EP4 prostanoid receptor-mediated vasodilatation of human middle cerebral arteries

Abstract

1. Dilatation of the cerebral vasculature is recognised to be involved in the pathophysiology of migraine. Furthermore, elevated levels of prostaglandin E(2) (PGE(2)) occur in the blood, plasma and saliva of migraineurs during an attack, suggestive of a contributory role. In the present study, we have characterised the prostanoid receptors involved in the relaxation and contraction of human middle cerebral arteries in vitro. 2. In the presence of indomethacin (3 microm) and the TP receptor antagonist GR32191 (1 microM), PGE(2) was found to relax phenylephrine precontracted cerebral arterial rings in a concentration-dependent manner (mean pEC(50) 8.0+/-0.1, n=5). 3. Establishment of a rank order of potency using the EP(4)>EP(2) agonist 11-deoxy PGE(1), and the EP(2)>EP(4) agonist PGE(1)-OH (mean pEC(50) of 7.6+/-0.1 (n=6) and 6.4+/-0.1 (n=4), respectively), suggested the presence of functional EP(4) receptors. Furthermore, the selective EP(2) receptor agonist butaprost at concentrations <1 microM failed to relax the tissues. 4. Blockade of EP(4) receptors with the EP(4) receptor antagonists AH23848 and EP(4)A caused significant rightward displacements in PGE(2) concentration-response curves, exhibiting pA(2) and pK(B) values of 5.7+/-0.1, n=3, and 8.4, n=3, respectively. 5. The IP receptor agonists iloprost and cicaprost relaxed phenylephrine precontracted cerebral arterial rings (mean pEC(50) values 8.3+/-0.1 (n=4) and 8.1+/-0.1 (n=9), respectively). In contrast, the DP and FP receptor agonists PGD(2) and PGF(2 alpha) failed to cause appreciable relaxation or contraction at concentrations of up to 30 microm. In the absence of phenylephrine contraction and GR32191, the TP receptor agonist U46619 caused concentration-dependent contraction of cerebral artery (mean pEC(50) 7.4+/-0.3, n=3). 6. These data demonstrate the presence of prostanoid EP(4) receptors mediating PGE(2) vasodilatation of human middle cerebral artery. IP receptors mediating relaxation and TP receptors mediating contraction were also functionally demonstrated.

Figure 1

Typical response to PGE₂ on phenylephrine precontracted human middle cerebral artery. The figure shows concentration-dependent relaxation in response to increasing concentrations of PGE₂ (0.1 nM–3 μM in half-log increments).

Figure 2

Characterisation of PGE₂ relaxatory response using a range of prostanoid ligands on human middle cerebral artery. (a) The EP₄>EP₂ agonist 11-deoxy PGE₁ and the EP₂>EP₄ agonist PGE₁-OH (1 nM–10 μM), caused relaxation of cerebral artery rings, with a rank order suggestive of the presence of functional EP₄ receptors. Butaprost was without significant effect at concentrations below 1 μM. (b) The IP receptor agonists iloprost and cicaprost (0.3 nM–10 μM) equipotently caused concentration-dependent relaxations of phenylephrine precontracted cerebral rings. PGD₂ (1 nM–100 μM) produced only weak relaxatory responses at high concentrations. Data are expressed as mean±s.e.m.% of phenylephrine contraction.

Figure 3

Effects of EP₄ receptor antagonists on PGE₂-mediated relaxation of phenylephrine pre-contracted human middle cerebral artery. Cerebral artery rings were preincubated for 60 min with (a) AH23848 (10 μM) or (b) different concentrations of EP₄A (30, 300, 1000 and 3000 nm), and then cumulatively concentration-dependently relaxed with PGE₂ in the presence of GR32191 (1 μM). Data are expressed as percentage of the phenylephrine contraction, and are given as mean±s.e.m. for n=3/4 donors. (c) Schild plot of the antagonist effect of EP₄A on responses to PGE₂ in human middle cerebral artery.