kjpp The Korean Journal of Physiology & Pharmacology Korean J Physiol Pharmacol 1226-4512 2093-3827 Korean J Physiol Pharmacol 10.4196/kjpp.2010.14.5.257 kjpp-14-257 Original Article Neuroprotective Effect of Visnagin on Kainic Acid-induced Neuronal Cell Death in the Mice Hippocampus KwonMin-Soo 2 ∗ LeeJin-Koo 1 ∗ ParkSoo-Hyun 1 SimYun-Beom 1 JungJun-Sub 1 WonMoo-Ho 3 KimSeon-Mi 1 SuhHong-Won 1 Department of Pharmacology, Institute of Natural Medicine, College of Medicine, Hallym University, Chuncheon 200-702, Korea Department of Aerospace Medical Research, Aerospace Medical Center, ROKAF (Republic of Korea Air Force), Cheongwon 363-842, Korea Department of Anatomy and Neurobiology, and Institute of Neurodegeneration and Neuroregeneration, College of Medicine, Hallym University, Chuncheon 200-702, Korea Corresponding to: Hong-Won Suh, Department of Pharmacology, College of Medicine, Hallym University, 1, Okchun-dong, Chuncheon 200-702, Korea. (Tel) 82-33-248-2614, (Fax) 82-33-248-2612, (E-mail) hwsuh@hallym.ac.kr

Equally contributed to first author.

 022010 18022010 14 5 257 263 11052010 03092010 10092010 Copyright © 2010 Korean J Physiol Pharmacol 2010 This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. Abstract

Visnagin (4-methoxy-7-methyl-5H-furo[3,2-g][1]-benzopyran-5-one), which is an active principle extracted from the fruits of Ammi visnaga, has been used as a treatment for low blood-pressure and blocked blood vessel contraction by inhibition of calcium influx into blood cells. However, the neuroprotective effect of visnagin was not clearly known until now. Thus, we investigated whether visnagin has a neuroprotective effect against kainic acid (KA)-induced neuronal cell death. In the cresyl violet staining, pre-treatment or post-treatment visnagin (100 mg/kg, p.o. or i.p.) showed a neuroprotective effect on KA (0.1 μg) toxicity. KA-induced gliosis and proinflammatory marker (IL-1β, TNF-α, IL-6, and COX-2) inductions were also suppressed by visnagin administration. These results suggest that visnagin has a neuroprotective effect in terms of suppressing KA-induced pathogenesis in the brain, and that these neuroprotective effects are associated with its anti-inflammatory effects.

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The effect of visnagin administered orally or intraperitoneally on KA-induced neuronal death in the hippocampus. The location of the hippocampal CA3 region performing cresyl violet-positive neuronal count is indicated (A) referring Franklin [18]. I.c.v. kainic acid injecton induced neuronal cell death in the pyramidal cells in the CA3 region of hippocampus (B). Mice were administered visnagin orally (C) or intraperitoneally (D) 20 min (– 20) prior to KA (0.1 μg/5 μl) injection or 0, 20 (+ 20), 40 (+40), 60 (+ 60) min after KA treatment (0.1 μg/5 μl). And then, the cresyl violet staining was performed at 1 day after KA. The vertical bars indicate the standard error of mean. ∗p<0.05, ∗∗p<0.01, ∗∗∗p<0.001 (KA 0.1 vs other groups). The mice number of each group was 10.

The time course alteration of proinflammatory cytokines mRNA on KA injection in the hippocampus. Alteration of IL-1β, TNF-α (B), IL-6 (C), and COX-2 (D) mRNA at 1, 3, 6, 12, 24 hrs after intracerebral (i.c.v.) KA (0.1 μg/5 μl) injectio was examined in the hippocampus. Control group (CON) was injected with i.c.v. PBS. The mice number of each group was 3. Experiments were conducted independently three times, giving a total of nine per group in the final statistical analysis. ∗∗p<0.01(CON vs other groups), ∗∗∗p<0.001 (CON vs other groups).

The effect of visnagin on proinflammatory cytokines increased by KA in the hippocampus. Alteration of –1β, TNF-α (B), IL-6 (C), and COX-2 (D) at 1 (A), 3 hrs (B, D), or 6 hrs (C) after intracerebroventricular (i.c.v.) KA injection was examined in the hippocampus. The increased IL-1β, TNF-α (B), IL-6 (C), and COX-2 (D) by KA was decreased by visnagin treated prior to 20 min. The mice number of each group was 3. Experiments were conducted independently three times, giving a total of nine per group in the final statistical analysis. ∗p<0.05, ∗∗p<0.01, ∗∗∗p<0.001 (CON vs KA), ++p<0.01 (Vis vs Vis + KA). CON, vehicle (i.p.) + PBS (i.c.v.); KA, vehicle (i.p.) + KA (i.c.v.); Vis, Visnagin (i.p.) + PBS (i.c.v.).

The effect of visnagin on the GFAP, OX-42 expression induced by KA in hippocampal CA3 region. OX-42 and GFAP immunoreactivities (A) in the hippocampus were examined at 24 hr after i.c.v. injection of KA (0.1 μg/5 μl). Animals (N=5 per each group) were pretreated orally with either PBS or visnagin (100 mg/kg) 20 min prior to KA (0.1 μg/5 μl). To quantify, we counted GFAP or OX-42 IR in each section (B). CON (a): vehicle (i.p.) + PBS (i.c.v.), KA (b): vehicle (i.p.) + KA (i.c.v.), Vis (c): Visnagin (i.p.) + PBS (i.c.v.), Vis + KA (d): Visnagin (i.p.) + KA (i.c.v.). Antibody against OX-42 and GFAP was used at 1: 10,000 dilution for immunostaining. ∗p<0.05 (KA vs Vis + KA), ∗∗p<0.01 (KA vs Vis + KA).