The role of SP in joint inflammation is unclear (Keeble & Brain, in press). We have developed techniques to quantify the contribution of substance P (SP) to plasma extravasation (index of oedema formation) and intravascular volume (index of blood flow/volume change) in murine joint inflammmation.

Female wild type (WT) Sv129+C57BL/6 mice or tachykinin NK₁ receptor knockout (KO) mice were administered ¹²⁵I-albumin i.v. under urethane anaesthesia and immediately injected intra-articularly with SP (100 pmol, 10 µl) and saline (contralateral joint, 10 µl) either into naïve joints or joints that had been treated with complete Freund's adjuvant (CFA, 10 µg; 10 µl) 4.5 hours previously. Plasma extravasation was allowed to develop for 30 min and radioactivity in the joints was then measured using a collimated gamma probe. In separate experiments, the ¹²⁵I-albumin was administered 2 min before the meaurement of radioactivity to determine the intravascular volume of the joints. ¹²⁵I-albumin accumulation was expressed as a ratio of counts in the SP-treated joint compared to the saline-treated joint. Knee diameter was taken at the end of all experiments. Statistics used were either one sample or two sample student's t tests.

Exogenous SP caused a significant increase in plasma extravasation after 30 min in the naïve WT joint (1.21 ± 0.05, p<0.05) without affecting the intravascular volume (1.03 ± 0.05). This response was abolished in NK₁ KO mice (1.02 ± 0.03). By comparison, SP had no effect on knee swelling in either WT (SP, 3.55 ± 0.03 mm; saline, 3.54 ± 0.02 mm) or KO mice (SP, 3.59 ± 0.03 mm; saline, 3.57 ± 0.03 mm). In the CFA-pretreated WT joint, SP produced a significant increase in both plasma extravasation (1.11 ± 0.02; p<0.05) and knee swelling (SP, 3.66 ± 0.07 mm; saline, 3.58 ± 0.07 mm; p<0.05). Again, no effect of SP was observed in KO mice (SP, 3.55 ± 0.05 mm; saline, 3.55 ± 0.06 mm).

Further experiments were performed in which WT and KO mice were injected with CFA (10 µg, 10 µl) and saline (contralateral joint, 10 µl) and inflammation was allowed to develop for 5 h. In knee joints treated with CFA alone, plasma extravasation and knee swelling was similar in WT and KO mice after 5 h. However, the intravascular volume of the CFA-treated joint in KO animals was significantly less than that of the saline control joint and that of WT animals at this time point (WT, 1.05 ± 0.02; KO, 0.89 ± 0.02; p<0.05).

The results show that exogenous SP causes acute plasma extravasation in the naïve and CFA-pretreated mouse joint via activation of the NK₁ receptor. Furthermore, SP potentiated CFA-induced knee swelling. In contrast, the results suggest that endogenous tachykinin-mediated activation of NK₁ receptors does not mediate CFA-induced plasma extravasation or knee swelling. However, the NK₁ receptor may affect blood flow during the early stages of joint disease as intravascular volume measurements were reduced in KO mice.

Keeble J.K. & Brain S.D. Neurocience. Letts., in press.

J.Keeble is funded by the Arthritis Research Campaign.