Figure 6. Simvastatin attenuated KA-induced seizure behavior in rats. The rats were orally treated with simvastatin (1 mg/kg) for 0.5 h after KA injection (1.0 ml of KA. i.c.v). The seizure behavior test (seizure score) was performed during 4–6 months after KA injection (1.0 ml of KA. i.c.v). Simvastatin attenuated the seizure score. Data were presented as means6standard deviation. *P,0.05 versus the saline group. doi:10.1371/journal.pone.0024966.g006 In the future study, it will be intriguing to assess the loss of inhibi- tory interneurons and neurotransmitter expression after simvasta- tin treatments. Possible mechanism underlying anti-excitotoxic effect of simvastatin The possible mechanisms underlying the beneficial effect of statins on neurodegenerative and cardiovascular diseases have been extensively investigated and discussed in previous literature. While the study of the effect of statins in epileptic conditions has been limited, previous research has shown the evidence supporting the anti-excitotoxic effect of statins [17], which might explain the beneficial effect of simvastin after KA-induced injury. For instant, in vitro study has shown that statins, especially rosuvastatin and simvastatin, protect cultured neurons from NDMA-induced excitotoxic death [31]. Atorvastatin has also been shown to protect primary cortical neurons from glutamate-induced excitotoxicity possibly by atten- uating glutamate-induced intracellular calcium signaling that modulating NMDA receptor function [71]. Anti-excitotoxic and anti-seizure effects of statins have been further studied in KA model of TLE. Pre-treatment with atorvastatin efficiently reduced KA-induced seizure activities and neuronal loss [35]. Moreover, a very recent in vivo study, which compared 5 commercially available statins in KA model of TLE, suggests that simvastatin is the most effective statin in attenuating KA-induced excitotoxicity and improving memory [30]. Our results in the present study further support the anti-excitotoxicity effect of statins in epileptic conditions and this protective effect could last until 6 months after KA-induced SE. Considering previous studies together with our results, we suggest that the roles of statins in regulating glutamate receptor and inhibiting injury-induced inflammatory response may contribut to the beneficial effect in the epileptic conditions. Simvastatin shows a long-term neuroprotective effect on epileptic brain Previous studies have investigated effects of two statins, atoro- vastatin [35], [46], [72] and lovastatin [73], [74], on neuronal Simvastatin Restrains Exacerbation of Epilepsy protection during epileptogenesis and attenuating seizure behavior in various animal models, including pentylentetrazole-kindled rats, quinolinic acid-induced seizure, pilocarpine-induced status epilep- tic and kainic-acid induced seizure. A very recent study has compared 9 statins by analyzing several parameters that could be related to neuroprotection and suggests that simvastatin presents the best characteristics for preventing neurodegenerative condi- tions, due to its high BBB penetration capacity and cholesterol lowering effect on neurons [29]. Moreover, study performed by the same group also shows that simvastatin is the most efficient statin in protecting against KA-induced excitotoxicity and memory deficits [30]. However, all animal studies above have mainly focused on the transient effect of the statin administration, ranging from 3 days to 2 weeks after SE. In the present study, we evaluated the long-term neuroprotection of simvastatin at 4, 5 and 6 months after KA-induced SE and our data showed a consistent result as transient outcome, suggesting that a 14-day administra- tion of simvastatin has a beneficial effect that can last for 6 months. Our data also support previous studies of transient outcome of statins administration. This long-term effect of simvastamin suggests that it may have therapeutic potential in the treatment of neurodegenerative disease involving excitotoxicity. Conclusion Our data showed that 2-week administration of simvastatin immediately after KA-induced seizure lead to reduced astrocytosis, attenuated neuronal loss, decreased abnormal MFS and reduced seizure activity in the brain at 4 to 6 month after KA lesion, suggesting a long-term protective role of simvastatin in the epileptic brain. This study supports the possibility of using simvastatin administration as a neuroprotective intervention for retarding progression of the epileptogenesis and a therapeutic strategy for epilepsy. Materials and Methods Experimental Groups Adult male Wister rats (200–250 g) were used in our experiment and all animals were maintained under 12 h light/dark-cycle with free access to food and water in a temperature-controlled room. All animal procedures were in accordance to guidelines of Ethics Committee of the First Clinical College of Harbin Medical University (201001). Rats were divided into 3 groups: 1) SE were induced in rats with KA injection followed by oral administration of saline starting at 0.5 h after SE once a day for 14 consecutive days; 2) Rats were subjected to KA-lesion followed by oral administration of simvastatin (1 mg/kg/day) starting at 0.5 h after SE for 14 consecutive days. The dose was selected according to our previous study [75]; 3) Rats were subjected to intracerebro- ventricular saline injection as a control. Rats with KA-lesion were sacrificed at 3 days, 4 and 6 months after simvastatin- or saline- administration. Rat cerebral hemispheres were homogenized for enzyme-linked immunosorbent assay (ELISA) at 3 day after KA- lesion. Kainic acid lesion Rats were anesthetized with chloral hydrate (40 mg/kg body weight, i.p.), then immobilized in a stereotaxic apparatus (WPI Stoelting, USA) with a plane of incision bar set at 3.360.33 mm below the interaural line. After making a midline incision, the skull was exposed and one burr hole was drilled according to the coordinates: 3.7 mm posterior to the bregma, 4.1 mm lateral to the midline, and 3.5 mm under the dura. Each rat was injected with 1.0 ml of KA (0.5 mg in 1.0 ml Saline) in the right lateral PLoS ONE | www.plosone.org 7 September 2011 | Volume 6 | Issue 9 | e24966