Characterization of tachykinin receptors mediating plasma extravasation and vasodilatation in normal and acutely inflamed knee joints of the rat
Authors: Lam F.Y. , Wong M.C.S. 
Published: July 2012

Inflammatory actions of tachykinins in normal rat knee joints were compared with those of animals with acutely inflamed joints induced by intra‐articular injection of 2% carrageenan. Plasma protein extravasation in rat knee joints, measured by protein micro‐turbidimetry, was induced by intra‐articular perfusion of selective tachykinin receptor agonists. Changes in joint blood flow, measured by laser Doppler perfusion imaging, were produced by topical applications of selective tachykinin receptor agonists to the joint capsule.

Carrageenan‐injected rat knee joints showed significantly higher (P < 0.001) basal plasma extravasation (56 ± 4 μg ml−1, n = 5) than normal rat knee joints (10 ± 4 μg ml−1, n = 6). Intra‐articular perfusion of the selective neurokinin1 (NK1) receptor agonist [Sar9, Met(O2)11]‐substance P (0.8 nmol min−1) for 60 min elevated the basal plasma extravasation to 90 ± 17 μg ml−1 (n = 6, P < 0.001) in normal joints, and to 150 ± 14 μg ml−1 (n = 5, P < 0.001) in inflamed joints. Perfusion of the selective NK1 receptor antagonist N2‐[(4R)‐4‐hydroxy‐1‐(1‐methyl‐1H‐indol‐3‐yl)carbonyl‐L‐prolyl]‐N‐methyl‐N‐phenylmethyl‐3‐(2‐naphthyl)‐L‐alaninamide (FK888; 0.8 nmol min−1) for 20 min followed by co‐perfusion with the NK1 receptor agonist (0.8 nmol min−1) produced complete inhibition of the NK1 receptor agonist‐induced plasma extravasation in the two groups of animals (for both groups; n = 3, P < 0.001).

Intra‐articular perfusion of the selective NK2 receptor agonist [Nle10]‐neurokinin A4–10 (0.8 nmol min−1) and the selective NK3 receptor agonist [MePhe7]‐neurokinin B (0.8 nmol min1) produced no increase in plasma extravasation in normal or in inflamed rat knee joints (n = 4 and 11, P > 0.05).

Topical bolus applications of the NK1 receptor agonist [Sar9, Met(O2)11]‐substance P onto normal joint capsules produced dose‐dependent vasodilatation expressed as a voltage increase from control level. The maximum increase in blood flow was 2.05 ± 0.21 V from a basal voltage of 3.42 ± 0.07 V (n = 13, P < 0.001). To a much lesser extent, administration of the NK2 receptor agonist [Nle10]‐neurokinin A4–10 also produced dose‐dependent vasodilatation with a maximum increase of 0.46 ± 0.08 V from a basal level of 3.38 ± 0.1 V (n = 7, P < 0.01). Animals with acutely inflamed joints showed enhanced vasodilator responses to the NK1 and NK2 receptor agonists (for both: P vs non‐inflamed joints < 0.001). Thus, the NK1 and NK2 receptor agonists produced maximum increases of 2.56 ± 0.19 V (basal level = 5.84 ± 0.07 V; n = 7, P < 0.001) and 1.97 ± 0.26 V (basal level = 6.31 ± 0.23 V; n = 11, P < 0.001), respectively. The NK3 receptor agonist [MePhe7]‐neurokinin B produced no change in blood flow in normal or in inflamed rat knee joints (n = 7 and 5, P > 0.05).

Bolus administration of the NK1 receptor antagonist FK888 (10 pmol) alone followed 5 min later by another dose of 10 pmol FK888 (i.e. total dose of 2 × 10 pmol) applied together with the NK1 receptor selective agonist [Sar9, Met(O2)11]‐substance P produced partial, but significant inhibition of the NK1 receptor agonist‐induced vasodilatation in both normal (maximum response reduced by 51.9 ± 5.4%; n = 6, P < 0.001) and inflamed rat knee joints (maximum response reduced by 49.3 ± 6.1%; n = 5, P < 0.001). The NK2 receptor agonist [Nle10]‐neurokinin A4–10‐induced vasodilator responses in inflamed joints were not affected by this treatment (n = 6, P > 0.05). However, with two higher doses of FK888 (both 1 nmol), the NK1 and the NK2 receptor agonist‐induced vasodilator responses were abolished in the two groups of animals (n = 6–8, P < 0.005).

Administration of two doses of the selective NK2 receptor antagonist (S)‐N‐methyl‐N‐[4‐acetylamino‐4‐phenylpiperidino)‐2‐(3,4‐dichlorophenyl)‐butyl]benzamide (SR48968; both 10 pmol) produced no change in NK1 receptor agonist‐induced vasodilatation in normal (n = 11, P > 0.05) or in inflamed joints (n = 4, P > 0.05). However, the vasodilator responses to the NK1 receptor agonist [Sar9, Met(O2)11]‐substance P in inflamed joints were abolished by two bolus doses of SR48968 (1 nmol), with the exception of the vasodilator response to the highest dose of the NK1 receptor agonist (100 pmol) tested, where a small but significant (P < 0.001) vasodilator response (0.58 ± 0.04 V; basal level = 5.37 ± 0.12 V; n = 6) was still present. In contrast, the NK2 receptor agonist [Nle10]‐neurokinin A4–10‐induced vasodilator responses were abolished by the two low doses of SR489868 (10 pmol) in normal (n = 6, P < 0.005) and in inflamed joints (n = 8, P < 0.001), except that to a high dose of the NK2 receptor agonist (100 pmol), where a small but significant (P < 0.001) vasodilator response (0.45 ± 0.22 V; basal level = 5.52 ± 0.05 V; n = 8) still remained in the inflamed joint. This residual vasodilator response in the inflamed joint was also resistant to higher doses of SR48968 (two doses at 1 nmol) (0.53 ± 0.11 V; basal level = 5.57 ± 0.12 V; n = 7, P < 0.001), but was abolished by prior co‐administration of the NK2 receptor antagonist SR48968 and the NK1 receptor antagonist FK888 both at 1 nmol (n = 4, P > 0.05).

The present findings indicate that NK1 receptors are important in mediating tachykinin‐induced plasma extravasation and vasodilatation in normal and acutely‐inflamed rat knee joints. In normal joints, NK2 receptor‐mediated vasodilatation is of minor importance, but it has greater significance in inflamed joints. The data do not support a role for NK3 receptors in mediating tachykinin‐induced vasodilatation or plasma extravasation. It is speculated that suppression of NK1 and NK2 receptor‐mediated responses might be useful in attenuating inflammatory responses in the rat knee joint where tachykinin release is implicated.