*Corresponding author: Mehdi Khaksari. School of Medicine, Shahroud University of Medical Sciences, Shahroud, Iran. Tel: +98-2332395054; Fax: +98-2332395054; Email: Khaksari417@yahoo.com Iranian Journal of Basic Medical Sciences ijbms.mums.ac.ir Usnic acid improves memory impairment after cerebral ischemia/ reperfusion injuries by anti-neuroinflammatory, anti-oxidant, and anti-apoptotic properties Sohaila Erfani 1 , Tahereh Valadbeigi 1 , Nahid Aboutaleb 2, 3 , Naser Karimi 4 , Ali Moghimi 5 , Mehdi Khaksari 6 * 1 Department of Biology, Faculty of Science, Ilam University, Ilam, Iran 2 Physiology Research Center and Department of Physiology, Faculty of Medicine, Iran University of Medical Sciences, Tehran, Iran 3 Department of Physiology, Faculty of Medicine, Iran University of Medical Sciences, Tehran, Iran 4 Department of Biology, Faculty of Science, Razi University of Kermanshah, Kermanshah, Iran 5 Rayan Center for Neuroscience and Behavior, Department of Biology, Faculty of Science, Ferdowsi University of Mashhad, Mashhad, Iran 6 School of Medicine, Shahroud University of Medical Sciences, Shahroud, Iran A R T I C L E I N F O A B S T R A C T Article type: Original article Objective(s): Cerebral ischemia/reperfusion causes complex pathological mechanisms that lead to brain tissue damage. Usnic acid is a lichen secondary metabolite that has many different biological properties including anti-inflammatory and anti-oxidant activities. Therefore, the objective of the current study was to investigate the neuroprotective effects of usnic acid on apoptotic cell death, neuroinflammation, anti-oxidant enzyme activities, and oxidative stress levels after transient cerebral ischemia/reperfusion. Materials and Methods: Forty-two male Wistar rats were randomly assigned to three groups (sham, ischemia/reperfusion, and ischemia/reperfusion+usnic acid). Ischemia was induced by 20 min occlusion of common carotid arteries. Injection of usnic acid (25 mg/kg, intraperitoneally) and saline was done at the beginning of reperfusion time. Morris water maze was applied to assess spatial memory. The protein expression amount was measured using immunohistochemical and immunofluorescence staining. Spectrophotometric assay was performed to determine the levels of anti-oxidant enzymes. Results: Usnic acid significantly reduced caspase-3, glial fibrillary acidic protein- positive and ionized calcium-binding adaptor molecule 1-positive cells (P<0.001) and enhanced spatial memory disorders (P<0.05) due to brain ischemia. In addition, treatment with usnic acid improves effects in the antioxidant system following cerebral ischemia (P<0.05). Conclusion: Our findings indicate that usnic acid has neuroprotective properties, which possibly is applicable as a promising candidate for cerebral injuries caused by ischemia. Article history: Received: Sep 19, 2019 Accepted: Jun 13, 2020 Keywords: Apoptosis Cerebral ischemia Lichen secondary- metabolites Neuroinflammation Spatial memory ►Please cite this article as: Erfani S, Valadbeigi T, Aboutaleb N, Karimi N, Moghimi A, Khaksari M. Usnic acid improves memory impairment after cerebral ischemia/ reperfusion injuries by anti-neuroinflammatory, anti-oxidant, and anti-apoptotic properties. Iran J Basic Med Sci 2020; 23:1225-1231. doi: 10.22038/ijbms.2020.43280.10165 Introduction Lichens are some of the most important sources of biologically active compounds that have been applied in traditional medicine owing to their precious therapeutic properties (1). These organisms have many different biological activities including anti-inflammatory, anti- oxidant, antipyretic, analgesic, anticancer, antimicrobial, antidiabetic, and wound healing properties (2). The most commonly used genus of lichen is Usnea Dill. ex Adans, which has applications in medicine worldwide. For example, Usnea longissima Ach. is a type of lichen species that is extensively used for injury treatment caused by bone fractures and inflammatory skin eruptions (3). Macrolichens are sources of phenolic secondary substances (e.g. phenols, quinones, dibenzofurans, depsides, depsones, depsidones, γ-lactones, pulvinic acid derivatives, and xanthones). Dibenzofuran of usnic acid (2,6-Diacetyl-7,9-dihydroxy-8,9 b-dimethyl-1,3 (2H,9bH)-dibenzofurandione) is known as a lichen secondary metabolite which is generated to form a yellow pigment in several lichen species (4). G. Amo et al. indicated that usnic acid has inhibitory effects on reactive oxygen species (ROS) synthesis due to hydrogen peroxide in a human astrocytoma cell line, and this anti- oxidant ability leads to improvement of cell viability (5). Moreover, usnic acid shows anti-inflammatory impact by reduction of pro-inflammatory cytokine expression, like tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL- 6) and IL-1β, also increases IL-10 expression as an anti- inflammatory cytokine in LPS-stimulated inflammatory cellular model (6). Odabasoglu et al. showed that usnic acid has gastroprotective effects via increasing superoxide dismutase (SOD), glutathione peroxidase (GPx), and glutathione (GSH) activities and decreasing lipid peroxidation (LPO) level in indomethacin-induced gastric ulcer tissues in an animal model (7). Cerebral ischemia/reperfusion (I/R) is the third most common factor of death in developed countries which induces considerable mortality and disability worldwide. It causes complex pathological mechanisms that can lead to tissue damage (8). Excitotoxicity,