-dopa administration. Overall the m-trifluoromethylbenzyl, phenylethyl, p-chlorophenylethyl and p-methoxyphenylethyl ester prodrugs produced greater elevations of striatal and tuberculum olfactorium HVA, but not DOPAC, compared with L-dopa. The administration of the 2-tetrahydropyranyl-methyl derivative only enhanced striatal HVA and striatal and tuberculum olfactorium DOPAC concentrations. Changes of HVA and DOPAC tissue concentrations following administration of the 2-hydroxypropyl, n-propyl, methyl, ethyl and 2-(1-methoxy)propyl ester prodrugs were comparable with those produced by the administration of L-dopa itself. The alterations in striatal and tuberculum olfactorium HVA and DOPAC levels observed did not correlate with the ability of these compounds to elicit locomotor activity in reserpinized mice.     

Elevation of cyclic AMP in C-1300 murine neuroblastoma by adenosine and related compounds and the antagonism of this response by methylxanthines.

Adenosine, in the presence of the phosphodiesterase inhibitor. Ro 20-1724, produces large increases in the intracellular content of cyclic adenosine 3',5'-monophosphate of murine neuroblastoma cells (clone N2a). This response is not blocked by dipyridamole, an antagonist of adenosine uptake, but is antagonized by certain methylxanthines. The structure activity relationships for agonistic and antagonistic activity were determined and the following observations made: (1) the N⁶ is necessary for the binding of adenosine to its receptor, but not for antagonist activity; (2) the substitution of a methyl group at the 1 position increases antagonistic activity of xanthines (from no activity to measurable activity) but greatly decreases the affinity of adenosine for its receptor; and (3) the sugar moiety is important for the binding of adenosine to its receptor, but not for antagonistic activity. The dissimilarities in the structure-activity relationships for agonistic activity and antagonist activity are interpreted to indicate that this antagonism is not competitive in nature.     

Adenosine-induced secretion in the canine trachea: modification by methylxanthines and adenosine derivatives.

Adenosine alone at 0.1 and 1.0 mg per tracheal segment stimulated mucus secretion by 52% and 88%, respectively, compared to baseline (P less than 0.0001). The site of the potent secretagogue effect of adenosine in canine trachea was consistent with A2 activation. A2 site activation and enhanced secretion were also induced by N-ethylcarboxamide adenosine and dipyridamole. N6-R-phenylisopropyl adenosine (PIA) and 2',5'-dideoxyadenosine (ddAdo) inhibited the adenosine-induced secretion (35% and 42%, respectively). However, when PIA or ddAdo were administered in conjunction with the potent phosphodiesterase inhibitor, methylisobutylxanthine (MIX), the effects of PIA were potentiated and the effects of ddAdo were reversed, yielding stimulation (A2) and antagonism (A1) of secretion, respectively. 8-Phenyltheophylline by aerosol was a very potent antagonist of the secretagogue effect of adenosine (70% inhibition; P less than 0.00001).     

The anticonvulsant action of methazolamide in mice: antagonism by various inhibitors of dopamine beta-hydroxylase.

The dopamine beta-hydroxylase inhibitors, FLA-63 and picolinic acid, antagonized the anticonvulsant action of methazolamine in mice; disulfiram and pyrimidinethiol were inactive. FLA-63 and picolinic acid, but not disulfiram or pyrimidinethiol prevented pheniprazine restoration of the anticonvulsant action of methazolamide in reserpinized mice. The present findings clearly demonstrate that differences exist among inhibitors of dopamine beta-hydroxylase regarding their ability to antagonize the anticonvulsant action of methazolamide under various test conditions.     

Observation of the grooming behaviour of reserpinized white mice as a method for investigating reserpine antagonism and synergism

The grooming movements of reserpinized white mice, dusted with pulverized charcoal, were studied under the influence of psychotropic drugs. With the help of this test it is possible to show differences in the antireserpine properties of different psychotropic substances. Only d-amphetamine, desipramine and Leo 640 (Chopramine, a new potential antidepressive) showed positive results in this test and stimulated the grooming movements again. In order to find the position of this new test in the test-battery the results were compared with those obtained in the reserpine-temperature and the ptosis tests for mice.     

Effects of combinations of methylxanthines and adenosine analogs on locomotor activity in control and chronic caffeine-treated mice

Chronic caffeine ingestion (CCI) by male NIH Swiss strain mice results in a prolonged reduction in locomotor activity and alterations in response to caffeine, other xanthines, and adenosine analogs. Caffeine, the A₁ selective 8-cyclopentyltheophylline (CPT), and the A₂-selective 3,7-dimethyl-1-propargylxanthine (DMPX) remain stimulatory and the bell-shaped locomotor dose-response curves are left-shifted after CCI. The depressant effects of methylxanthines that are potent phosphodiesterase inhibitors remain after CCI. After CCI, mice became more sensitive to depressant effects of A₁, mixed A₁/A₂, and A₂ agonists. In the presence of caffeine the A₁-selective agonist N⁶-cyclohexyladenosine (CHA), the mixed A₁/A₂ agonist 5′-N-ethylcarboxamidoadenosine and the A₂-selective agonist 2-[(2-aminoethylamino)-carbonylethylphenylethylamino]-5′-N-ethylcarboxamidoadenosine (APEC) all have dose-response curves, appearing to consist of initial depressant effects, then stimulatory effects, and finally pronounced depressant effects. The phasic character is reduced or absent after CCI. Synergistic depressant effects of combinations of CHA and APEC also appear reduced after CCI. © 1993 wiley-Liss, Inc.     

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.     

Effects of dopamine receptor agonists on passive avoidance learning in mice: interaction of dopamine D1 and D2 receptors.

The present study examined the effects of dopamine D1 and D2 receptor agonists on the acquisition stage of passive avoidance learning and on locomotor activity in mice. The D2 agonist, RU 24213 (1-10 mg/kg s.c.), and the non-selective agonist, apomorphine (0.3-3 mg/kg s.c.), but not the D1 agonist, SKF 38393 (1-10 mg/kg s.c.), impaired learning and activated locomotion. RU 24213 (1 mg/kg s.c.) was more effective in impairing learning than in activating locomotion. The concurrent administration of SKF 38393 (10 mg/kg i.p.) and RU 24213 (1 and 3 mg/kg s.c.) produced a synergistic effect in both behavioral situations. The D1 antagonist, SCH 23390 (0.025 mg/kg i.p.), slightly inhibited the effects of apomorphine and of the combination of SKF 38393 and RU 24213 on learning but not on locomotion. The D2 antagonist, (-)-sulpiride (40 mg/kg i.p.), completely blocked these effects in both situations. These results suggest that dopamine receptor agonists impair passive avoidance learning through the D2 receptor, and that D1 and D2 receptors act synergistically in this impairment, as they do in their effects on locomotion. The involvement of D1 and D2 receptors is qualitatively similar in each of these behaviors, although some small differences may exist.     

Comparison of the behavioral effects of adenosine agonists and dopamine antagonists in mice

The adenosine agonists 5-N-ethylcarboxamidea-denosine (NECA), 2-chloroadenosine (2-CLA), N⁶-cyclohexyladenosine (CHA), N⁶-cyclopentyladenosine (CPA), 2-(phenylamino)adenosine (CV-1808) and R and S isomers of N⁶-phenylisopropyladenosine (R-PIA and S-PIA) decreased spontaneous locomotor activity in mice and, except for CPA, did so at doses that did not impair motor coordination, a profile shared by dopamine antagonists. CV-1808, the only agent with higher affinity for A₂ as compared with A₁ adenosine receptors, displayed the largest separation between locomotor inhibitory and ataxic potency. Like dopamine antagonists, NECA and CV-1808 also decreased hyperactivity caused by d-amphetamine at doses that did not cause ataxia whereas A₁-selective adenosine agonists reduced amphetamine's effects only at ataxic doses. Unlike dopamine antagonists, adenosine agonists inhibited apomorphine-induced cage climbing only at doses that caused ataxia. Involvement of central adenosine receptors in these effects was suggested by the significant correlation obtained between potency for locomotor inhibition after IP and ICV administration. Affinity for A₁ but not A₂ adenosine receptors was significantly correlated with potency for inducing ataxia. These results suggest that the behavioral profile of adenosine agonists in mice is related to their affinity for A₁ and A₂ adenosine receptors and indicate that adenosine agonists produce certain behavioral effects that are similar to those seen with dopamine antagonists.     

Steric aspects of agonism and antagonism at beta-adrenoceptors: experiments with the enantiomers of clenbuterol

The enantiomers of clenbuterol, a beta 2-selective adrenoceptor agonist with partial agonistic activity, were examined with respect to their ability to react in vitro on adrenoceptors in the trachea (mostly beta 2), the soleus muscle (beta 2) and in the papillary muscle of the left ventricle (beta 1) from the guinea-pig. (-)-Clenbuterol relaxed the carbachol contracted trachea and depressed the subtetanic contractions of the soleus muscle in a concentration-dependent manner. (+)-Clenbuterol was at least 1,000 times less potent in this respect. Both isomers inhibited competitively the effect of isoprenaline on the trachea, the (-)-isomer being about 100 times more active than the (+)-isomer. None of the isomers showed any detectable positive inotropic effect on the papillary muscle but both inhibited competitively the response to isoprenaline. Also in this respect (-)-clenbuterol was more potent than (+)-clenbuterol. It is concluded that the beta 2-agonistic as well as the beta 1-antagonistic effect of clenbuterol resides in the (-)-isomer and that the (+)-isomer does not seem to contribute to the pharmacological effects displayed by racemic clenbuterol.     

TRPV1 activation is not an all-or-none event: TRPV1 partial agonism/antagonism and its regulatory modulation

TRPV1 has emerged as a promising therapeutic target for pain as well as a broad range of other conditions such as asthma or urge incontinence. The identification of resiniferatoxin as an ultrapotent ligand partially able to dissect the acute activation of TRPV1 from subsequent desensitization and the subsequent intense efforts in medicinal chemistry have revealed that TRPV1 affords a dramatic landscape of opportunities for pharmacological manipulation. While agonism and antagonism have represented the primary directions for drug development, the pharmacological complexity of TRPV1 affords additional opportunities. Partial agonism/partial antagonism, its modulation by signaling pathways, variable desensitization, and slow kinetics of action can all be exploited through drug design.     

The rise of body temperature induced by the stimulation of dopamine D1 receptors is increased in acutely reserpinized mice

In naive mice, the selective D1 agonist, SK&F 38393 (7.5-30 mg/kg s.c.), induced a significant rise of body temperature (0.5-1 degree C) which was antagonized by SCH 23390 (100 micrograms/kg s.c.) and by flupenthixol (0.4 mg/kg i.p.). In mice treated with reserpine (5 mg/kg s.c.) 18 h before testing, which on its own caused intense hypothermia (10-12 degrees C), SK&F 38393 (1.87-30 mg/kg s.c.) induced a dose-dependent and more marked rise of body temperature (5-7 degrees C). Similarly, SK&F 38393 (30 mg/kg s.c.) partially prevented reserpine-induced hypothermia. The central origin of the SK&F 38393 effects in reserpine-treated mice is indicated by the rise of body temperature induced by the i.c.v. administration of the drug (12.5-50 micrograms per mice). The SK&F 38393-induced rise of body temperature in acutely reserpinized mice was antagonized by SCH 23390 (50-200 micrograms/kg s.c.), clozapine (1.87-30 mg/kg i.p.) or chlorpromazine (2-32 mg/kg i.p.) but not by metoclopramide (25 or 100 mg/kg i.p.) or amisulpride (12.5 or 50 mg/kg). In naive mice, apomorphine (1 mg/kg s.c.) or LY 171555 (0.4 mg/kg s.c.) induced hypothermia which was antagonized by amisulpride (12.5 mg/kg i.p.); a transiently increased body temperature was even measured 30 min after apomorphine injection in amisulpride-treated mice. Apomorphine (1 mg/kg s.c.) induced a rise of body temperature in acutely reserpinized mice which was significantly reduced by SCH 23390 (50 and 200 micrograms/kg s.c.) and significantly increased by amisulpride (12.5 and 50 mg/kg i.p.). These data suggest that pharmacologically different dopamine receptor subtypes mediate different effects on body temperature in mice: D1 dopamine receptors mediate a rise of body temperature which is increased in hypothermic reserpinized animals and dopamine receptors of the D4 subtype mediate the decrease of body temperature in naive mice.     

Postsynaptic dopamine DA1- and DA2-receptors in jejunal arteries of rabbits.

Changes in the external diameter of rabbit isolated jejunal arteries were measured by a photoelectric device. Both norepinephrine and dopamine produced vasoconstriction; prazosin, but not idazoxan, antagonized these effects. Arteries preconstricted with norepinephrine were dilated by SKF 38393 and LY 171555. SCH 23390 antagonized both compounds, however, in the case of LY 171555 with a lower potency. Sulpiride also interacted with LY 171555. When arteries were preconstricted with PGF2 alpha in the presence of prazosin, idazoxan, and propranolol, SKF 38393 caused vasodilation, while LY 171555 and dopamine were inactive. We conclude that in jejunal arteries both DA1- and DA2-receptors may be present at the vascular smooth muscle.     

Enhancement of tetrodotoxin-induced axonal blockade by adenosine, adenosine analogues, dibutyryl cyclic AMP and methylxanthines in the frog sciatic nerve.

The effects of adenosine, adenosine analogues (N6-cyclohexyladenosine (CHA), L-N6-phenylisopropyladenosine (L-PIA), D-N6-phenylisopropyladenosine (D-PIA), N6-methyladenosine and 2-chloroadenosine), adenine, inosine, hypoxanthine, cyclic AMP and its analogue the dibutyryl cyclic AMP (db cyclic AMP), and methylxanthines (theophylline, caffeine and isobutylmethylxanthine (Ibmx) on compound action potentials were investigated in de-sheathed sciatic nerve preparations of the frog. Adenosine and its analogues enhanced, in a concentration-dependent manner, the inhibitory action of tetrodotoxin (TTX) on nerve conduction. The order of potencies was: CHA greater than D-PIA greater than L-PIA greater than N6-methyladenosine greater than 2-chloroadenosine greater than adenosine. The adenosine metabolites, inosine and hypoxanthine, were inactive on TTX-induced axonal blockade. Adenine enhanced the inhibitory action of TTX on nerve conduction, but was less effective than adenosine. db Cyclic AMP, but not cyclic AMP, mimicked the inhibitory effect of adenosine on nerve conduction. Methylxanthines did not antagonize the effect of adenosine on TTX-induced axonal block and in high concentrations also mimicked the effect of adenosine on nerve conduction. The possibility of adenosine acting on TTX-induced axonal block through an adenosine receptor positively coupled to adenylate cyclase is discussed.     

1-Methylisoguanosine: interaction with central adenosine receptors and lack of antagonism of its in vivo effects by a benzodiazepine antagonist

1-Methylisoguanosine, a marine natural product analogue of adenosine, with moderate activity as a benzodiazepine receptor ligand, has previously been shown to have muscle-relaxant and hypothermic activity in mice in vivo. The present experiments showed that the benzodiazepine antagonist Ro15-1788 did not block the in vivo muscle-relaxant and hypothermic effects of 1-methylisoguanosine, suggesting that these particular actions are not due to interactions with benzodiazepine receptors. When applied by microiontophoresis near spontaneously-active neurones or neurones activated by ACh, DL-homocysteate or glutamate in the ventrobasal thalamus of anaesthetized rats, 1-methylisoguanosine had a depressant action; it was similar to adenosine in potency and in its ability to be antagonized by 8-(parasulphophenyl)theophylline. The depression was usually longer lasting than that caused by adenosine, consistent with previous neurochemical data showing it to be resistant to adenosine deaminase and a poor substrate for the uptake system for adenosine in the CNS. These results suggest that the major pharmacological/behavioural actions of 1-methylisoguanosine in vivo are more likely to be caused by an interaction with adenosine receptors, rather than with benzodiazepine sites.