Inhibition of D-amino-Acid oxidase activity induces pain relief in mice

Abstract

(1). We investigated the effects of inhibiting D: -amino-acid oxidase (DAO) activity on nociceptive responses through the use of mutant ddY/DAO(-) mice, which lack DAO activity, and through the application of a selective inhibitor of DAO, sodium benzoate, in the tail flick test, hot-plate test, formalin test, and acetic acid-induced writhing test. (2). Compared with normal ddY/DAO+ mice, ddY/DAO(- )mice showed significantly prolonged tail withdrawal latency in the tail flick test and licking/jumping latency in the hot-plate test, as well as significantly reduced duration of licking/biting in the late phase of the formalin test and the number of abdominal writhing in the acetic acid-induced writhing test. (3). In addition, we investigated the effects of sodium benzoate in Kunming mice having normal DAO activity. (4). Intravenous administration of sodium benzoate (400 mg/kg) significantly inhibited pain responses of the late phase of the formalin test and abdominal writhing responses in the acetic acid-induced writhing test, with no effects on the early phase flinch responses in the formalin test, nociceptive responses in the tail flick test, or hot-plate test. (5). These results suggest that DAO acts as a pro-nociceptive factor in pain, particularly chronic pain, transmission and modulation, and may be a target for pain treatment.

Fig. 1

Kidney DAO activity in ddY/DAO⁻ mice, ddY/DAO⁺ mice, and Kunming mice. DAO activity was expressed as nmol pyruvate formed per min/mg wet weight of the kidneys. Data were presented as means ± S.E.M. (**P < 0.01 compared with ddY/DAO⁺ mice; two-tailed Student’s t-test; n = 10 in each group)

Fig. 2

Acute pain responses in ddY/DAO⁺ and ddY/DAO⁻ mice. Mutant ddY/DAO⁻ mice demonstrated significantly prolonged nociceptive response latencies than ddY/DAO⁺ mice in the tail flick test (50 ± 0.5°C) (A) and hot-plate test (55 ± 0.1°C) (B). Data were presented as means ± S.E.M. (**P < 0.01 compared with ddY/DAO⁺ mice; two-tailed Student’s t-test; n = 10 in each genotype)

Fig. 3

Depressed pain sensitivity in ddY/DAO⁻ mice in formalin test. Intraplantar injection of 5% formalin (10 μl) resulted in depressed nociceptive responses in ddY/DAO⁻ mice compared to ddY/DAO⁺ mice. In the late phase (10–30 min), ddY/DAO⁻ mice displayed a shorter duration of nociceptive behavior than wild-type controls. Values were means ± S.E.M. (n = 10 in each group). **P < 0.01 compared with ddY/DAO⁺ mice, two-tailed Student’s t-test

Fig. 4

Depressed pain sensitivity of ddY/DAO⁻ mice in acetic acid-induced writhing test. Writhing was induced by an intraperitoneal injection of acetic acid (1.5%, 6 ml/kg) and evaluated by the number of writhing movements in 15 min of observation immediately after the acetic acid injection. The results were expressed as means ± S.E.M. (n = 8 in each group). **P < 0.01 compared with ddY/DAO⁺ mice, two-tailed Student’s t-test

Fig. 5

Effects of intravenous (i.v.) injection of sodium benzoate (400 mg/kg) on pain threshold in Kuming mice in the tail flick test (50 ± 0.5°C) (A), and hot-plate test (55 ± 0.1°C) (B). Data were presented as means ± S.E.M. (n = 10 in each group)

Fig. 6

Effects of sodium benzoate on formalin-induced nociceptive respinses in Kunming mice. Animals were pre-treated with sodium benzoate (400 mg/kg, i.v.) 30 min prior to formalin injection (5%, 10 μl). Total time of the licking responses (sec) during the early phase (0–5 min) and the late phase (15–30 min) to formalin injection was presented as the means ± S.E.M. (*P < 0.05 compared with normal saline animals; two-tailed Student’s t-test; n = 8 in each group)

Fig. 7

Effects of sodium benzoate on acetic acid-induced writhing responses in Kunming mice. Writhing was induced by acetic acid (1.5%, 6 ml/kg, i.p.) and evaluated by recording the number of writhing movements in 15 min of observation period immediately after the acetic acid injection. Animals were pre-treated with sodium benzoate (400 mg/kg, i.v.) 30 min prior to the injection of acetic acid. Data were presented as means ± S.E.M. (**P < 0.01 compared with the normal saline control; two-tailed Student’s t-test; n = 8 in each group)