Kahweol, a natural diterpene from coffee, induces peripheral antinociception by endocannabinoid system activation

Abstract

Kahweol is a compound derived from coffee with reported antinociceptive effects. Based on the few reports that exist in the literature regarding the mechanisms involved in kahweol-induced peripheral antinociceptive action, this study proposed to investigate the contribution of the endocannabinoid system to the peripheral antinociception induced in rats by kahweol. Hyperalgesia was induced by intraplantar injection of prostaglandin E2(PGE2) and was measured with the paw pressure test. Kahweol and the drugs to test the cannabinoid system were administered locally into the right hind paw. The endocannabinoids were purified by open-bed chromatography on silica and measured by LC-MS. Kahweol (80 µg/paw) induced peripheral antinociception against PGE2-induced hyperalgesia. This effect was reversed by the intraplantar injection of the CB1 cannabinoid receptor antagonist AM251 (20, 40, and 80 μg/paw), but not by the CB2 cannabinoid receptor antagonist AM630 (100 μg/paw). Treatment with the endocannabinoid reuptake inhibitor VDM11 (2.5 μg/paw) intensified the peripheral antinociceptive effect induced by low-dose kahweol (40 μg/paw). The monoacylglycerol lipase (MAGL) inhibitor, JZL184 (4 μg/paw), and the dual MAGL/fatty acid amide hydrolase (FAAH) inhibitor, MAFP (0.5 μg/paw), potentiated the peripheral antinociceptive effect of low-dose kahweol. Furthermore, kahweol increased the levels of the endocannabinoid anandamide, but not of the other endocannabinoid 2-arachidonoylglycerol nor of anandamide-related N-acylethanolamines, in the plantar surface of the rat paw. Our results suggested that kahweol induced peripheral antinociception via anandamide release and activation of CB1 cannabinoid receptors and this compound could be used to develop new drugs for pain relief.

Figure 1. Schematic diagram of the experimental protocol. A, Kahweol (Kah) was administered 175 min after the local administration of prostaglandin E₂(PGE₂) (2 µg) and the antinociceptive response was measured prior to and 180 min, 195 min, and 210 min after PGE₂ injection. B, Kah was administered in the right hind paw 175 min after local injection of PGE₂. The cannabinoid drugs AM251, AM630, MAFP, JZL184, or VDM11 were given 10 min prior (165 min) to Kah intraplantar administration and measurements were performed prior to and 180 min after PGE₂ administration.

Figure 2. Temporal development of the effect induced by intraplantar administration of kahweol on hyperalgesia. Kah was administered 175 min after the local administration of prostaglandin E₂(PGE₂) (2 µg) and the antinociceptive response was measured prior to and 180 min, 195 min, and 210 min after PGE₂ administration. Kah 20: kahweol 20 µg/paw; Kah 40: kahweol 40 µg/paw; Kah 80: kahweol 80 µg/paw; Veh (vehicle) 1: 2% ethanol in saline; Veh 2: 10% DMSO in saline. Data are reported as means±SE (n=5). *P<0.05 compared with PGE₂+Veh 2. ^oP<0.05, between the experimental periods 180 min and 195 min. ^#P<0.05, between the experimental periods 195 min and 210 min. ⁺P<0.05, between the experimental periods 180 min and 210 min (ANOVA and Bonferroni's test).

Figure 3. The CB₁ receptor antagonist blocked kahweol-induced peripheral antinociception in hyperalgesic paws. The antinociceptive response was measured by the paw pressure test. Prostaglandin E₂(PGE₂) injection (2 µg/paw) was done at time 0, AM251 (20, 40, 80 µg/paw) was injected at time 165 min, and kahweol (Kah; 80 µg/paw) was given at 175 min. Measurements were made prior to and 180 min after PGE₂ administration. Data are reported as means±SE (n=5) of Δ nociceptive threshold measured in grams (g). *P<0.05 compared to PGE₂ + Veh 2 + Veh 3; ^#P<0.05 compared to PGE₂ + Veh 2 + Kah 80)-injected groups (ANOVA and Bonferroni's test). Veh (vehicle) 1: 2% ethanol in saline; Veh 2: 10% DMSO in saline; Veh 3: sterile saline solution (0.9% NaCl).

Figure 4. The CB₂ receptor antagonist did not block kahweol-induced peripheral antinociception in hyperalgesic paws. The antinociceptive response was measured by the paw pressure test. Prostaglandin E₂(PGE₂) injection (2 µg/paw) was done at time 0, AM630 (100 µg/paw) was injected at time 165 min, and kahweol (Kah; 80 µg/paw) was given at 175 min. Measurements were made prior to and 180 min after PGE₂ administration. Data are reported as means±SE (n=5) of Δ nociceptive threshold measured in grams (g). *P<0.05 compared to PGE₂ + Veh 2 + Veh 3-injected group (ANOVA and Bonferroni's test). There was no significant difference between (PGE₂ + Veh 2 + Kah 80) and (PGE₂ + AM630 100 + Kah 80)-injected groups. Veh (vehicle) 2: 10% DMSO in saline; Veh 3: sterile saline solution (0.9% NaCl).

Figure 5. Pretreatment with VDM11, JZL184, and MAFP potentiated the peripheral antinociceptive action of kahweol. The antinociceptive response was measured by the paw pressure test. Prostaglandin E₂(PGE₂) injection (2 µg/paw) was done at time 0. VDM11 (2.5 µg/paw; A), JZL184 (4 µg/paw; B), or MAFP (0.5 µg/paw; C) was injected at time 165 min, and kahweol (Kah; 80 µg/paw) was given at 175 min. Measurements were made prior to and 180 min after PGE₂ administration. Data are reported as means±SE (n=5) of Δ nociceptive threshold measured in grams (g). *P<0.05 compared to (PGE₂ + Veh 3 + Veh 4/Veh 5/Veh 6); ^#P<0.05 compared to (PGE₂ + Kah 80 + Veh 4/Veh 5/Veh 6)-injected groups (ANOVA and Bonferroni's test). Veh (vehicle) 1: 2% ethanol in saline; Veh 3: sterile saline solution (0.9% NaCl); Veh 4: 10% tocrisolve in saline; Veh 5: 3% ethanol in saline; Veh 6: 20% DMSO in saline.

Figure 6. Increase of anandamide (AEA) levels (A), but not 2-arachidonoylglycerol (2-AG) (B), oleoylethanolamide (OEA) (C), and palmitoylethanolamide (PEA) (D) on kahweol injection. Kahweol (Kah; 80 μg/paw) was injected into the paw and 5 min later the paw tissues were collected for analysis. Data are reported as means±SE (n=5). *P<0.05 compared to Naive- and Saline (SAL)-injected groups (ANOVA and Bonferroni's test).