Possible mechanisms involved in the anti-nociceptive effects of hydro-ethanolic leaf extract of Ziziphus abyssinica

Abstract

Context: Various parts of Ziziphus abyssinica Hochst ex. A. Rich (Rhamnaceae) have been used in Ghanaian and African traditional medicine as an analgesic. However, there are little scientific data to support the anti-nociceptive effects of the hydro-ethanolic leaf extract of Ziziphus abyssinica (EthE) as well as the possible mechanisms involved in its anti-nociceptive effects.

Purpose: To predict possible nociceptive pathways involved in the anti-nociceptive effects of EthE.

Materials and methods: The effect of EthE (30, 100 and 300 mg/kg) on intraplantar injection of pain mediators such as interleukin-1β, tumour necrosis factor-α, prostaglandin E₂ and bradykinin was evaluated in male Sprague Dawley rats using Randall-Selitto test for 5 h. The effect of specific antagonists to the opioidergic, adenosinergic, ATP-sensitive K⁺ channels, nitric oxide, serotonergic, muscarinic, adrenergic and voltage-gated calcium channel on the anti-nociceptive effect of EthE (100 mg/kg) was evaluated using the formalin test in male imprinting control region (ICR) mice for 1 h.

Results: Pretreatment of the rats with EthE significantly reversed the hypernociception induced by intraplantar injection of TNF-α (F_4,120 = 10.86, p < 0.0001), IL-1β (F_4,120 = 14.71, p < 0.0001), bradykinin (F_4,80 = 12.52, p < 0.0001) and prostaglandin E₂ (F_5,144 = 6.165, p = 0.0001). The anti-nociceptive effect exhibited by EthE in the formalin test was reversed by systemic administration of N^G-l-nitro-arginine methyl ester, naloxone, theophylline and glibenclamide.

Conclusions: EthE inhibits hypernociception induced by TNF-α, IL-1β, bradykinin and prostaglandin E₂. EthE exhibited anti-nociceptive effects possibly mediated through opioidergic, adenosinergic, ATP-sensitive potassium channels and nitric oxide cyclic GMP pathways.

Keywords: IL-1β; TNF-α; bradykinin; nociception; prostaglandin.

Figure 1.

Effect of pretreatment of rats with EthE (30–300 mg/kg, p.o.) and morphine (3 mg/kg, i.p.) on TNF-α-induced hypernociception. Each datum represents the mean of five animals and the error bars indicate SEM. The symbols * and † indicate significance levels compared to respective control groups: (a) represent the time-course curves ***p < 0.001, **p < 0.01 (two-way ANOVA followed by Bonferroni’s post hoc test), whereas (b) represents total anti-nociceptive effects (AUC) ††p < 0.01 and †p < 0.05 (one-way ANOVA followed by Newman–Keuls post hoc test).

Figure 2.

Effect of pretreatment of rats with EthE (30–300 mg/kg, p.o.) and morphine (3 mg/kg, i.p.) on IL-1β-induced hypernociception. Each datum represents the mean of five animals and the error bars indicate SEM. The symbols * and † indicate significance levels compared to respective control groups: (a) represents the time-course curves **p < 0.01, *p < 0.05 (two-way ANOVA followed by Bonferroni’s post hoc test), whereas (b) represents total anti-nociceptive effects (AUC) ††p < 0.01 and †p < 0.05 (one-way ANOVA followed by Newman–Keuls post hoc test).

Figure 3.

Effect of pretreatment of rats with EthE (30–300 mg/kg, p.o.) and morphine (3 mg/kg, i.p.) on bradykinin-induced hypernociception. Each datum represents the mean of five animals and the error bars indicate SEM. The symbols * and † indicate significance levels compared to respective control groups: (a) represents the time-course curves ***p < 0.001, **p < 0.01, *p < 0.05 (two-way ANOVA followed by Bonferroni’s post hoc test), whereas (b) represents total anti-nociceptive effects (AUC) †††p < 0.001 and ††p < 0.01 (one-way ANOVA followed by Newman–Keuls post hoc test).

Figure 4.

Effect of pretreatment of rats with EthE (30–300 mg/kg, p.o.), morphine (3 mg/kg, i.p.) and diclofenac (10 mg/kg, i.p.) on PGE₂-induced hypernociception. Each datum represents the mean of five animals and the error bars indicate SEM. The symbols * and † indicate significance levels compared to respective control groups: (a) represents the time-course curves **p < 0.01, *p < 0.05 (two-way ANOVA followed by Bonferroni’s post hoc test), whereas (b) represents total anti-nociceptive effects (AUC) ††p < 0.01 (one-way ANOVA followed by Newman–Keuls post hoc test).

Figure 5.

Effect of pretreatment of mice with (a) yohimbine (3 mg/kg, p.o.), (b) nifedipine (10 mg/kg, p.o.), (c) atropine (5 mg/kg, i.p.), (d) naloxone (2 mg/kg, i.p.), (e) granisetron (2 mg/kg, p.o.), (f) L-NAME (10 mg/kg, i.p.), (g) glibenclamide (8 mg/kg, p.o.) and (h) theophylline (10 mg/kg, i.p.) on the nociceptive scores of EthE (100 mg/kg, p.o.) on the time-course curves of formalin-induced nociceptive test. Each point represents the mean of five animals and the error bars indicate SEM. ***p < 0.001, **p < 0.01 and *p < 0.05 compared to the control group at same time points (two-way ANOVA followed by Bonferroni’s post hoc test).

Figure 6.

Effect of pretreatment of mice with yohimbine (3 mg/kg, p.o.), nifedipine (10 mg/kg, p.o.), atropine (5 mg/kg, i.p.), naloxone (2 mg/kg i.p.), granisetron (2 mg/kg, p.o.), L-NAME (10 mg/kg, i.p), glibenclamide (8 mg/kg, p.o.) and theophylline (10 mg/kg, i.p.) on the total nociceptive score of EthE (100 mg/kg, p.o.) in phase 1 and phase 2 of formalin-induced nociception. Each column represents the mean of five animals and the error bars indicate SEM. ***p < 0.001 compared to control group and †††p < 0.001, ††p < 0.01 and †p < 0.05 compared to EthE-alone-treated group (one-way ANOVA followed by Newman–Keuls post hoc test).

Figure 7.

Effect of pretreatment of mice with (a) yohimbine (3 mg/kg, p.o.), (b) nifedipine (10 mg/kg, p.o.), (c) atropine (5 mg/kg, i.p.), (d) naloxone (2 mg/kg, i.p.), (e) granisetron (2 mg/kg, p.o.), (f) L-NAME (10 mg/kg, i.p.), (g) glibenclamide (8 mg/kg, p.o.) and (h) theophylline (10 mg/kg i.p.) on the total nociceptive score of morphine (3 mg/kg, i.p.) on the time-course curves of formalin-induced nociceptive test. Each point represents the mean of five animals and the error bars indicate SEM. ***p < 0.001, **p < 0.01 and *p < 0.05 compared the control group at same time points (two-way ANOVA followed by Bonferroni’s post hoc test).

Figure 8.

Effect of pretreatment of mice with yohimbine (3 mg/kg, p.o.), nifedipine (10 mg/kg, p.o.), atropine (5 mg/kg, i.p.), naloxone (2 mg/kg i.p.), granisetron (2 mg/kg, p.o.), L-NAME (10 mg/kg, i.p), glibenclamide (8 mg/kg, p.o.) and theophylline (10 mg/kg, i.p.) on the total nociceptive scores of morphine (3 mg/kg, i.p.) in phase 1 and phase 2 of formalin-induced nociception. Each column represents the mean of five animals and the error bars indicate SEM. ***p < 0.001 compared to control group and †††p < 0.001, ††p < 0.01 and †p ≤ 0.05 compared to morphine-alone-treated group (one-way ANOVA followed by Newman–Keuls post hoc test).