KATP channel openers for the treatment of airways hyperreactivity.
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Adenosine Triphosphate↗
Biomedical subjects
J R Fozard
Publications and source records attributed to J R Fozard.
Adenosine receptor ligands: potential as therapeutic agents in asthma and COPD.
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Animals↗
SCA 40: studies of the relaxant effects on cryopreserved human airway and vascular smooth muscle.
1. 6-Bromo-8-methylaminoimidazol[1,2-a]pyrazine-2carbonitrile (SCA 40) has been claimed to induce relaxation in guinea-pig trachea by opening high conductance, calcium-activated potassium (BKCa) channels. The mechanism of action of SCA 40 has now been further investigated in ring preparations from cryopreserved human airway and vascular smooth muscle preparations in vitro. 2. Human bronchi with spontaneous tone relaxed in response to SCA 40 in a biphasic way. A high affinity component (pD2 8.61 +/- 0.21; mean +/- s.e.mean) accounted for 30% of the response and a low affinity component (pD2 6.53 +/- 0.14) for the remaining 70%. In contrast, in bronchi contracted with carbachol, 1 microM, the concentration-response curve to SCA 40 was monophasic and yielded a pD2 of 6.31 +/- 0.29. 3. SCA 40 relaxed pulmonary and mesenteric arteries and peripheral veins which had been precontracted by 10 nM U46619 nearly completely and in a monophasic way; the pD2 values were 6.37 +/- 0.08, 6.17 +/- 0.15 and 5.45 +/- 0.25, respectively. 4. Lemakalim, an opener of ATP-dependent potassium (KATP) channels, also relaxed human bronchi under spontaneous tone and the vascular tissues. NS 1619, a recognised opener of BKca channels, was inactive up to 10 microM on bronchial and vascular tissues. 5. The SCA 40-induced relaxation of human bronchi was reduced concentration-dependently in the presence of high potassium chloride (20 and 80 mM). However, in the presence of 80 mM KCl and nifedipine, 30 nM, SCA 40 fully relaxed the remaining contractile response with pD2 values of 8.08 +/- 0.13 and 5.27 +/- 0.13 for the high and low affinity component, respectively. 6. Relaxation responses to SCA 40 in human bronchi were resistant to blockade by glibenclamide at concentrations up to 10 microM (which blocked the relaxant response to lemakalim), quinine (30 microM), apamin (100 nM), tetraethylammonium (0.1-1 mM) and charybdotoxin (10-100 nM), thus excluding the involvement of a variety of K+ channels including KATP and KCa channels. 7. In bronchi contracted with carbachol, 1 microM, the nature of the interaction between SCA 40 and the beta 2-adrenoceptor agonist, salbutamol, was synergistic. 8. These experiments establish that SCA 40 is a potent relaxant of human bronchial smooth muscle manifesting spontaneous tone. A low affinity relaxant component has its counterpart in the relaxation seen in both human arterial and venous smooth muscle. The consensus of the evidence suggests that K+ channel opening is not the basis of the relaxant response to SCA 40. Furthermore, BKCa channels appear to be of minor importance in the regulation of human airway smooth muscle tone. Our data suggest that inhibition of an adenosine 3':5'-cyclic monophosphate phosphodiesterase may contribute, at least to the low affinity relaxant component of SCA 40. However, the exact mechanism mediating the SCA 40-induced relaxation of human airways remains to be defined.
Albuterol↗
Mast cell degranulation following adenosine A3 receptor activation in rats.
The present studies were carried out to provide further evidence for the hypothesis that the hypotensive response to adenosine A3 receptor activation in the anaesthetized rat involves mediator release from mast cells. Male Sprague-Dawley rats were anaesthetized and given just supramaximal hypotensive doses of either the non-selective A3 receptor agonist, N6(-2)-(4-aminophenyl)ethyladenosine (APNEA: 100 micrograms/kg, preceded by the A1/A2 receptor antagonist, 8-p-(sulphophenyl)theophylline, to "isolate' the A3 receptor-mediated component of the response), the mast cell degranulating agent, compound 48/80 (300 micrograms/kg) or the vehicle for APNEA, intravenously. Blood was withdrawn from a carotid artery between 2 and 10 min after the injection and plasma and serum histamine concentrations measured. Samples of connective tissue (surrounding the abdominal musculature), thymus, mesenteric lymph node, kidney, skin and diaphragm were removed for histological analysis. The plasma and serum histamine concentrations were markedly and significantly higher in the APNEA- or compound 48/80-treated animals compared to vehicle-treated controls. Consistent with this, a substantially greater proportion of mast cells was seen to be undergoing degranulation in all tissues removed from animals treated with APNEA or compound 48/80 compared to those from rats treated with vehicle. Thus, adenosine A3 receptor activation results in rapid mast cell degranulation in the anaesthetized rat. The data provide further evidence of a key role for the mast cell in adenosine A3 receptor-mediated hypotension in this species.
Adenosine↗
Subtypes of the type 4 cAMP phosphodiesterases: structure, regulation and selective inhibition.
The 'famille nombreuse' of cyclic nucleotide phosphodiesterases, responsible for degrading the ubiquitous second messenger molecules, cAMP and cGMP, maintains its place as a major focus of interest for many research laboratories in both academia and industry. The increase in knowledge of the primary sequences, plus the availability of selective inhibitors, are rapidly improving our insight into the structure, regulation and function of these pivotal enzymes of cellular homeostasis. Here, Thomas Müller, Peter Engels and John Fozard focus on family 4 of the phosphodiesterases, which is of particular interest owing to both the number of genes (and splice variants) and the emergence of selective inhibitors, which are enabling the functional significance of these enzymes to be defined.
3',5'-Cyclic-AMP Phosphodiesterases↗
Salbutamol-induced airway hyperreactivity in guinea pigs is not due to a loss of its bronchodilator effect.
Guinea pigs were treated for 10 days with (+/-)-salbutamol (0.2 mg/kg/day, delivered from subcutaneously implanted osmotic minipumps). Airway reactivity to intravenously administered histamine, methacholine and bombesin was substantially increased in salbutamol-treated guinea pigs relative to controls. In the same animals, the potency of intravenously administered salbutamol to reverse bombesin-induced bronchoconstriction remained unchanged thus exactly reflecting effects in man. In conclusion, subchronic administration of salbutamol at low doses to guinea pigs increases airway reactivity. Since the bronchorelaxant effect of salbutamol remained unchanged, desensitisation of beta-adrenoceptors on airway smooth muscle is unlikely to account for this effect.
Adrenergic beta-Agonists↗
Further analysis of the mechanisms underlying the tracheal relaxant action of SCA40.
1. SCA40 (1nM-10 microM), isoprenaline (1-300 nM) and levcromakalim (100 nM-10 microM) each produced concentration-dependent suppression of the spontaneous tone of guinea-pig isolated trachea. Propranolol (1 microM) markedly (approximately 150 fold) antagonized isoprenaline but did not antagonize SCA40. The tracheal relaxant action of SCA40 was unaffected by suramin (100 microM) or 8-(p)-sulphophenyltheophylline (8-SPT; 140 microM). 2. An isosmolar, K(+)-rich (80 mM) Krebs solution increased tracheal tone, antagonized SCA40 (approximately 60 fold), antagonized isoprenaline (approximately 20 fold) and very profoundly depressed the log concentration-effect curve for levcromakalim. Nifedipine (1 microM) did not itself modify the relaxant actions of SCA40, isoprenaline or levcromakalim. However, nifedipine prevented the rise in tissue tone and the antagonism of SCA40 and isoprenaline induced by the K(+)-rich medium. In contrast, nifedipine did not prevent the equivalent antagonism of levcromakalim. 3. Charybdotoxin (100 nM) increased tracheal tone, antagonized SCA40 (approximately 4 fold) and antagonized isoprenaline (approximately 3 fold). Nifedipine (1 microM) prevented the rise in tissue tone and the antagonism of SCA40 and isoprenaline induced by charybdotoxin. 4. Quinine (30 microM) caused little or no change in tissue tone and did not modify the relaxant action of isoprenaline. However, quinine antagonized SCA40 (approximately 2 fold). Nifedipine (1 microM) prevented the antagonism of SCA40 induced by quinine. 5. Tested on spontaneously-beating guinea-pig isolated atria SCA40 (1 nM-10 microM) increased the rate of beating in a concentration-dependent manner. Over the concentration-range 1 microM-10 microM, SCA40 also caused an increase in the force of atrial contraction. 6. Intracellular electrophysiological recording from guinea-pig isolated trachealis showed that the relaxant effects of SCA40 (1 micro M) were often accompanied by the suppression of spontaneous electrical slow waves but no change in resting membrane potential. When the concentration of SCA40 was raised to 10 micro M, its relaxant activity was accompanied both by slow wave suppression and by plasmalemmal hyperpolarization.7. SCA40 (10 nM- 100 micro M) more potently inhibited the activity of cyclic AMP phosphodiesterase (PDE)than that of cyclic GMP PDE derived from homogenates of guinea-pig trachealis. Theophylline(1 micro M- 1O mM) also inhibited these enzymes but was less potent than SCA40 in each case and did not exhibit selectivity for inhibition of cyclic AMP hydrolysis.8. Tested against the activity of the isoenzymes of cyclic nucleotide PDE derived from human blood cells and lung tissue, SCA40 proved highly potent against the type III isoenzyme. It was markedly less potent against the type IV and type V isoenzymes and even less potent against the isoenzymes types I and II.9. It is concluded that the tracheal relaxant action of SCA40 (1 nM- 1 micro M) does not involve the activation of beta-adrenoceptors or P1 or P2 purinoceptors. Furthermore, this action is unlikely to depend upon the opening of BKca channels with consequent cellular hyperpolarization and voltage-dependent inhibition of Ca2+ influx. The tracheal relaxant action of SCA40 (up to 1 micro M) is more likely to depend upon its selective inhibition of the type III isoenzyme of cyclic nucleotide PDE. At concentrations above 1 micro M, SCA40 exerts more general inhibition of the isoenzymes of cyclic nucleotide PDE and may then promote the opening of BKca channels.
Animals↗
A role for mast cells in adenosine A3 receptor-mediated hypotension in the rat.
1. The adenosine A3 receptor agonist, N6-2-(4-aminophenyl)ethyladenosine (APNEA) induces hypotension in the anaesthetized rat. The present experiments were carried out to explore the role of mast cells in the response. 2. Intravenous injection of APNEA (1-30 micrograms kg-1 to rats in which the A3 receptor-mediated response had been isolated by pretreatment with 8-(p-sulphophenyl) theophylline (8-SPT)), induced dose-related falls in blood pressure accompanied at higher doses by small falls in heart rate. Responses to the mast cell degranulating agent, compound 48/80 (10-300 micrograms kg-1, i.v.) were qualitatively similar to those to APNEA. 3. Pretreatment with sodium cromoglycate (0.25-20 mg kg-1, i.v.) induced dose-related, although incomplete, blockade of the hypotensive responses to APNEA. At 20 mg kg-1, sodium cromoglycate also inhibited the cardiovascular response to compound 48/80 but had no effects on those to the selective A1 receptor agonist, N6-cyclopentyladenosine (CPA) or the selective A2A receptor agonist, 2-[p-(2-carboxyethyl)phenylamino]-5'-N-ethylcarboxamidoadenosine (CGS 21680). Lodoxamide (0.01-20 mg kg-1) also blocked selectively but incompletely the response to APNEA. 4. The cardiovascular responses to compound 48/80 (10-300 micrograms kg-1, i.v.) were markedly suppressed in animals which had received repeated doses of the compound by the intraperitoneal route. Similarly APNEA was essentially devoid of cardiovascular activity in such preparations. In contrast, responses to CPA were similar in animals treated repeatedly with compound 48/80 to those obtained in control animals. 5. Plasma and serum histamine concentrations were markedly increased associated with the pronounced hypotensive responses induced by intravenous injections of APNEA (30 or 100 microg kg-1) in the presence of 8-SPT, or compound 48/80 (300 microg kg-1).6. Taken together the data implicate the mast cell in a key role in adenosine A3 receptor-mediated hypotension in the anaesthetized rat.
Adenosine↗
BW-A522 blocks adenosine A3 receptor-mediated hypotensive responses in the rat.
The effects of 3-(3-iodo-4-aminobenzyl)-8-(4-oxyacetate)-1-propylxanthine (I-ABOPX; BW-A522), which has nanomolar affinity for the recently cloned human and sheep adenosine A3 receptor, on the putative A3 receptor mediated hypotensive response to N6-2-(4-aminophenyl)ethyl adenosine (APNEA) in the rat have been investigated. Following blockade of A1 and A2 receptors with 8-(p-sulphophenyl)theophylline, BW-A522, 10 and 40 mg/kg i.v., blocked dose-dependently and surmountable the hypotensive response to APNEA. The results provide direct evidence of an A3 receptor in the cardiovascular system of the rat which induces hypotension when activated.
Adenosine↗
5-Hydroxytryptamine (5-HT) and the initiation of migraine: new perspectives.
The hypothesis that 5-hydroxytryptamine (5-HT) acting through 5-HT2C receptors is a key factor in the initiation of migraine has been re-evaluated in the light of recent basic and clinical scientific developments. The key findings are that nitric oxide is an important trigger for migraine, that 5-HT2B/5-HT2C receptors are present on endothelial cells and trigger nitric oxide release when activated and that supersensitivity of the 5-HT2B/5-HT2C receptor is a neurochemical feature predisposing to headache. Taken together the data bring new perspectives to the role of 5-HT acting through 5-HT2C (or closely similar) receptors in the initiation of migraine.
Animals↗
Cardiovascular effects of cyclosporine A and OG 37-325 after chronic administration to conscious rats: role of endothelin.
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Animals↗
International Union of Pharmacology classification of receptors for 5-hydroxytryptamine (Serotonin).
It is evident that in the last decade or so, a vast amount of new information has become available concerning the various 5-HT receptor types and their characteristics. This derives from two main research approaches, operational pharmacology, using selective ligands (both agonists and antagonists), and, more recently, molecular biology. Although the scientific community continues to deliberate about the hierarchy of criteria for neurotransmitter receptor characterisation, there seems good agreement between the two approaches regarding 5-HT receptor classification. In addition, the information regarding transduction mechanisms and second messengers is also entirely consistent. Thus, on the basis of these essential criteria for receptor characterisation and classification, there are at least three main groups or classes of 5-HT receptor: 5-HT1, 5-HT2, and 5-HT3. Each group is not only operationally but also structurally distinct, with each receptor group having its own distinct transducing system. The more recently identified 5-HT4 receptor almost undoubtedly represents a fourth 5-HT receptor class on the basis of operational and transductional data, but this will only be definitively shown when the cDNA for the receptor has been cloned and the amino acid sequence of the protein is known. Although those 5-HT receptors that have been fully characterised and classified to date (and, hence, named with confidence) would seem to mediate the majority of the actions of 5-HT throughout the mammalian body, not all receptors for 5-HT are fully encompassed within our scheme of classification. These apparent anomalies must be recognised and need further study. They may or may not represent new groups of 5-HT receptor or subtypes of already known groups of 5-HT receptor. Even though the cDNAs for the 5-ht1E, 5-ht1F, 5-ht5, 5-ht6, and 5-ht7 receptors have been cloned and their amino acid sequence defined, more data are necessary concerning their operational and transductional characteristics before one can be confident of the suitability of their appellations. Therefore, it is important to rationalise in concert all of the available data from studies involving both operational approaches of the classical pharmacological type and those from molecular and cellular biology.(ABSTRACT TRUNCATED AT 400 WORDS)
Animals↗
Effect of pertussis toxin treatment on the putative adenosine A3 receptor-mediated hypotensive response in the rat.
The effect of pertussis toxin pretreatment on the putative A3 receptor-mediated hypotensive response to N6-2-(4-aminophenyl)ethyladenosine (APNEA) has been investigated in the anaesthetised rat. Pertussis toxin pretreatment essentially abolished the bradycardia induced by the prototype A1 receptor agonist, N6-cyclopentyladenosine, whereas the fall in blood pressure induced by the selective A2A receptor agonist, CGS 21680 was enhanced. Pertussis toxin substantially reduced the hypotensive response to APNEA. In this respect, the mechanism of action resembles that of the cloned A3 receptor which when expressed in CHO cells couples negatively to adenylate cyclase by a pertussis toxin-sensitive mechanism. The data provide further evidence that adenosine A3 receptors mediate the hypotensive response to APNEA in the rat.
Adenosine↗
SDZ 216-525, a selective and potent 5-HT1A receptor antagonist.
The pharmacological properties of SDZ 216-525, methyl 4-(4-[4-(1,1,3-trioxo-2H-1,2-benzoisothiazol-2-yl)butyl]-1-p iperazinyl)1H- indole-2-carboxylate, a new selective and potent 5-HT1A receptor antagonist, are described in vitro (and comparisons made with those of MDL 73005 and NAN 190, two putative 5-HT1A receptor antagonists) and in vivo. In radioligand binding studies, SDZ 216-525 showed high affinity and selectivity for 5-HT1A sites (pKD = 9.2) as compared to 5-HT1B, 5-HT1C, 5-HT1D, 5-HT2 and 5-HT3 sites (pKD = 6.0, 7.2, 7.5, 5.2 and 5.4, respectively). The affinity of the compound for alpha 1, alpha 2, beta 1 and beta 2 adrenoceptors, and dopamine D2 receptors was at least 50-100 times lower than for 5-HT1A sites. The effects of SDZ 216-525, MDL 73005 and NAN 190 on 5-HT1 receptor-linked second messengers were characterised in the following tests: inhibition of forskolin-stimulated adenylate cyclase activity in calf hippocampus (5-HT1A), rat substantia nigra (5-HT1B) and calf substantia nigra (5-HT1D) and stimulation of inositol phosphate production in pig choroid plexus (5-HT1C). SDZ 216-525 potently antagonised the effects of 8-OH-DPAT (8-hydroxy-2-[N-dipropyl-amino]-tetralin) on 5-HT1A receptors (pKB = 10) and displayed no intrinsic activity in this test, whereas it behaved at best as a weak antagonist on the other receptor models (pKB values < 6.9).(ABSTRACT TRUNCATED AT 250 WORDS)
8-Hydroxy-2-(di-n-propylamino)tetralin↗
The cardiovascular effects of selective adenosine A1 and A2 receptor agonists in the pithed rat: no role for glibenclamide-sensitive potassium channels.
The cardiovascular effects of N6-cyclopentyladenosine (CPA), a selective adenosine A1 receptor agonist and 2-[p-carboxyethyl)phenylethylamino]-5'-N-ethylcarboxamidoadenosine (CGS 21,680), a selective A2 receptor agonist have been investigated in the pithed rat with blood pressure raised to normal levels with angiotensin II. Cumulative intravenous administration of CPA, 0.3-10 micrograms/kg, induced dose-related falls in blood pressure and heart rate; over the same dose range CGS 21,680 induced hypotension but no bradycardia. Pretreatment with a maximally effective dose of the A1/A2 receptor antagonist 8-(p-sulphophenyl) theophylline (8-SPT) blocked the bradycardiac effects of CPA (92-fold) more effectively than its hypotensive activity (5.1-fold); the vasodepressor effects of CGS 21,680 were blocked 19-fold by 8-SPT. Glibenclamide, a blocker of ATP-sensitive potassium (K+ATP) channels, administered intravenously at 20 mg/kg markedly attenuated the vasodepressor effects of the potassium channel opener, (-)-(3S,4R)-3,4-dihydro-3-hydroxy-2,2-dimethyl-4-(3-oxo-cyclopent- 1-enyloxy)-2H-1-benzopyran-6-carbonitrile (SDZ PCO 400). In contrast, neither the hypotensive nor the bradycardic effects of CPA nor the fall in blood pressure following CGS 21,680 was significantly affected by pretreatment with glibenclamide. These results indicate that a significant component of the blood pressure fall induced by CPA and CGS 21,680 in the pithed rat with blood pressure supported by angiotensin II occurs by a mechanism which is insensitive to 8-SPT and unlikely, therefore, to be mediated by A1 or A2 receptors. Moreover, in contrast to the prevailing literature, the cardiovascular effects arising from adenosine receptor activation in this model are not mediated by glibenclamide-sensitive, K+ATP channels.
Adenosine↗
Adenosine A3 receptors: two into one won't go.
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Animals↗
Adenosine A3 receptors mediate hypotension in the angiotensin II-supported circulation of the pithed rat.
The cardiovascular effects of N6-2-(4-aminophenyl)ethyladenosine (APNEA), which when radiolabelled with 125I shows high affinity for the newly described adenosine A3 receptor, have been investigated in the angiotensin II-supported circulation of the pithed rat. APNEA induces hypotensive responses which are unaffected by high doses (20-40 mg kg-1) of the broad spectrum, adenosine receptor antagonist, 8-(p-sulphophenyl)theophylline (8-SPT). 8-SPT-resistant falls in blood pressure are also seen, in the absence of bradycardia, with 5'-N-ethylcarboxamidoadenosine (NECA) and the R- and S-enantiomers of N6-phenylisopropyladenosine (PIA). Xanthine insensitivity, high potencies of APNEA, NECA and R-PIA, and an enantiomeric selectivity favouring R- over S-PIA are distinguishing features of the adenosine A3 receptor. We suggest that hypotension in the pithed rat may be a functional correlate of this site.
Adenosine↗
The novel 5-HT1A receptor antagonist, SDZ 216-525, decreases 5-HT release in rat hippocampus in vivo.
1. Recent evidence suggests that the novel compound SDZ 216-525 is a selective and possibly silent 5-HT1A receptor antagonist. Here we have examined the action of SDZ 216-525 on central 5-HT1A autoreceptor function. The experiments involved measurement of drug effects on extracellular 5-HT in the ventral hippocampus of the chloral hydrate anaesthetized rat by use of microdialysis. 2. Acute injection of SDZ 216-525 (0.1, 0.3, 1.0 and 3 mg kg-1, s.c.) caused a dose-related decrease in 5-HT output with an estimated ED50 of at least 0.3 mg kg-1. This ED50 value is 20-30 times greater than ED50 values previously obtained for 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) and NAN-190. In comparison, SDZ 216-525 is reported to have slightly higher affinity for the 5-HT1A site than 8-OH-DPAT and NAN-190. 3. The inhibitory effect of SDZ 216-525 (1 mg kg-1, s.c.) on 5-HT was blocked by the 5-HT1/beta-adrenoceptor antagonist, (-)-pindolol (8 mg kg-1, s.c.) but not by a combination of the beta 1- and beta 2-selective adrenoceptor antagonists metoprolol and ICI 118,551 (4 mg kg-1, each). 4. Although in several experimental models SDZ 216-525 has high affinity, selectivity and lacks intrinsic activity at the 5-HT1A receptor, our experiments show that the drug decreases extracellular 5-HT in ventral hippocampus of the chloral hydrate anaesthetized rat via a pindolol-sensitive mechanism. We conclude that either SDZ 216-525 promotes (with low potency in vivo) 5-HT1A receptor/G-protein interactions, or that the 5-HTlA autoreceptor is a 5-HT1A receptor subtype different from the postsynaptic 5-HT1A receptor.
8-Hydroxy-2-(di-n-propylamino)tetralin↗