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Prostanoid EP3 receptors mediate the high-synergy contractile action of PGE2 analogues on rat femoral artery
Deletion of genes for prostanoid EP receptor subtypes in the mouse indicates that PGE2 causes peripheral vasoconstriction by activating EP3 receptors (Audoly et al., 1999; Zhang et al., 1999); the mechanism of this effect is of interest to us. Our previous studies on guinea-pig aorta showed that the EP3 agonist sulprostone induced direct contraction, which was little affected by nifedipine, and showed modest synergy with other vasoconstrictors ( Shum et al., 2003 ). We now report on the contractile activity of PGE2 and its analogues on rat femoral artery. Femoral arteries were removed from male Sprague-Dawley rats (250 - 300 g). Four ring preparations were set up in a wire myograph system in Krebs-Henseleit solution gassed with 95% O2 / 5%CO2 at 37 oC and containing 1 µM indomethacin. The basic protocol involved addition of the TP antagonist GR 32191 (1 µM) and the NOS inhibitor L-NAME (100 µM), followed by a priming dose of phenylephrine, U-46619 or K+, followed by cumulative addition of PGE2 analogue. Data were analyzed by one-factor ANOVA / Bonferroni post-hoc testing. The weak contractile activity of sulprostone (0.1 – 400 nM, ≤10% of tissue maximum) was dramatically enhanced when the preparation was primed with another contractile agent (30 – 300 nM phenylephrine, 3 – 10 nM U-46619, 10 –30 mM K+). Both the pEC50 and maximum response of sulprostone increased with increasing priming level and fitting the phenylephrine priming response - pEC50 data with a hyperbola gave a pEC50 asymptote of 9.42 (95% CL 9.30 – 9.57, n = 73). Similar profiles were obtained for U-46619 and K+ priming. PGE2 and 17-phenyl PGE2 (EP1 > EP3 agonist) were about 10 times less potent than sulprostone (EP3 > EP1 agonist). Nifedipine (100 nM) abolished contraction induced by phenylephrine and K+ alone and their synergistic interactions with sulprostone, but only partially inhibited contraction to U-46619 alone. In the presence of 300 nM nifedipine, there was no synergism between sulprostone and contraction induced by a higher concentration of U-46619 (10 – 30 nM). The novel EP3 receptor antagonist L-798106 (Juteau et al., 2001; Clarke et al., 2004) caused a parallel right-shift of the log concentration-response curve of sulprostone enhanced by 50 nM phenylephrine. The pEC50 values for control, DMSO vehicle, 0.2 and 1 µM L-798106 treatments were 9.01 ± 0.17, 8.96 ± 0.12, 8.22 ± 0.17 and 6.96 ± 0.12 (s.e. mean, n = 6) respectively; the L-798106 means were significantly different from the vehicle mean (P <0.01 and P <0.001 respectively). The corresponding pA2 values were 7.35 and 8.00, similar to values of 7.48 and 7.82 reported for EP3 antagonism by L-798106 in guinea-pig vas deferens and trachea respectively (Clarke et al., 2004). In contrast, the potent EP1 antagonist SC-51322 (Hallinan et al., 2001; Durocher et al., 2000) did not significantly affect (P >0.05) the synergistic interactions of sulprostone and 17-phenyl PGE2 with 100 nM phenylephrine; pEC50 values for control, ethanol vehicle and 1 µM SC-51322 treatments were 9.15 ± 0.21, 9.26 ± 0.23 and 9.12 ± 0.12 (n = 5) and 8.33 ± 0.18, 8.49 ± 0.17 and 8.20 ± 0.18 (n = 4) respectively. These findings indicate that prostanoid EP3 receptors mediate the high-synergy contractile action of PGE2 analogues on rat femoral artery.
Audoly LP et al. (1999). Am J Physiol, 277, H924-H930. We thank GlaxoSmithKline UK for the gift of L-798106. |
