Analysis of sphingosine 1-phosphate receptors involved in constriction of isolated cerebral arteries with receptor null mice and pharmacological tools - PubMed (original) (raw)

Analysis of sphingosine 1-phosphate receptors involved in constriction of isolated cerebral arteries with receptor null mice and pharmacological tools

S Salomone et al. Br J Pharmacol. 2008 Jan.

Abstract

Background and purpose: Sphingosine 1-phosphate (S1P) selectively and potently constricts isolated cerebral arteries, but this response has not been pharmacologically characterized.

Experimental approach: The receptor subtype(s) involved in S1P-induced cerebrovascular constriction were characterized using genetic (S1P(2) and S1P(3) receptor null mice) and pharmacological tools (phospho-FTY720, a S1P(1/3/4/5) receptor agonist; SEW2871, a S1P(1) receptor agonist, JTE-013, a S1P(2) receptor antagonist, VPC23019, a S1P(1/3) receptor antagonist). Isolated basilar or peripheral (femoral, mesenteric resistance) arteries, from either rat or mouse, were studied in a wire myograph.

Key results: S1P concentration-dependently constricted basilar artery in rat, wild-type (WT) and S1P(2) null mice, but barely affected vascular tone in S1P(3) null mice. Vasoconstriction to U46619 (a thromboxane analogue) or to endothelin-1 did not differ between WT, S1P(2) and S1P(3) null mice. JTE-013 inhibited not only S1P-induced vasoconstriction, but also KCl-, U46619- and endothelin-1-induced constriction. This effect was observed in WT as well as in S1P(2) null mice. VPC23019 increased the concentration-dependent vasoconstriction to S1P in both rat and mouse basilar arteries with intact endothelium, but not in rat basilar artery without endothelium. Phospho-FTY720 concentration-dependently constricted rat basilar arteries, but not femoral or mesenteric resistance arteries, while SEW2871 did not induce any response in the same arteries.

Conclusions and implications: S1P constricts cerebral arteries through S1P(3) receptors. The purported S1P(2) receptor antagonist JTE-013 does not appear to be selective, at least in rodents. Enhancement of S1P-induced contraction by VPC23019 might be related to blockade of S1P(1) receptors and NO generation.

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Figures

Figure 1

Figure 1

Vasoconstriction to sphingosine 1-phosphate (S1P) in isolated basilar artery from wild-type (WT) or S1P2 or S1P3 receptor null mice. (a) Typical recording showing the contractile responses to cumulative concentrations of S1P in basilar artery isolated from WT mice; arrows indicate the addition of each concentration of S1P, from 10−8 to 3 × 10−5

M

, in half-log steps, to the organ bath; contractile response to high (100 m

M

) K+-induced depolarization is shown for comparison. (b) Effect of the S1P solvent (transient vasoconstriction, when injecting 50 and 150 μl) in basilar artery isolated from WT mice. (c) Typical recording showing the contractile response to S1P in basilar artery isolated from S1P3−/− mice. (d) Averaged contractile responses to cumulative concentrations of S1P in WT, S1P2−/−, S1P3−/− and S1P3+/− mice, expressed in % of high (100 m

M

) K+-induced contractions in the same preparations: mean and s.e.mean. The corresponding pharmacological parameters are given in Table 1.

Figure 2

Figure 2

Effect of JTE-013 on the responsiveness of mouse basilar artery to sphingosine 1-phosphate (S1P) and other agonists. (a–c) Averaged contractile responses to cumulative concentrations of U46619, S1P or endothelin-1 (ET-1) in basilar arteries from wild-type (WT) mice, in the presence or absence of 10 μ

M

JTE-013. Contractile responses are expressed in % of high (100 m

M

) K+-induced contractions in the same preparations in the absence of JTE-013. Values are mean and s.e.mean; _n_=5. (d, e) Typical recordings (experiment repeated twice with similar results) showing the contractile responses to cumulative concentrations of U46619 and S1P, in the absence (d) or presence of 10 μ

M

JTE-013 (e), in basilar artery isolated from S1P2−/− mice; arrows indicate the addition of each concentration of U46619 (10−8 to 3 × 10−6

M

) or S1P (10−8 to 3 × 10−5

M

), in half-log steps, to the organ bath; contractile responses to high (100 m

M

) K+-induced depolarization in the same preparations are shown for comparison. When used, JTE-013 was preincubated for 30 min before challenging with U46619, S1P or ET-1.

Figure 3

Figure 3

Effect of VPC23019 on the responsiveness of basilar artery from rat (a) and mouse (b) to sphingosine 1-phosphate (S1P). Contractile responses to cumulative concentrations of S1P, expressed in % of high (100 m

M

) K+-induced contractions in the same preparations, are shown as mean and s.e.mean (_n_=7–9). Isolated basilar arteries were incubated for 30 min with VPC23019 (10 μ

M

), vehicle (5 μl dimethyl sulphoxide; final concentration: 0.1%) or 0.1 m

M

_N_-ω-nitro-

L

-arginine methyl ester hydrochloride (

L

-NAME) before being challenged with cumulative concentrations of S1P. Nonlinear fits for each individual data set were compared by F-test.

Figure 4

Figure 4

Vasoconstriction to phospho-FTY720 in isolated rat basilar artery. (a) Typical recording showing the contractile responses to cumulative concentrations of phospho-FTY720; arrows indicate the addition of each concentration of phospho-FTY720, from 10−8 to 10−5

M

, in half-log steps, to the organ bath; the contractile response to 5-hydroxytryptamine (5-HT, 1 μ

M

) is shown for comparison. (b) Averaged contractile responses to cumulative concentrations of phospho-FTY720: mean and s.e.mean (_n_=12). The corresponding pharmacological parameters are given in the text.

Figure 5

Figure 5

Absence of vasoconstriction (a, b) or vasodilatation (c–e) to SEW2871 in different arteries isolated from rat. Typical recordings show the lack of effect of SEW2871, either as single concentration or as cumulative concentrations (by half-log steps), in comparison with positive controls for vasoconstriction, such as high (100 m

M

) K+-induced depolarization and 5-hydroxytryptamine (5-HT, 1 μ

M

) in basilar artery (a) and phenylephrine (PE, 10 μ

M

) in mesenteric resistance artery (b). For endothelium-dependent vasodilatation, acetylcholine (ACh; 1 μ

M

) was used as a positive control in basilar (c), femoral (d) and mesenteric resistance arteries (e). Each preparation/condition was tested 3–4 times with similar results.

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