DP Receptor Antagonist BW A868C Research Articles

Prostaglandin D 2 mediates neuronal damage by amyloid-β or prions which activates microglial cells

Microglial cells killed neurons damaged following incubation with sub-lethal concentrations of peptides derived from either the human prion protein (HuPrP82-146) or amyloid-β 1–42 (a peptide found in Alzheimer's disease). HuPrP82-146 or amyloid-β 1–42 induced phenotypic changes in neurons that caused them to bind a CD14-IgG chimera. In co-cultures microglial cells produced interleukin (IL)-6 in response to HuPrP82-146 or amyloid-β 1–42 damaged neurons. The binding of the CD14-IgG chimera to HuPrP82-146 or amyloid-β 1–42 damaged neurons was reduced by pre-treatment with cyclo-oxygenase (COX)-1 inhibitors and in co-cultures, COX-1 inhibitors significantly increased neuronal survival. Studies with individual prostaglandins demonstrated that the addition of prostaglandin D 2, or prostaglandin E 2, but not other prostaglandins (F 2α, H 2, I 2 or 15-dJ 2), mimicked the effects of amyloid-β 1–42 on neurons. Thus, prostaglandin D 2 or E 2 damaged neurons bound the CD14-IgG chimera, and in co-cultures prostaglandin D 2 damaged neurons activated microglial cells. These effects were mediated via the DP prostanoid receptor; DP receptor agonists BW245C or SQ27986 induced neuronal damage, while the DP receptor antagonist BWA868C was neuroprotective in co-cultures. These results indicate that prostaglandin D 2, produced following activation of COX-1 by sub-lethal concentrations of HuPrP82-146 or amyloid-β 1–42, causes phenotypic changes in neurons that activates microglial cells and leads to neuronal loss.

Piglet saphenous vein contains multiple relaxatory prostanoid receptors: evidence for EP4, EP2, DP and IP receptor subtypes

Prostaglandin E(2) produced endothelium-independent relaxation of phenylephrine- and 5-HT-contracted piglet saphenous vein (PSV; pEC(50)=8.6+/-0.2; n=6). The prostanoid EP(4) receptor antagonist GW627368X (30-300 nM) produced parallel rightward displacement of PGE(2) concentration-effect (E/[A]) curves (pK(b)=9.2+/-0.2; slope=1). Higher concentrations of GW627368X did not produce further rightward shifts, revealing the presence of non-EP(4) prostanoid receptors. In all, 18 other prostanoid receptor agonists relaxed PSV in a concentration-related manner. Relative potencies of agonists most sensitive to 10 muM GW627368X (and therefore predominantly activating EP(4) receptors) correlated well with those at human recombinant EP(4) receptors in human embryonic kidney (HEK-293) cells (r(2)=0.74). In the presence of 10 microM GW627368X, the rank order of agonist relative potency matched that of the human recombinant EP(2) receptor in Chinese hamster ovary cells (r(2)=0.72). Iloprost, cicaprost and PGI(2) relaxed PSV maximally and were antagonised by 10 microM GW627368X, demonstrating that they were full EP(4) receptor agonists. Residual responses to these compounds in the presence of GW627368X suggested the presence of IP receptors.BW245C relaxed PSV maximally (pEC(50)=6.8+/-0.1). In the presence of 10 microM GW627368X, BW245C produced biphasic E/[A] curves (phase one pEC(50)=6.6; alpha=24%; phase two pEC(50)=5.1; alpha=112%). Phase two was antagonised by the DP receptor antagonist BW A868C (1 microM), demonstrating that BW245C is an agonist at DP and EP4 receptors. We conclude that PSV contains EP(4), EP(2), DP and IP receptors; IP receptor agonists are also porcine EP(4) receptor agonists.

Prostaglandin D2 is a potent chemoattractant for human eosinophils that acts via a novel DP receptor.

Prostaglandin D2 (PGD2) is released following exposure of asthmatics to allergen and acts via the adenylyl cyclase-coupled receptor for PGD2 (DP receptor). In this study, it is reported that human eosinophils possess this receptor, which would be expected to inhibit their activation. In contrast, it was found that prostaglandin D2 is a potent stimulator of eosinophil chemotaxis, actin polymerization, CD11b expression, and L-selectin shedding. These responses are specific for eosinophils, as neutrophils display little or no response to prostaglandin D2. They were not due to interaction with receptors for other prostanoids, as prostaglandins E2 and F(2alpha), U46619 (a thromboxane A2 analogue), and carbaprostacyclin (a prostacyclin analogue) displayed little or no activity. Furthermore, they were not shared by the selective DP receptor agonist BW245C and were not prevented by the selective DP receptor antagonist BWA868C, indicating that they were not mediated by DP receptors. In contrast, the prostaglandin D2 metabolite 13,14-dihydro-15-oxoprostaglandin D2 induced eosinophil activation but did not stimulate DP receptor-mediated adenosine 3',5'-cyclic monophosphate (cAMP) formation. These results indicate that in addition to the classic inhibitory DP1 receptor, eosinophils possess a second, novel DP2 receptor that is associated with PGD2-induced cell activation. These 2 receptors appear to interact to regulate eosinophil responses to PGD2, as blockade of DP1 receptor-mediated cAMP production by BWA868C resulted in enhanced DP2 receptor-mediated stimulation of CD11b expression. The balance between DP1 and DP2 receptors could determine the degree to which prostaglandin D2 can activate eosinophils and may play a role in eosinophil recruitment in asthma.

Role of the parasite-derived prostaglandin D2 in the inhibition of epidermal Langerhans cell migration during schistosomiasis infection.

Epidermal Langerhans cells (LCs) play a key role in immune defense mechanisms and in numerous immunological disorders. In this report, we show that percutaneous infection of C57BL/6 mice with the helminth parasite Schistosoma mansoni leads to the activation of LCs but, surprisingly, to their retention in the epidermis. Moreover, using an experimental model of LC migration induced by tumor necrosis factor (TNF)-alpha, we show that parasites transiently impair the departure of LCs from the epidermis and their subsequent accumulation as dendritic cells in the draining lymph nodes. The inhibitory effect is mediated by soluble lipophilic factors released by the parasites and not by host-derived antiinflammatory cytokines, such as interleukin-10. We find that prostaglandin (PG)D2, but not the other major eicosanoids produced by the parasites, specifically impedes the TNF-alpha-triggered migration of LCs through the adenylate cyclase-coupled PGD2 receptor (DP receptor). Moreover, the potent DP receptor antagonist BW A868C restores LC migration in infected mice. Finally, in a model of contact allergen-induced LC migration, we show that activation of the DP receptor not only inhibits LC emigration but also dramatically reduces the contact hypersensitivity responses after challenge. Taken together, we propose that the inhibition of LC migration could represent an additional stratagem for the schistosomes to escape the host immune system and that PGD2 may play a key role in the control of cutaneous immune responses.

Effects of some isoprostanes on the human umbilical artery in vitro.

1. Cumulative concentration-effect curves for the selective prostanoid TP receptor agonist U46619 and six isoprostanes were constructed in the human isolated umbilical artery. 2. All compounds except 8-iso-PGF3 alpha produced concentration-dependent contractions. The contractile response to the isoprostanes increased with each cumulative addition up to a point, after which subsequent addition reduced the contraction below the previous level. This 'downturn' in the concentration-effect curve did not occur with U46619. 3. The potencies of the compounds tested were as follows (pEC50 +/- s.e.mean): U46619, 6.7 +/- 0.2; 8-iso-PGE2, 6.5 +/- 0.1; 8-iso-PGF2 alpha, 5.8 +/- 0.2; 8-iso-PGE1, 5.4 +/- 0.1; 8-iso-PGF1 alpha, 5.0 +/- 0.1; 8-iso-PGF2 beta > 4.8; 8-iso-PGF3 alpha >> 4.8 (n = 4-17). Neither 8-iso-PGF2 beta nor 8-iso-PGF3 alpha at 44 microM had a significant effect on cumulative concentration-effect curves to U46619. 4. The selective TP receptor antagonist GR32191 (0.1 microM) caused rightward shifts in the concentration-effect curves to all the active compounds. pA2 values for GR32191 against U46619, 8-iso-PGE2, 8-iso-PGF2 alpha, 8-iso-PGE1 were 7.6 +/- 0.2, 9 +/- 1, 8.2 +/- 0.3 and 7.7 +/- 0.3, respectively (n = 4). 5. Neither N omega-nitro-L-arginine methyl ester (100 microM) nor the selective DP receptor antagonist BW A868C (50 nM) affected the complex concentration-effect curve to 8-iso-PGE2 (n = 3). 6. Stable contractions to U46619 (1-3 microM) were unaffected by anandamide at concentrations up to 60 microM.

Inhibitory prostanoid EP receptors in human non-pregnant myometrium

Prostanoid EP receptor agonists relaxed cloprostenol-stimulated contraction of human non-pregnant myometrium in vitro with pEC 50 values of ( n=4): prostaglandin E 2, 7.8±0.2>1-OH prostaglandin E 1, 7.2±0.3>misoprostol, 6.6±0.1>16,16-dimethyl prostaglandin E 2, 6.3±0.7>butaprost, 5.7±0.3>11-deoxy prostaglandin E 1, 5.5±0.2=AH13205 ((±)- trans-2-[4-(1hydroxyhexyl)phenyl]-5-oxocyclopentaneheptanoic acid), 5.5±0.2. The EP 4 receptor antagonist AH23848B ([1α(z), 2β5α]-(±)-7-[5-[[(1,1′-biphenyl)-4-yl]methoxy]-2-(4-morpholinyl)-3-oxo-cyclopentyl]-4-heptenoic acid) (29 μM) had no effect on concentration–effect curves to the EP receptor agonists. The mixed prostanoid receptor antagonist AH6809 (6-isopropoxy-9-oxaxanthene-2-carboxylic acid) competitively antagonised prostaglandin E 2 with a p A 2 of 5.6±0.2. AH6809 (42 μM) antagonised misoprostol, 11-deoxy prostaglandin E 1, and the prostanoid DP receptor agonist BW245C (5-(6-carboxyhexyl)-1-(3-cyclohexyl-3-hydroxypropyl)hydantoin) with apparent p A 2 values of 5.6±0.3, 5.1±0.9 and 5.9±0.4 ( n=4), respectively, but was ineffective against the IP receptor agonist cicaprost ( n=4). The prostanoid DP receptor antagonist BW A868C (3-benzyl-5-(6-carboxyhexyl)-1-(2-cyclohexyl-2-hydroxyethylamino)hydantoin) (50 nM) had no effect on responses to prostaglandin E 2 or misoprostol. The presence of an AH6809-sensitive, AH23848B- and BW A868C-insensitive mechanism is consistent with the hypothesis that inhibitory EP receptor agonists cause relaxation of human non-pregnant myometrium by an EP 2 receptor-mediated process.

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.

Inhibition of nociceptin-induced allodynia in conscious mice by prostaglandin D2.

1. We recently showed that intrathecal administration of nociceptin induced allodynia by innocuous tactile stimuli and hyperalgesia by noxious thermal stimuli in conscious mice. In the present study, we examined the effect of prostaglandins on nociceptin-induced allodynia and hyperalgesia. 2. Prostaglandin D2 (PGD2) blocked the allodynia induced by nociceptin in a dose-dependent manner with an IC50 of 26 ng kg(-1), but did not affect the nociceptin-induced hyperalgesia at doses up to 500 ng kg(-1). BW 245C (an agonist for PGD (DP) receptor) blocked the allodynia with an IC50 of 83 ng kg(-1). 3. The blockade of nociceptin-induced allodynia by PGD2 was reversed by the potent and selective DP-receptor antagonist BW A868C in a dose-dependent manner with an ED50 of 42.8 ng kg(-1). 4. Glycine (500 ng kg[-1]) almost completely blocked the nociceptin-induced allodynia. A synergistic effect on the inhibition of nociceptin-evoked allodynia was observed between glycine and PGD2 at below effective doses. 5. Dibutyryl cyclic AMP, but not dibutyryl cyclic GMP, blocked the nociceptin-induced allodynia with an IC50 of 2.9 microg kg(-1). 6. PGE2, PGF2alpha, butaprost (an EP2 agonist) and cicaprost (a PGI receptor agonist) did not affect the nociceptin-induced allodynia. 7. These results demonstrate that PGD2 inhibits the nociceptin-evoked allodynia through DP receptors in the spinal cord and that glycine may be involved in this inhibition.

Effects of BW A868C, a selective prostaglandin DP receptor antagonist, in dog isolated vascular preparations

The effects of the selective prostaglandin DP receptor antagonist BW A868C ((±)-3-benzyl-5-(6-carboxyhexyl)-1-(2-cyclohexyl-2-hydroxyethylamino)-hydantoin), 3.0 nM to 0.3 μM) were examined against prostaglandin D2 and BW245C ((±)-5-(6-carboxyhexyl)-1-(3-cyclohexyl-3-hydroxypropyl)-hydantoin)-induced smooth muscle relaxation on dog isolated vascular preparations pre-contracted with a sub-maximal concentration of KCl (50 mM). In dog dorsal nasal vein BW245C was found to be more potent than prostaglandin D2 with p[A]50 estimates of 7.6 ± 0.1 (S.E.M., n = 8) and 5.8 ± 0.1 (n = 5), respectively. BW A868C, up to 0.3 μM, displaced the relaxant concentration-effect curves to BW245C in dog dorsal nasal vein in an apparently competitive manner with parallel rightward shifts and no significant changes in the upper asymptotes. The data were analysed by using a modified Schild equation which not only gives equal weight to all agonist concentration-effect data but also allows a direct plot in Clark plot space. A pKB estimate of 7.3 ± 0.8 (n = 20) was obtained with a unity Schild plot slope (b = 1.0 ± 0.1). This affinity estimate, however, is lower than the values previously reported in other studies. The affinity estimates of BW A868C against BW245C and prostaglandin D2 obtained from dog dorsal nasal vein, major palatine artery and saphenous vein were found to be consistent. The relatively low affinity estimates of BW A868C at DP receptors as observed in the present study may be due to species- or tissue-related variations or may be indicative of the possible existence of DP receptor subtypes.

Agents That Elevate cAMP Inhibit Human Neutrophil Apoptosis

Neutrophil apoptosis, determined after 20 h in culture using standard criteria and shedding of cell surface CD16 (FcγRIII), is dramatically inhibited, in a concentration-dependent manner, by the cAMP analogs, dibutyryl-cAMP and 8-Br-cAMP, and the adenylyl cyclase activator, forskolin. Furthermore, the stable receptor-directed PGD2 mimetic, ZK 118,182, and the PGE2 mimetic, Il-deoxy PGE1, similarly inhibited apoptosis. The DP-receptor antagonist BW A868C blocked the effect of ZK 118,182 and the protein kinase A inhibitor H-89 reversed the inhibition of apoptosis induced by dibutyryl-cAMP. These results clearly show that neutrophil apoptosis is markedly attenuated by cAMP elevating agents. This nucleotide second messenger may play a fundamental role in controlling neutrophil longevity and pharmacological regulation of cAMP levels or actions may influence neutrophil apoptosis in vivo.

Effects of the isoprostane, 8-epi-prostaglandin F 2α, on the contractility of the human myometrium in vitro

8-epi-prostaglandin F 2α stimulated contraction of human myometrial strips obtained from five different donors at the time of hysterectomy with a pEC 50 value of 6.3 ± 0.5. In paired strips from the same donors the pEC 50 value for the selective TP receptor agonist U46619 ([-[1a,4a,5b( Z),6a(1 E,3 S∗)]]-7-[6-(3-hydroxy-1-octenyl)-2-oxabicyclo[2.2.1]hept-5-yl]-5-heptenoci acid) was 8.3 ± 0.4. In strips from four other donors 8-epi-prostaglandin F 2α was ineffective whereas the pEC 50 for U46619 was 6.9 ± 0.3. Responses to 8-epi-prostaglandin F 2α were unaffected by the selective DP receptor antagonist BW A868C (3-benzyl-5-(6-carboxyhexyl)-1-(2-cyclohexyl-2-hydroxyethylamino)hydantoin) at 50 nM but were blocked by the selective TP receptor antagonist L670596 ((−)6,8-difluoro-9- p-methylsulfonyl benzyl-1,2,3,4-tetrahydrocarbazol-1-yl-acetic acid) at 50 nM. The pIC 50 values obtained when the TP receptor antagonists GR 32191 ([1 R-[1α( Z),2β,3β,5α]]-(+)-7-[5-[[(1,1'-biphenyl)-4-yl]methoxy]-3-hydroxy-2-(1-piperidinyl)cyclopentyl]-4-heptenoic acid), ICI D1542 ((4( Z)-6-[(2 S,4 S,5 R)-2-[1-methyl-1-(2-nitro-4-tolyloxy)ethyl]-4-(3-pyridyl)-1,3-dioxan-5-yl]hex-4-enoci acid), ICI 192605 (4(Z)-6-[(2,4,5- cis)-2-(2-chlorophenyl)-4-(2-hydroxyphenyl)-1,3-dioxan-5-yl]hexenoic acid), L670596 and SQ 29548 ([1 S-(1α,2β(5Z),3β,4α]]-7-[3-[[2-[(phenylamino)carbonyl]hydrazino]methyl]-7-oxabicyclo[2.2.1]hept-2-yl]-5-heptenoic acid) were added cumulatively to strips pre-contracted with an EC 80 concentration of 8-epi-prostaglandin F 2α were not significantly different from those obtained when an EC 80 concentration of U46619 was used. The effects of 8-epi-prostaglandin F 2α on strips pre-contracted with an EC 80 concentration of U46619 were not different from those of U46619 itself. It is concluded that in the non-pregnant human myometrium 8-epi-prostaglandin F 2α is a medium potency contractile agonist acting predominantly at the TP receptor.

Functional characterization of the prostanoid DP receptor in human myometrium

Spontaneous contractile activity of strips of human myometrium obtained from non-pregnant donors at the time of hysterectomy was inhibited by the selective prostanoid DP receptor agonists BW 245C (5-(6-carboxyhexyl)-1-(3-cyclohexyl-3-hydroxypropyl)hydantoin) and ZK110841 ((5 Z,13 E)-(9 R,11 R,15 S)-9β-chlor-15-cyclohexyl-11,15-dihydroxy-16,17,18,19,20-pentanor-5,13-prostadienoic acid) with pEC 50 values of 8.4 and 7.3 respectively but prostaglandin D 2 produced a biphasic effect. In the presence of the TP receptor antagonist L670596 ((−)-6,8-difluoro-9- p-methylsulfonyl benzyl-1,2,3,4-tetrahydrocarbazol-1-yl-acetic acid), contractile activity induced by the FP receptor agonist, cloprostenol ([1 R-[1α( Z),2β(1 E,3 R∗),3α,5α]]-7-[2-[4-(3-chlorophenoxy-3-hydroxy-7-butenyl]-3,5-dihydroxycyclopentyl]-5-heptenoic acid), was inhibited by BW 245C ( pEC 50 = 7.5), ZK110841 ( pEC 50 = 6.7) and prostaglandin D 2 ( pEC 50 = 6.3). Under these conditions both prostaglandin J 2 and 9α,11β-prostaglandin F 2 were inhibitory partial agonists. All compounds were antagonized by the selective DP receptor antagonist BW A868C (3-benzyl-5-(6-carboxyhexyl)-1-(2-cyclohexyl-2-hydroxyethylamino)hydantoin), but the pK B values were both concentration-dependent ( pK B versus BW 245C at 10 nM = 9.1, at 50 nM = 8.3) and agonist-dependent ( pK B at 10 nM versus BW 245C = 9.1, versus ZK110841 = 7.4). Both agonist and antagonist potencies support the existence of DP receptors in human myometrium. The concentration and agonist dependence of the action of BW A868C suggests that putative DP receptor agonists relax human myometrium by more than one mechanism. These observations may be explained by the existence of subtypes of DP receptor in human myometrium.

Prostanoid receptors of the EP3 subtype mediate the inhibitory effect of prostaglandin E2 on noradrenaline release in the mouse brain cortex.

Mouse or rat brain cortex slices were preincubated with 3H-noradrenaline and superfused with physiological salt solution containing desipramine. We studied the effects of prostaglandin E2 (PGE2), prostaglandin D2 (PGD2) and related drugs on the electrically evoked (50 mA, 2 ms, 0.3 Hz) tritium overflow. PGE2 inhibited the electrically evoked tritium overflow from mouse brain cortex slices; the maximum effect of PGE2 (79%) was attenuated by the alpha 2-adrenoceptor agonist talipexole (to 52%) and enhanced by the alpha 2-adrenoceptor antagonist rauwolscine (to 92%). Rauwolscine was added to the superfusion medium in all subsequent experiments. The effect of PGE2 was readily reversible upon withdrawal from the medium and remained constant upon prolonged exposure of the tissue to the prostanoid. Studies with EP receptor agonists, mimicking the inhibitory effect of PGE2, showed the following potencies (pIC50); sulprostone (8.22); misoprostol (8.00); PGE2 (7.74); PGE1 (7.61); iloprost (5.86). The concentration-response curve of PGE2 was marginally shifted to the right by the EP1 receptor antagonist AH 6809 (6-isopropoxy-9-oxoxanthene-2- carboxylic acid; apparent pA2 3.97) and by the TP receptor antagonist vapiprost (4.50). AH 6809, by itself, did not affect the evoked overflow whereas vapiprost increased it. PGD2 inhibited the evoked overflow at high concentrations (pIC50 4.90); this effect was not altered by the DP receptor antagonist BW A868C (3-benzyl-5-(6-carboxyhexyl)-1-(2-cyclohexyl-2- hydroxyethylamino)hydantoin), which, by itself, did not affect the evoked overflow. Indometacin slightly increased the evoked overflow and tended to increase the inhibitory effect of PGE2. PGE2 inhibited the electrically evoked tritium overflow also in rat brain cortex slices.(ABSTRACT TRUNCATED AT 250 WORDS)

Identification of a single (FP) receptor associated with prostanoid-induced Ca 2+ signals in swiss 3T3 cells

Thus far, the prostanoid FP-receptor has been characterized only on the basis of agonist studies. It is currently classified as a receptor having particular sensitivity to prostaglandin F 2α (PGF 2α) but with the ability to recognize prostaglandins D 2 and E 2 (PGD 2 and PGE 2). We have re-examined this concept by studying second messenger Ca 2+ signals to PGF 2α, PGD 2 and PGE 2, and performing radioligand binding studies in Swiss 3T3 cells. The same rank order of potency was obtained for both the Ca 2+ transient signal and competition for PGF 2α binding sites. The potency rank order, PGF 2 α >PGD 2>PGE 2, was identical to that obtained from functional studies in isolated tissues, such as the cat iris. Additional support for the concept that PGF 2α, PGD 2, and PGE 2 interact with a single receptor to elicit a Ca 2+ signal was provided by successive addition studies. Thus, cells pretreated with a supramaximal concentration of PGF 2α exhibited little or no response to subsequent administration of PGD 2 or PGE 2. Likewise, cells pretreated with a large concentration PGD 2 or PGE 2 exhibited minimal responsiveness to successive addition of the corresponding alternative prostaglandins. Pretreatment with a maximally effective concentration of PGF 2α, PGD 2, or PGE 2 rendered the cells refractory to the FP-receptor selective agonist fluprostenol, which further supports the hypothesis that Ca 2+ transient signals in response to prostanoids in Swiss 3T3 cells are mediated by the FP-receptor. The Ca 2+ transient responses to PGF 2α, PGD 2, and PGE 2 also exhibited a similar modest reduction when extracellular Ca 2+ was removed. Finally, the DP-receptor antagonist BW A868C did not block the Ca 2+ transient response to PGD 2, indicating an absence of DP-receptor involvement. Moreover, Ca 2+ responses to the thromboxane A 2 mimetic U-46619 were unaffected by the TP-antagonist BM 13505, which indicates no involvement of the TP-receptor. These results support the contention that the FP-receptor has particular sensitivity to PGF 2α but will also recognize PGD 2 and PGE 2.

Prostaglandin Receptors Coupled to Adenylyl Cyclase in the Iris-Ciliary Body of Rabbits, Cats and Cows

Stimulation of adenylyl cyclase by prostaglandin (PG) receptor agonists was examined in the iris-ciliary body of rabbits, cats and cows. Iris-ciliary bodies were incubated with or without agonists in Krebs pyruvate buffer containing protease and cyclooxygenase inhibitors. The formation of cAMP in a 15-min period in response to the agonists was measured. In the rabbit iris-ciliary body, all PG receptor agonists at concentrations of 0·062-10·0 μM with the exception of sulprostone stimulated adenylyl cyclase. In this species, 17-phenyl trinor PGE 2 and PGF 2α stimulated adenylyl cyclase but only at high concentrations (> 5 μM). The order of potency of the agonists was dimethyl PGE 2 > PGE 2 > 11-deoxy PGE 1 = PGD 2 > iloprost > PGF 2α. The DP receptor antagonist BWA868C had no effect on EP agonist responses, indicating that PGE 2, dimethyl PGE 2 and 11-deoxy PGE 1 were not stimulating DP receptors. In the cat, PGE 2, dimethyl PGE 2 and 11-deoxy PGE 1, were all equally potent in stimulating adenylyl cyclase. The order of potency in the cow was PGE 2 > dimethyl PGE 2 > 11-deoxy PGE 1. However, PGD 2, iloprost, 17-phenyl trinor PGE 2 and PGF 2α were inactive in this species. The results of this study indicate that EP receptor agonists stimulate adenylyl cyclase in the iris-ciliary body of the three species: stimulation by other agonists depends on the species. In the rabbit, the EP 2 receptor subtype appears to predominate; such a predominance of EP 2 receptors was not observed in the cat and cow.

In vitro characterization of prostanoid receptors on human myometrium at term pregnancy.

1. Prostanoid receptors present on the pregnant human myometrium in vitro have been characterized according to the receptor classification proposed by Coleman et al. (1984) using natural prostanoids and synthetic, selective analogues and antagonists where available. 2. Prostaglandin E2 (PGE2) produced a biphasic effect consisting of an initial excitation followed by a dose-related inhibition. The EP2/EP3-receptor agonists, rioprostil and misoprostol, produced similar effects to PGE2, however, the excitatory event of the misoprostol response was related to dose. The EP1/EP3-receptor agonist, sulprostone, evoked a purely excitatory response which was unaffected by AH6809. The selective EP2-receptor agonist butaprost produced a long-lasting dose-dependent inhibition of activity. The results from these prostanoids indicated that inhibitory EP2- and excitatory EP3-receptors are present on myometrium from pregnant donors at term. 3. PGF2 alpha and the synthetic FP-receptor agonist, fluprostenol, caused equipotent excitatory effects, indicating the presence of contractile FP-receptors. 4. PGD2 produced a biphasic effect of which the inhibition appeared dose-related and was antagonized by the selective DP-receptor antagonist BW A868C. The selective DP-receptor agonist, BW245C, produced a potent inhibitory effect that was competitively antagonized by BW A868C (pA2 = 8.6). 5. PGI2 produced a biphasic response qualitatively similar to PGE2. The EP1/IP-receptor agonist, iloprost, produced an occasional unquantifiable excitation and dose-related inhibition. The selective IP-receptor prostanoid, cicaprost, evoked only an inhibitory response. 6. The stable thromboxane A2 (TXA2)-mimetic, U46619, produced potent excitation which was competitively antagonized by the TP-receptor antagonist, GR32191 (pA2 = 7.2). 7. The prostanoids tested indicate that a heterogeneous population of prostanoid receptors are presen ton human myometrium from pregnant donors. It may be concluded that excitation is EP3-, FP- and TP-receptor-mediated and inhibition is EP2-, DP- and IP-receptor-mediated. Comparison of data obtained from non-pregnant specimens indicates that the lower segment tissue from pregnant donors demonstrated more pronounced responses to EP2 and IP-receptor activation.

Further studies on ocular responses to DP receptor stimulation

Prostaglandin D 2 (PGD 2) and the selective DP receptor agonist BW 245C have been previously shown to lower intraocular pressure in rabbits, while PGD 2, but not BW 245C, caused plasma extravasation, eosinophil infiltration, and goblet cell depletion. In these present studies definition of the ocular pharmacology of prostaglandin D 2 (PGD 2) has been extended by using a further selective DP receptor agonist SQ 27986 and a potent and selective DP receptor antagonist BW A868C. In cats and rabbits SQ 27986 caused ocular hypotension. The ocular hypotensive effect of PGD 2 in rabbits was blocked by pretreatment with the DP receptor antagonist BW A868C, whereas the activities of PGE 2 and PGF 2α remained unaltered. THe singular involvement of the DP receptor in changes in rabbit intraocular pressure evoked by PGD 2 was thereby verified by using the antagonist BW A868C. In terms of effects on the ocular surface, SQ 27986 caused no increase in conjunctival microvascular permeability, no eosinophil infiltration, and no depletion of the goblet cell population. These findings reinforce the concept that selective DP receptor agonists may be useful for lowering intraocular pressure without causing ocular surface pathology. PGD 2 induced increases in conjunctival microvascular permeability were inhibited by BW A868C, despite the fact that DP receptor agonists failed to evoke a plasma exudation response. This finding was unexpected and suggests a possible subdivision of the DP receptor designation.