An EP receptor with a novel pharmacological profile in the T-cell line Jurkat.

1. Comparison of the rank order of potency of the natural prostanoids prostaglandin E2 (PGE2), PGD2, PGF2 alpha and carbaprostacyclin in stimulating cyclic AMP in Jurkat cells is consistent with the presence of an EP receptor. 2. Lack of responsiveness to the EP1/EP3 selective agonist, sulprostone, and the EP2 agonists, butaprost and AH 13205, indicates that this receptor is not of the EP1, EP2 or EP3 subtypes. 3. Inhibition of PGE2-stimulated cyclic AMP by the EP4 antagonist, AH 23848 is non-competitive, unlike the competitive antagonism reported in the pig saphenous vein EP4 preparation. Furthermore, 16,16-dimethyl PGE2 is 100 fold less potent than PGE2 in Jurkat cells, while these agonists are equipotent in the rabbit jugular vein purported EP4 preparation. In addition, 1-OH PGE1, which also is active in the rabbit jugular vein preparation, is inactive in Jurkat cells at concentrations up to 1 x 10(-4) M. These data are not wholly consistent with any adenylate cyclase coupled EP receptor described to date. 4. It is postulated that an EP receptor, positively coupled to adenylate cyclase, with a unique pharmacological profile is present in Jurkat cells.     

Prostaglandin E(2) increases cyclic AMP and inhibits endothelin-1 production/secretion by guinea-pig tracheal epithelial cells through EP(4) receptors

Prostaglandin E(2) (PGE(2)) increased adenosine 3' : 5'-cyclic monophosphate (cyclic AMP) formation in tracheal epithelial cells and concomitantly decreased the production/secretion of immunoreactive endothelin (irET). Naturally occurring prostanoids and selective and non-selective EP receptor agonists showed the following rank order of potency in stimulating cyclic AMP generation by epithelial cells: PGE(2) (EP-selective)>16,16-dimethyl PGE(2) (EP-selective)>11-deoxy PGE(2) (EP-selective)>iloprost (IP/EP(1)/EP(3)-selective), butaprost (EP(2)-selective), PGD(2) (DP-selective), PGF(2alpha) (FP-selective). The lack of responsiveness of the latter prostanoids indicated that the prostanoid receptor present in these cells is not of the DP, FP, IP, EP(1), EP(2) or EP(3) subtype. Pre-incubating the cells with the selective TP/EP(4)-receptor antagonists AH23848B and AH22921X antagonized the PGE(2)-evoked cyclic AMP generation. This suggested that EP(4) receptors mediate PGE(2) effects. However, in addition to any antagonistic effects at EP(4)-receptors, both compounds, to a different extent, modified cyclic AMP metabolism. The selective EP(1), DP and EP(2) receptor antagonist (AH6809) failed to inhibit PGE(2)-evoked cyclic AMP generation which confirmed that the EP(2) receptor subtype did not contribute to the change in cyclic AMP formation in these cells. The PGE(2)-induced inhibition of irET production by guinea-pig tracheal epithelial cells was due to cyclic AMP generation and activation of the cyclic AMP-dependent protein kinase since this effect was reverted by the cyclic AMP antagonist Rp-cAMPS. These results provide the first evidence supporting the existence of a functional prostaglandin E(2) receptor that shares the pharmacological features of the EP(4)-receptor subtype in guinea-pig tracheal epithelial cells. These receptors modulate cyclic AMP formation as well as ET-1 production/secretion in these cells.     

Mechanisms underlying the relaxation response induced by bradykinin in the epithelium-intact guinea-pig trachea in vitro

In this study, we investigated some of the signalling pathways involved in bradykinin (BK)-induced relaxation in epithelium-intact strips of the guinea-pig trachea (GPT + E). BK induced time- and concentration-dependent relaxation of GPT + E. Similar responses were observed for prostaglandin E2 (PGE2) or the combination of subthreshold concentrations of BK plus PGE2. The nonselective cyclooxygenase (COX) inhibitors indomethacin or pyroxicam, or the selective COX-2 inhibitors DFU, NS 398 or rofecoxib, but not the selective COX-1 inhibitor SC 560, all abolished BK-induced relaxation. The tyrosine kinase inhibitors herbimycin A and AG 490 also abolished BK-induced relaxation in GPT + E. The nonselective nitric oxide synthase (NOS) inhibitor 7-NINA concentration-dependently inhibited BK effects. BK-induced relaxation was prevented by the selective antagonists for EP3 (L 826266), but not by EP1 (SC 19221), EP1/EP2 (AH 6809) or EP4 (L161982) receptor antagonists. Otherwise, the selective inhibitors of protein kinases A, G and C, mitogen-activated protein kinases, phospholipases C and A2, nuclear factor-kappaB or potassium channels all failed to significantly interfere with BK-mediated relaxation.BK caused a marked increase in PGE2 levels, an effect that was prevented by NS 398, HOE 140 or AG 490. COX-2 expression did not differ in preparations with or without epithelium, and it was not changed by BK stimulation. However, incubation with BK significantly increased the endothelial NOS (eNOS) and neuronal NOS (nNOS) expression, independent of the epithelium integrity. Our results indicate that BK-induced relaxation in GPT + E depends on prostanoids (probably PGE2 acting via EP3 receptors) and NO release and seems to involve complex interactions between kinin B2 receptors, COX-2, nNOS, eNOS and tyrosine kinases.     

Characterization of the prostanoid receptor types involved in mediating calcitonin gene-related peptide release from cultured rat trigeminal neurones.

1. Prostaglandins and the vasodilator neuropeptide, calcitonin-gene related peptide (CGRP), have both been implicated in the pathogenesis of migraine headache. We have used primary cultures of adult rat trigeminal neurones to examine the effects of prostanoids on CGRP release in vitro. 2. CGRP release was stimulated by prostaglandin E2 (PGE2) and the IP receptor agonist, carbaprostacyclin (cPGI2). These responses were extracellular calcium-dependent, and the PGE2-induced CGRP release was unaltered by inhibition of nitric oxide synthase (NOS), ATP receptor blockade, or the addition of adenosine deaminase. 3. Increases in CGRP levels were also observed in response to prostaglandin D2 (PGD2), and the EP2 receptor selective agonist, butaprost. No increases in CGRP release were observed in response to prostaglandin F2alpha (PGF2alpha) or the TP receptor selective agonist, U46619, or the EP3 receptor selective agonist, GR63799X. 4. The selective DP receptor antagonist, BWA868C, antagonized the PGD2-, but not PGE2- or cPGI2-induced release. Furthermore, the EP1 selective antagonist, ZM325802, failed to antagonize the PGE2-induced CGRP release from these cells. 5. These data indicate that activation of DP, EP and IP receptors can each cause CGRP release from trigeminal neurones, and suggest that the predominant EP receptor subtype involved may be the EP2 receptor. Together with evidence that the cyclo-oxygenase inhibitor, aspirin, particularly when administered intravenously is effective in treating acute migraine, these findings further suggest a role for prostaglandins in migraine pathophysiology.     

Interleukin-1beta-induced mucin production in human airway epithelium is mediated by cyclooxygenase-2, prostaglandin E2 receptors, and cyclic AMP-protein kinase A signaling

We reported recently that interleukin (IL)-1beta exposure resulted in a prolonged increase in MUC5AC mucin production in normal, well differentiated, human tracheobronchial epithelial (NHTBE) cell cultures, without significantly increasing MUC5AC mRNA (Am J Physiol 286:L320-L330, 2004). The goal of the present study was to elucidate the signaling pathways involved in IL-1beta-induced MUC5AC production. We found that IL-1beta increased cyclooxygenase-2 (COX-2) mRNA expression and prostaglandin (PG) E(2) production and that the COX-2 inhibitor celecoxib suppressed IL-1beta-induced MUC5AC production. Addition of exogenous PGE(2) to NHTBE cultures also increased MUC5AC production and IL-1beta-induced Muc5ac hypersecretion in tracheas from wild-type but not from COX-2-/- mice. NHTBE cells expressed all four E-prostanoid (EP) receptor subtypes and misoprostol, an EP2 and EP4 agonist, increased MUC5AC production, whereas sulprostone, an EP1 and EP3 agonist, did not. Furthermore, specific protein kinase A (PKA) inhibitors blocked IL-1beta and PGE(2)-induced MUC5AC production. However, neither inhibition of epidermal growth factor receptor (EGFR) activation with the tyrosine kinase inhibitor 4-(3-chloroanilino)-6,7-dimethoxyquinazoline HCl (AG-1478) or EGFR blocking antibody nor inhibition of extracellular signal-regulated kinase/P-38 mitogen activated protein kinases with specific inhibitors blocked IL-1beta stimulation of MUC5AC mucin production. We also observed that tumor necrosis factor (TNF)-alpha, platelet activating factor (PAF), and lipopolysaccharide (LPS) induced COX-2 and increased MUC5AC production that was blocked by celecoxib, suggesting a common signaling pathway of inflammatory mediator-induced MUC5AC production in NHTBE cells. We conclude that the induction of MUC5AC by IL-1beta, TNF-alpha, PAF, and LPS involves COX-2- generated PGE(2), activation of EP2 and/or EP4 receptor(s), and cAMP-PKA-mediated signaling.     

Characterization of the prostaglandin E2 sensitive (EP)-receptor in the rat isolated trachea.

1. Using a range of natural and synthetic prostanoid receptor agonists and antagonists, we have shown that the rat isolated trachea contains a heterogeneous population of prostaglandin receptor sub-types mediating both relaxation and contraction of the smooth muscle. Prostaglandin E2 (PGE2) elicits smooth muscle relaxation of pre-contracted preparations, the responses being well defined, with a mean potency (p[A50]) of 7.81 +/- 0.05. 2. 11-deoxy PGE1 16,16-dimethyl PGE2 and misoprostol were all full agonists at this receptor, whilst AH13205 was a low potency agonist, and sulprostone was inactive. 3. The EP1 receptor antagonist, AH6809 (5 microM), and the selective DP receptor antagonist, BW A868C (0.1 microM), had no significant effect on the concentration-effect (E/[A]) curves to PGE2. 4. The putative EP4-receptor antagonist, AH23848B, produced non-competitive antagonism of the PGE2 response curves; pA2 values of 5.07 +/- 0.15 and 5.24 +/- 0.19 were obtained at concentrations of 30 microM and 100 microM respectively. 5. The synthetic thromboxane A2 mimetic, U46619, caused smooth muscle contractions, with a mean p[A50] of 6.90 +/- 0.11. This response was antagonized by the TP receptor antagonist, GR32191B, yielding a mean pA2 of 8.31. 6. At concentrations of 1 microM and above, prostaglandin D2 (PGD2) and the IP-receptor agonist, cicaprost, generally elicited concentration-dependent relaxations of the rat trachea. Prostaglandin F2 alpha (PGF2 alpha) was without affinity or efficacy. 7. These data suggest that the rat isolated trachea contains EP-receptors, TP-receptors, and few, if any DP, IP or FP-receptors.(ABSTRACT TRUNCATED AT 250 WORDS)     

Partial agonism of taprostene at prostanoid IP receptors in vascular preparations from guinea-pig, rat, and mouse

This study investigates whether incomplete relaxation of vascular smooth muscle preparations induced by the prostacyclin analogue taprostene is due to partial agonism at prostanoid IP receptors. In the presence of the prostanoid EP4 receptor antagonist AH 23848, 3 microM taprostene induced 45% relaxation of phenylephrine-contracted guinea-pig saphenous vein rings and displaced log concentration-response curves for the prostacyclin analogues AFP-07, TEI-9063, and cicaprost to the right, parallel to their predicted addition curves. In contrast, taprostene interacted additively with prostaglandin E2 (PGE2), ONO-AE1-259 (selective EP2 agonist), and acetylcholine. Similarly, on rat tail artery contracted with phenylephrine, 3 microM taprostene (20% relaxation) opposed AFP-07- but not PGE2-induced relaxation. However, under U-46619-induced tone (AH 23848 absent), taprostene antagonized AFP-07 and cicaprost more than TEI-9063, suggesting that the latter has more than one relaxation mechanism. The presence of a sensitive EP3 contractile system in mouse aorta interfered with IP receptor-mediated relaxation. By generating tone with phenylephrine and the potent EP3 agonist sulprostone, it was possible to show that 3 microM taprostene (15% relaxation) selectively opposed relaxations induced by AFP-07, TEI-9063, and cicaprost. Our experiments indicate that taprostene is a partial agonist at prostanoid IP receptors, and may be a lead to an IP receptor antagonist.     

In vitro characterization of prostanoid EP-receptors in the non-pregnant human myometrium.

1. Prostaglandin receptors of the PGE type have been characterized in the non-pregnant human myometrium in vitro according to the scheme of Coleman et al. (1984) by use of the agonists PGE2, sulprostone, rioprostil, AY23626, butaprost, misoprostol, 16,16-dimethylprostaglandin E2, enprostil and iloprost, and, the antagonist AH6809. 2. All prostanoids tested were active in non-pregnant human myometrium either as stimulators and/or inhibitors of spontaneous activity or both. Biphasic responses to PGE2 indicate that at least two receptor types of the EP-receptor exist, one mediating relaxation and the other mediating contraction. 3. Further evidence for the EP-receptor mediating excitation and relaxation was provided by the action of the EP2-/EP3-receptor selective prostanoids rioprostil, AY23626 and misoprostol, and the EP1-/EP2-receptor selective agonist 16,16-dimethylprostaglandin E2. 4. Butaprost, an EP2-receptor selective agonist, produced potent inhibition of spontaneous activity in the tissue which was generally longer-lasting than that evoked by the natural prostanoid PGE2. 5. The EP1-/EP3-receptor selective agonist sulprostone and the EP3-receptor agonist enprostil produced potent contractile responses supporting the presence of contractile EP3-receptors in the non-pregnant human myometrium in vitro. 6. The EP1-/IP-receptor selective agonist, iloprost, produced mixed responses in non-pregnant human myometrium. The contractile response was inhibited by the EP1-receptor antagonist AH6809. However, responses to the EP1-/EP3-receptor selective agonist sulprostone were unaffected by AH6809 which may indicate that only a small population of EP1-receptors is present. 7. Therefore it would seem that a heterogeneous population of EP-receptors is present in the non-pregnant human myometrium.     

Interleukin-1beta induces MUC2 and MUC5AC synthesis through cyclooxygenase-2 in NCI-H292 cells

Interleukin-1beta (IL-1beta) has been implicated in the pathogenesis of inflammatory diseases of the airway. In this study, we investigated the regulation of MUC2 and MUC5AC expression and of their regulatory mechanisms through cyclooxygenase-2 (COX-2) and prostaglandin E(2) (PGE(2)). Cells activated by IL-1beta showed increased COX-2, MUC2, and MUC5AC expressions at both the mRNA and protein levels. Mucin production was blocked by the selective COX-2 inhibitor NS398, and PGE(2) directly induced MUC2 and MUC5AC expression at both the mRNA and protein levels in a dose-dependent manner. These results suggest a role for PGE(2) in IL-1beta-induced mucin synthesis in NCI-H292 cells. To investigate the roles of molecules upstream of COX-2 in mucin regulation, we examined the role of mitogen-activated protein kinases (MAPKs). Cells activated by IL-1beta showed increased extracellular signal-regulated kinase (ERK)1/2 and p38 phosphorylation, and IL-1beta-induced MUC2 and MUC5AC production was blocked by the ERK pathway inhibitor PD98059 or the p38 inhibitor SB203580. The inhibition of both MAPKs reduced IL-1beta-induced COX-2 expression and PGE(2) synthesis. Furthermore, the addition of PGE(2) to cells overcame the inhibitory effects of both MAPK inhibitors in IL-1beta-induced mucin production. These results indicate that in human pulmonary epithelial cells, IL-1beta activates ERK or p38 to induce COX-2 production, which in turn induces MUC2 and MUC5AC production.     

Effects of the selective EP4 antagonist, CJ-023,423 on chronic inflammation and bone destruction in rat adjuvant-induced arthritis

Prostaglandin E2 (PGE2) produced by cyclooxygenase (COX) is a potent pro-inflammatory mediator. We have recently discovered CJ-023,423, a highly selective antagonist of EP4 receptors, one of the PGE2 receptors. This agent is suitable for exploring the effects of blocking EP4 receptors following oral administration in rats. In this study, CJ-023,423 was used in rats with adjuvant-induced arthritis (AIA) to investigate the role of the EP4 receptor in chronic inflammation and bone destruction. These effects were compared with those of rofecoxib, a selective COX-2 inhibitor. CJ-023,423 had significant inhibitory effects on paw swelling, inflammatory biomarkers, synovial inflammation and bone destruction in AIA rats. In particular, the inhibitory effect on paw swelling in AIA rats was comparable to that of rofecoxib. These results suggest that PGE2 acting via the EP4 receptor is involved in the development of chronic inflammation and bone destruction, particularly with respect to oedema in AIA rats. This is the first study to confirm the in-vivo effects of EP4 receptor blockade on inflammation and bone destruction in AIA rats with a small-molecule compound.     

Prostanoid receptors involved in the relaxation of human bronchial preparations

1. Iloprost and cicaprost (IP-receptor agonists) induced relaxations in the histamine- (50 microM) contracted human bronchial preparations (pD2 values, 6.63+/-0.12 and 6.86+/-0.08; Emax values, 90+/-04 and 65+/-08% of the papaverine response for iloprost (n=6) and cicaprost (n=3), respectively). 2. Prostaglandin E2 (PGE2) and misoprostol (EP-receptor agonist) relaxed the histamine-contracted human bronchial preparations (pD2 values, 7.13+/-0.07 and 6.33+/-0.28; Emax values, 67+/-04 and 57+/-08% of the papaverine response for PGE2 (n=14) and misoprostol (n=4), respectively). In addition, both relaxations were inhibited by AH6809 (DP/EP1/EP2-receptor antagonist; 3 microM; n=5-6). 3. The PGE2-induced relaxations of human bronchial preparations were not modified by treatment with AH23848B (TP/EP4-receptor antagonist; 30 microM; n=4). 4. The contracted human bronchial preparations were significantly relaxed by prostaglandin D2 (PGD2) or by BW245C a DP-receptor agonist. However, these responses did not exceed 40% of the relaxation induced by papaverine. In addition, the relaxations induced by PGD2 were significantly inhibited by treatment with a DP-receptor antagonist BWA868C (0.1 microM; n=3). 5. These data suggest that the relaxation of human isolated bronchial preparations induced by prostanoids involved IP-, EP2- and to a lesser extent DP-receptors but not EP4-receptor.     

Elevation of plasma noradrenaline levels in urethane-anaesthetized rats by activation of central prostanoid EP3 receptors.

1. We studied the effects of intracerebroventricular (i.c.v.) administration of prostaglandin E2 (PGE2) and its receptor subtype ligands on plasma levels of catecholamines in urethane-anaesthetized rats. 2. Administration of PGE2 (0.15, 0.3 and 1.5 nmol per animal, i.c.v.) dose-dependently elevated plasma levels of noradrenaline (NA), while the levels of adrenaline were not affected. 3. Administration of sulprostone (EP3/EP1 agonist) and misoprostol (EP3/EP2 agonist) effectively elevated plasma NA levels in a dose-dependent manner (0.1, 0.3, and 1.0 nmol per animal). Butaprost (EP2 agonist) (0.3, 1.0 and 3.0 nmol per animal) was without effect. 17-Phenyl-omega-trinor PGE2 (EP1/EP3 agonist) effectively elevated plasma NA levels only at its highest dose (1.0 nmol per animal), but this elevation was not attenuated by pretreatment with SC-19220 (selective EP1 antagonist) (20 nmol per animal, i.c.v.). 4. The potency of these test agents in elevating plasma levels of NA was as follows; misoprostol > sulprostone > PGE2 > > 17-phenyl-omega-trinor PGE2 > > > butaprost. These results suggest that activation of central prostanoid EP3-receptors induces central sympathetic outflow in rats.     

Role of PGE(2) in protease-activated receptor-1, -2 and -4 mediated relaxation in the mouse isolated trachea

1. The potential mediator role of the prostanoid PGE(2) in airway smooth muscle relaxations induced by peptidic and proteolytic activators of PAR-1, PAR-2, PAR-3 and PAR-4 was investigated in carbachol-precontracted mouse isolated tracheal segments. 2. The tethered ligand domain sequences of murine PAR-1 (SFFLRN-NH(2)), PAR-2 (SLIGRL-NH(2)) and PAR-4 (GYPGKF-NH(2)), but not PAR-3 (SFNGGP-NH(2)), induced smooth muscle relaxation that was abolished by the non-selective cyclo-oxygenase (COX) inhibitor, indomethacin. The relative order for mean peak relaxation was SLIGRL-NH(2)>GYPGKF-NH(2) approximately amp; SFFLRN-NH(2)>SFNGGP-NH(2). 3. SFFLRN-NH(2), SLIGRL-NH(2) and GYPGKF-NH(2), but not SFNGGP-NH(2), induced significant PGE(2) release that was abolished by indomethacin. Like that for relaxation, the relative order for mean PGE(2) release was SLIGRL-NH(2)>GYPGKF-NH(2)>SFFLRN-NH(2)>SFNGGP-NH(2). 4. In dose-response studies, SLIGRL-NH(2) induced concentration-dependent increases in PGE(2) release (EC(50)=20.4 microM) and smooth muscle relaxation (EC(50)=15.8 microM). 5. The selective COX-2 inhibitor, nimesulide, but not the COX-1 inhibitor valeryl salicylate, significantly attenuated SLIGRL-NH(2)-induced smooth muscle relaxation and PGE(2) release. 6. Exogenously applied PGE(2) induced potent smooth muscle relaxation (EC(50)=60.3 nM) that was inhibited by the mixed DP/EP(1)/EP(2) prostanoid receptor antagonist, AH6809. SLIGRL-NH(2)-induced relaxation was also significantly inhibited by AH6809. 7. In summary, the results of this study strongly suggest that PAR-mediated relaxation in murine tracheal smooth muscle is dependent on the generation of the spasmolytic prostanoid, PGE(2). PAR-stimulated PGE(2) release appears to be generated preferentially by COX-2 rather than COX-1, and induces relaxation via activation of the EP(2) receptor.     

Prostaglandin E2 activates EP2 receptors to inhibit human lung mast cell degranulation

The prostanoid, PGE2, is known to inhibit human lung mast cell activity. The aim of the present study was to characterize the EP receptor that mediates this effect. PGE2 (pEC(50), 5.8+/-0.1) inhibited the IgE-mediated release of histamine from mast cells in a concentration-dependent manner. Alternative EP receptor agonists were studied. The EP2-selective agonist, butaprost (pEC50, 5.2+/-0.2), was an effective inhibitor of mediator release whereas the EP1/EP3 receptor agonist, sulprostone, and the EP1-selective agonist, 17-phenyl-trinor-PGE2, were ineffective. The DP agonist PGD2, the FP agonist PGF(2alpha), the IP agonist iloprost and the TP agonist U-46619 were ineffective inhibitors of IgE-mediated histamine release from mast cells. PGE2 induced a concentration-dependent increase in intracellular cAMP levels in mast cells. The effects of the EP1/EP2 receptor antagonist, AH6809, and the EP4 receptor antagonist, AH23848, on the PGE2-mediated inhibition of histamine release were determined. AH6809 (pK(B), 5.6+/-0.1) caused a modest rightward shift in the PGE2 concentration-response curve, whereas AH23848 was ineffective. Long-term (24 h) incubation of mast cells with either PGE2 or butaprost (EP2 agonist), but not sulprostone (EP1/EP3 agonist), caused a significant reduction in the subsequent ability of PGE2 to inhibit histamine release. Collectively, these data suggest that PGE2 mediates effects on human lung mast cells by interacting with EP2 receptors.     

Characterization of receptors involved in the direct and indirect actions of prostaglandins E and I on the guinea-pig ileum.

1. A study of the effects of prostaglandin E2 (PGE2) and eleven synthetic analogues on the guinea-pig isolated ileum preparation has revealed three distinct contractile actions, each associated with a different prostaglandin E (EP-) receptor subtype. In addition, PGI2 (prostacyclin) and its stable analogues can activate prostaglandin I (IP-) receptors to elicit both contraction and relaxation of the ileum. 2. Two of the PGE actions involve direct stimulation of the smooth muscle, being unaffected by 1 microM morphine treatment. One action is blocked by AH 6809 at micromolar concentrations and ICI 80205 and 16,16-dimethyl PGE2 are particularly potent agonists. Activation of EP1-receptors appears to be involved. The second action is unaffected by AH 6809; sulprostone and MB 28767 are potent agonists. Comparison with agonist potency rankings on the guinea-pig vas deferens indicates that EP3-receptors may be involved. 3. The third PGE effect and the stimulant PGI effect are blocked by morphine, indicating enteric neurones and/or sensory nerve terminals as sites of action. EP2-receptors may be involved in the PGE action, in view of the marked effect of morphine on the contractile actions of misoprostol, 11-deoxy PGE2-1-alcohol, 11-deoxy PGE1 and butaprost, all of which show some selectivity for EP2-receptors. The PGI action is most easily studied with cicaprost (EC25 = 1.3 nM), since iloprost, carbacyclin and to a lesser extent PGI2 also have agonist activity at EP1-receptors. 4. The contractile action of 17-phenyl-omega-trinor PGE2 on the ileum is unaffected by morphine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Potentiation of aggregation and inhibition of adenylate cyclase in human platelets by prostaglandin E analogues.

1. The 16-phenoxy prostaglandin E analogue sulprostone consistently potentiates primary aggregation waves induced by adenosine 5'-diphosphate (ADP), PAF and 11,9-epoxymethano PGH2 (U-46619) in platelet-rich plasma from human donors. The effect is not blocked by the TP-receptor antagonists, EP 092 and GR 32191. The high potency of sulprostone (threshold concentration = 4-10 nM) and the weak block of sulprostone potentiation by the EP1-receptor antagonist, AH 6809 (pA2 = 4.3) suggest the involvement of EP3-receptors as opposed to EP1- or EP2-subtypes. 2. Eight prostaglandin E (PGE) analogues were compared against sulprostone for their effects on PAF-induced aggregation in human platelet-rich plasma (PRP) in the presence of GR 32191 and the DP-receptor antagonist, BW A868C. PGE2 and 11-deoxy PGE2-1-alcohol showed evidence of both potentiating and inhibitory actions and butaprost showed only inhibitory activity at high concentrations. The remaining analogues always elicited potentiation, with the following potency ranking: sulprostone = 16,16-dimethyl PGE2 > MB 28767 > misoprostol > GR 63779X = 17-phenyl-omega-trinor PGE2. The results again indicate that EP3- rather than EP1- or EP2-receptors are involved. However, relative potentiating potency could be affected by differences in plasma protein binding and the very high sensitivity of the human platelet to prostacyclin (IP)-receptor-mediated inhibition (IC50 for the specific IP-receptor agonist cicaprost = 0.8 nM).(ABSTRACT TRUNCATED AT 250 WORDS)     

Ontogenic increase in PGE2 and PGF2 alpha receptor density in brain microvessels of pigs.

1. The hypothesis that the relative vasoconstrictor ineffectiveness of prostaglandin E2 (PGE2) and PGF2 alpha on cerebral vessels of newborn pigs might be due to fewer receptors for these prostanoids was tested by comparing receptors for PGE2 (EP) and PGF2 alpha (FP) in cerebral microvessels from newborn and adult pigs. 2. Specific binding of [3H]-PGE2 and [3H]-PGF2 alpha to membranes prepared from brain microvessels showed that EP and FP receptor density (Bmax) in tissues from newborn animals was less than 50% of that determined in tissues from adults. By contrast, estimates of affinity (KD) were unchanged. 3. Specifically bound [3H]-PGE2 to brain microvessels from both the newborn and adult was displaced by AH 6809 (EP1-selective antagonist) by 80-90%, and only by approximately 30-35% by both 11-deoxy PGE1 (EP2/EP3 agonist) and M&B 28,767 (EP3 agonist); butaprost (EP2 agonist) was completely ineffective. 4. PGE2, 17-phenyl trinor PGE2 (EP1 agonist), PGF2 alpha and fenprostalene (PGF2 alpha analogue) caused significantly less increase in inositol 1,4,5-triphosphate (IP3) in brain microvessels from the newborn than in those from adult pigs. The stimulation of IP3 by PGE2 and 17-phenyl trinor PGE2 was almost completely inhibited by the EP1 antagonist, AH 6809. 5. PGE2, 11-deoxy PGE1 and M&B 28,767 produced small reduction of adenosine 3':5'-cyclic monophosphate (cyclic AMP) production in adult vessels but no effect in newborn tissues. 6. The lower density of EP and FP receptors in microvessels of newborn pigs compared to adults may explain the reduced ability of PGE2 and PGF2 alpha to stimulate production of IP3 in tissues from newborn animals.(ABSTRACT TRUNCATED AT 250 WORDS)     

Prostanoid stimulation of anion secretion in guinea-pig gastric and ileal mucosa is mediated by different receptors.

1. The receptors that mediate stimulation of anion secretion by prostanoids in isolated preparations of guinea-pig gastric and ileal mucosa have been compared by use of selective prostanoid agonists and antagonists. 2. In gastric mucosa, the relative potency of agonists suggested that the control of anion secretion in this tissue was complex and may be mediated by EP2, EP3, and TP receptors. A role for TP receptors was confirmed with the TP-selective antagonist AH23848 which inhibited short circuit current responses to the TP receptor agonist U-46619 with a pA2 value of 8.44 but was without effect on responses to prostaglandin E2 (PGE2) or the EP selective agonist, sulprostone. 3. In ileal mucosa, the relative potency of agonists differed from that observed in gastric mucosa and was consistent with the view that anion secretion in this region of intestine was controlled by DP and EP2 receptors. 4. These studies suggest that anion secretion in gastric and ileal mucosa is controlled by different prostanoid receptor subtypes and so provide important information for the design of prostanoids which may protect gastric mucosa and that are free from side effects such as diarrhoea.     

Potent contractile actions of prostanoid EP3-receptor agonists on human isolated pulmonary artery.

1. In 13 of 15 experiments, prostaglandin E2 (PGE2) and sulprostone (a prostanoid EP1/EP3-receptor agonist) contracted isolated rings of human pulmonary artery at low concentrations (> or = 5 and > or = 0.5 nM respectively). Tissue was obtained from patients undergoing surgery mainly for carcinoma of the lung. Characterization of the receptors involved was complicated by loss of sensitivity to the contractile PGE action over the experimental period. In contrast, contractile responses to KCl, phenylephrine and the specific thromboxane (TP-) receptor agonist, U-46619, did not decrease with time. 2. The relative contractile potencies for seven PGE analogues, measured during the first few hours after setting up the preparations, were as follows: sulprostone > misoprostol = gemeprost > or = PGE2 > or = GR 63799X > 17-phenyl-omega-trinor PGE2 > or = 11-deoxy PGE1. This ranking indicates that an EP3-receptor is involved. 3. The contractile action of sulprostone was not blocked by the TP-receptor antagonists, EP 169 and GR 32191, and the EP1-receptor antagonist, AH 6809. 4. In two experiments, PGE2 (50 nM) reduced basal tone and sulprostone was a weak contractile agent. Phenylephrine-induced tone was also inhibited by PGE2 (EC50 = 5-20 nM), 11-deoxy PGE1 and butaprost (a selective EP2-receptor agonist); the latter prostanoids were about 2 and 4 times less potent than PGE2 respectively. Interactions with phenylephrine were different in experiments where PGE2 alone was contractile: PGE2 induced contraction superimposed on the phenylephrine response and 11-deoxy PGE1 induced either further contraction or had no effect.(ABSTRACT TRUNCATED AT 250 WORDS)