Peripheral Leptin Signaling Mediates Formalin-Induced Nociception

Abstract

Accumulating evidence suggests that obesity is associated with chronic pain. However, whether obesity is associated with acute inflammatory pain is unknown. Using a well-established obese mouse model induced by a high-fat diet, we found that: (1) the acute thermal pain sensory threshold did not change in obese mice; (2) the model obese mice had fewer nociceptive responses in formalin-induced inflammatory pain tests; restoring the obese mice to a chow diet for three weeks partly recovered their pain sensation; (3) leptin injection induced significant phosphorylation of STAT3 in control mice but not in obese mice, indicating the dysmodulation of topical leptin-leptin receptor signaling in these mice; and (4) leptin-leptin receptor signaling-deficient mice (ob/ob and db/db) or leptin-leptin receptor pathway blockade with a leptin receptor antagonist and the JAK2 inhibitor AG 490 in wild-type mice reduced their nociceptive responses in formalin tests. These results indicate that leptin plays a role in nociception induced by acute inflammation and that interference in the leptin-leptin receptor pathway could be a peripheral target against acute inflammatory pain.

Keywords: Formalin test; Leptin; Nociception; Obesity.

Fig. 1

Body weight, body composition, and plasma leptin levels of obese mice and chow diet mice. The high-fat diet caused increased weight gain (A) and fat gain (B) compared with chow diet controls; plasma leptin levels were increased significantly after 12 weeks on the high-fat diet administration (C); n = 4–8/group, mean ± SEM, high-fat diet vs chow diet, ***P < 0.001.

Fig. 2

Acute thermal nociceptive responses and spontaneous locomotor activity of obese mice and chow diet mice. The high-fat diet group’s responses to acute noxious heat (A Paw withdrawal latency; B Tail-flick latency) were the same as the chow diet controls; the spontaneous locomotor activity of both groups were not statistically different (C Time course; D Total distance traveled); n = 8/group, mean ± SEM.

Fig. 3

Formalin-induced nociceptive responses of obese mice and chow diet mice. The nociceptive behavior during the A first (0–9 min) and B second (10–45 min) phases after 0.25%, 0.5%, and 1% formalin injection was measured as the number of flinching motions made by the injected paw; n = 8/group, mean ± SEM, *P < 0.05, **P < 0.01, ***P < 0.001, high-fat diet vs chow diet.

Fig. 4

Formalin-induced nociceptive responses of mice 3 weeks after switching to a chow diet. A Flowchart of this study; the decreased nociceptive behavior of obese mice during the (B) first (0–9 min) and (C) second (10–45 min) phases after 1% formalin injection significantly recovered and was associated with decreased body weight (D), fat content (E), and leptin levels in plasma (F); n = 8/group, mean ± SEM, *P < 0.05, ** P < 0.01, ***P < 0.001, obese chow diet vs obese high fat diet.

Fig. 5

Formalin-induced inflammatory nociceptive responses of wild-type mice and leptin–leptin receptor signaling-deficient mice. The formalin-induced nociceptive behavior was decreased in ob/ob (A, B, and C) and db/db mice (D, E and F) during the first (0–9 min) and second (10–45 min) phases; n = 8/group, mean ± SEM; *P < 0.05, **P < 0.01, ob/ob or db/db mice vs wild-type mice.

Fig. 6

STAT activation in topical paw skin 2 h after leptin injection (0.1 µg/paw); p-STAT3/STAT3 expression in chow diet (A) and high-fat diet mice (B); n = 4/group, mean ± SEM, *P < 0.05 (student t test); NS, not significant; leptin vs saline group.

Fig. 7

Effects of leptin antagonist and AG 490 on formalin-induced nociceptive responses in normal mice; pretreatment with the leptin antagonist at 0.3 to 3 µg inhibited formalin-induced nociceptive behavior during the first (A 0–9 min) and second (B 10–45 min) phases in a dose-dependent manner; pretreatment with the leptin signaling pathway inhibitor AG 490 (JAK2 inhibitor) at 1 to 10 µg inhibited formalin-induced nociceptive behavior during the first (C 0–9 min) and second (D 10–45 min) phases in a dose-dependent manner; n = 7–8/group, mean ± SEM; *P < 0.05, **P < 0.01, leptin antagonist or AG 490 vs vehicle group.