Bror Jonzon

Adenosine Receptors Mediating Cyclic AMP Productioin the Rat Hippocampus

Abstract: In the transversely cut rat hippocampus, adenosine caused a dose‐dependent increase in the accumulation of [3H]cyclic AMP from [3H]ATP. Adenosine breakdown products were inactive. AMP was somewhat less effective than adenosine, and its effect could be partially, but not completely, abolished by α,β‐methylene‐ADP and GMP, which inhibited its metabolism by 5′‐nucleotidase. The effect of adenosine was unaffected by inhibitors of adenosine deaminase, but enhanced by several inhibitors of adenosine uptake. Some analogues of adenosine, including N6‐phenylisopropyladenosine (PIA), 2‐chloroadenosine and adenosine 5′‐ethylcarboxamide (NECA), were more active than adenosine, whereas others such as 2‐deoxyadenosine and 9‐(tetrahydro‐2‐furyl)adenine (SQ 22536) actually inhibited the response. The effect of PIA was highly stereospecific. The action of adenosine was inhibited by several alkylxanthines, the most potent of which was 8‐phenyltheophylline. [3H]Cyclohexyladenosine (CHA) bound specifically to cell membranes from the rat hippocampus. The extent of binding was similar to that found in other cortical areas. The relative potency of some adenosine analogues and alkylxanthines to displace labelled CHA was essentially similar to their potency as effectors of the cyclic AMP system. Adenosine contributed to the cyclic AMP‐elevating effect of α‐adrenoceptor‐stimulating drugs and several amino acids, but not to that seen with isoprenaline. The cyclic AMP increase seen following depolarization was only partially adenosine‐dependent. The present results demonstrate that the rat hippocampus contains adenosine receptors mediating cyclic AMP accumulation and that these receptors have similar characteristics to those mediating pyramidal cell depression. Adenosine‐induced cyclic AMP accumulation may be used as a biochemical correlate to electrophysiology and as a convenient parameter to assess the influence of drugs on adenosine mechanisms in the rat hippocampus.

Release of purines, noradrenaline, and GABA from rat hippocampal slices by field stimulation.

Labelled adenine, noradrenaline (NA), and γ‐aminobutyric acid (GABA) were taken up by the transversely cut hippocampal slice. [3H]NA and [14C]GABA were retained as such, [3H]‐ (or [14C]‐) adenine mainly as adenine nucleotides. There was a spontaneous overflow of all three types of compounds ranging from 0.1 (GABA) to 0.21 (NA) %/min. The rate of [3H]NA overflow increased rapidly during electrical field stimulation. The release rate was well maintained over a 15‐min period. The rate of [14C]GABA release also increased rapidly but it was not maintained over a 15‐min period even if uptake and/or metabolism was inhibited by nipecotic acid (1 mM) and aminooxyacetic acid (AOAA, 0.1 mM). The bulk of the purines was released after the stimulation period. For all compounds the amounts released were frequency‐ and calcium‐dependent. At a frequency of 3 Hz a 10 V stimulation was sufficient to cause a maximal [3H]NA release and 20 V to cause maximal [14C]GABA release, but 14C‐purine release was increased further by increasing the voltage to 40 V. The evoked purine release was inhibited by a nucleoside uptake inhibitor (dipyridamole). On stimulation of [3H]NA‐labelled slices the released radioactivity was composed of >95% unchanged NA. The specific activities of NA in the slice and in the superfusate were practically identical. In [3H]adenine‐labelled slices the released radioactivity was composed of adenosine, inosine, and hypoxanthine, but the activity in the slice of ATP, ADP, and AMP. The specific activity of the released purines was about four times higher than that of the purines retained in the slice but similar to that of the hypoxanthine in the slice. It is suggested that the purine release induced by electrical field stimulation differs in important respects from the release of a traditional neurotransmitter such as NA and that it depends on the intracellular formation of adenosine from a labile fraction of adenine nucleotides that is affected by neurotransmission.

On the mechanism by which methylxanthines enhance apomorphine-induced rotation behaviour in the rat

Methylxanthines, such as caffeine and theophylline, potentiate the rotation behaviour induced by dopamine receptor agonists in rats with unilateral lesions of the nigro-striatal pathway. In the present study we have examined the possibility that interaction with central adenosine mechanisms could influence rotation behaviour. Under in vitro conditions adenosine and N6-phenylisopropyl-adenosine (PIA) stimulate cyclic AMP accumulation. This effect was enhanced by the phosphodiesterase inhibitor rolipram, but blocked by alkylxanthines such as caffeine, theophylline and, particularly, 8-phenyl-theophylline. Rotation behaviour induced by apomorphine (0.05 mg/kg), was inhibited by PIA and rolipram and by a low dose of the adenosine deaminase inhibitor EHNA (2 mg/kg). By contrast, theophylline and 8-phenyl-theophylline caused a potentiation. The former drug stimulated rotation behaviour per se, while the latter did not. 8-Phenyl-theophylline entered the brain poorly and its concentration in brain it was less than 1/10 of theophylline. It is concluded that theophylline does not potentiate rotation behaviour secondarily to inhibition of phosphodiesterase. Antagonism of endogenous adenosine may partly explain the effect of methylxanthines. Possibly, some as yet unknown mechanism may also contribute to the effects of xanthine-derivatives on rotation behaviour.

Effect of adenosine receptor agonists and other compounds on cyclic AMP accumulation in forskolin-treated hippocampal slices

Summary1.The effect of adenosine analogues and some putative neurotransmitters have been studied on cyclic AMP accumulation in rat hippocampal slices treated with the adenylate cyclase activator forskolin.2.The effects of PGE2 and histamine were potentiated by forskolin (0.1 μM). Isoprenaline and NECA had essentially additive effects with 0.1 μM forskolin and serotonin (above 10−4 M) inhibited forskolin-stimulated cyclic AMP accumulation.3.The A1-adenosine receptor selective adenosine analogue R-PIA inhibited forskolin stimulated cyclic AMP accumulation in low doses and stimulated in high. NECA, adenosine and 2-chloroadenosine uniformly stimulated cyclic AMP accumulation. 2′,5′-dideoxyadenosine inhibited, but only at high concentrations.4.Both the stimulatory and the inhibitory effects of R-PIA were antagonized by 8-phenyltheophylline (10 μM). Enprofylline (100 μM) selectively inhibited the stimulatory effect. In the presence of enprofylline both 2-chloroadenosine showed an inhibitory effect on cyclic AMP accumulation.5.It is concluded that the forskolin-treated rat hippocampal slice is a useful preparation to study both stimulatory and inhibitory effects of transmitters and modulators on adenylate cyclase. The results also show that the rat hippocampus has both A1-receptors that are linked to inhibition of cyclic AMP accumulation and A2-receptors that are linked to stimulation. Furthermore, enprofylline is shown to selectively antagonize the stimulatory response, revealing inhibitory effects of compounds such as 2-chloroadenosine and adenosine.

Evidence that a novel 8-phenyl-substituted xanthine derivative is a cardioselective adenosine receptor antagonist in vivo.

The ability of caffeine, enprofylline (3-pro-pylxanthine), 8-phenyltheophylline, 8-p-sulphophenyl-theophylline, 8-(4-carboxymethyloxyphenyl)-l,3-dipro-pylxanthine, and 8-(4-carboxymethyloxyphenyl)-l,3-di-propylxanthine-2-aminoethylamide (XAC) to antagonize the effects of an adenosine analogue, N-5-ethylcarbox-amidoadenosine, on heart rate and blood pressure in anesthetized rats was examined. The first five xanthine derivatives were equally active in antagonizing the two responses. By contrast, XAC was ∼20 times more potent in antagonizing the heart rate response than the blood pressure response. Measurements of the concentration of XAC in plasma and brain indicate that it penetrates the central nervous system poorly. It is concluded that XAC is a cardioselective adenosine antagonist, and since adenosine is supposed to reduce heart rate via an effect on A,-receptors, and the blood pressure via A2-receptors, XAC may be a selective A,-adenosine receptor antagonist in vivo

Adenosine mechanisms are not affected by antidepressant concentrations of desipramine

We have evaluated the proposal that adenosine may mediate some of the effects of tricyclic antidepressant therapy. In‐vitro desipramine (DMI) (1–10 μm) did not affect adenosine or 2‐chloroadenosine‐induced inhibition of lipolysis or the adenosine stimulated formation of cyclic (c)AMP in the hippocampal slice. However, very high concentrations of desipramine (0·2–0·5 mm) as well as some detergents potentiated the stimulatory effect of adenosine on cAMP formation. The ATP, ADP and AMP contents in slices were unaffected as was the electrically evoked release of purines. Long‐term treatment in‐vivo with antidepressants in clinically relevant doses did not alter the sensitivity of adenosine receptor mediated cAMP formation in‐vitro while the β‐adrenoceptor‐mediated formation was depressed by desipramine or imipramine treatment but not by zimelidine or fluoxetine treatment. It is concluded that actions on central adenosine mechanisms are unlikely to play any important role in the therapeutic effects of tricyclic antidepressants.

Formation and Actions of Adenosine in the Rat Hippocampus, with Special Reference to the Interactions with Classical Transmitters

Adenosine and related compounds can probably play a transmitter or cotransmitter role at least in some tissues (e.g., Burnstock, 1985). However, this seems to be rare and the major functional role of adenosine not only in the periphery but also in the nervous system appears to be that of a modulator. Adenosine is released not only from nerve terminals but also from cell bodies and from glial cells or endothelial cells lining the cerebral blood vessels. Besides its role in blood-flow regulation, adenosine clearly plays a role in modulation of nervous activity, pre- and postjunctionally.

Nicotine Enhances Angina Pectoris-like Chest Pain and Atrioventricular Blockade Provoked by Intravenous Bolus of Adenosine in Healthy Volunteers

Summary: An attempt was made to study possible interaction between neuromodulation by adenosine and nicotine stimulatory effects. Dose–effect curves were made double blind in 7 nonsmoking, nonsnuffing healthy volunteers (25–49 years) before and during exposition to nicotine roughly corresponding to the nicotine of one cigarette, 2 mg ingested from a chewing gum (800 chews during 20 min). Chest pain was estimated by the Borg CR-10 scale. ECG was followed, and respiration was recorded continuously by spirometry. Maximal tolerable dose of adenosine was 12.7 ±3.0 mg. Chest pain increased dose dependently to 5.7 ± 1.7 units. Nicotine increased the pain response by 20 ± 15%, (p < 0.02). The total time with atrioventricular (AV) block provoked by adenosine increased with nicotine (7 ± 12%, p < 0.03) while increased ventilation provoked by adenosine was unaffected by nicotine. In conclusion, interaction between adenosine and nicotine was demonstrated. Nicotine enhanced both stimulatory (chest pain) and inhibitory actions (AV-block) of adenosine.

Effects of Adenosine Agonists on Camp Accumulation in Adult and Newborn Rat Aortic Smooth Muscle Cell Cultures

Abstract In aortic smooth muscle cell cultures of adult and newborn rats, the NECA-induced CAMP accumulation was enhanced in adults. The inhibitory effect of the A,-receptor is unlikely to be the main factor responsible for this large difference.

Adenosine Stimulated Respiration and Pain in Man: Examples of Adenosine Receptor Modulated Afferent Information

Cellular adenosine formation and release is enhanced by hypoxia or lack of energy supply. Adenosine acts in several ways to balance the supply to the cellular demands. In 1929 Drury and Szent-Gyorgyi showed that adenosine produced several biological effects such as hypotension and bradycardia. Since then different effects of adenosine have been studied. For example, apart from its vasodilatory effect adenosine inhibits lipolysis, aggregation of platelets, release of noradrenaline and acetylcholine, hippocampal pyramidal excitability, motor behaviour and even respiration. Some of the actions of adenosine may be due to uptake into cells and further metabolic effects but most of the effects are mediated via specific adenosine cell surface receptors. Methylxanthines such as caffeine and theophylline, in the clinical relevant concentration range, act as competetive adenosine receptor antagonists (for review see Fredholm and Jonzon 1987). Adenosine is rapidly eliminated mainly by cellular uptake. The half life of adenosine in blood is approximately 10 seconds. Dipyridamole blocks the carrier mediated transport of adenosine across the cell membrane and thereby enhances the extracellular concentration and the receptor mediated effects of administered adenosine. We have studied the respiratory stimulation and estimated the pain response of bolus injections of adenosine to humans.