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Scopolamine-induced memory impairment is alleviated by xanthotoxin: a role of acetylcholinesterase and oxidative stress processes. K Skalicka-Wozniak, Barbara Budzynska, Grazyna Biala, and Anna Boguszewska-Czubara ACS Chem. Neurosci., Just Accepted Manuscript • DOI: 10.1021/acschemneuro.8b00011 • Publication Date (Web): 29 Jan 2018 Downloaded from http://pubs.acs.org on February 7, 2018
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Scopolamine-induced memory impairment is alleviated by xanthotoxin: a role of acetylcholinesterase and oxidative stress processes.
Krystyna Skalicka-Wozniaka, Barbara Budzynskab*, Grazyna Bialaa, Anna BoguszewskaCzubarac
a
Department of Pharmacology and Pharmacodynamics, 4a Chodzki Str., Medical University
of Lublin, 20-093 Lublin, Poland b
Department of Pharmacognosy with Medicinal Plants Unit, 1 Chodzki Str., Medical
University of Lublin, 20-093 Lublin, Poland c
Department of Medicinal Chemistry, 4a Chodzki Str., Medical University of Lublin, 20-093
Lublin, Poland
Abstract Xanthotoxin, popularly occurring furanocoumarin, which can be found in plants from Apiaceae family, was isolated from fruits of Pastinaca sativa L. by mean of high-performance countercurrent chromatography and its effects on the scopolamine-induced cognitive deficits in male Swiss mice using the passive avoidance (PA) test was evaluated. To measure the acquisition of memory processes, xanthotoxin (1, 2.5, 5 mg/kg) was administered 30 min before PA test and scopolamine was administered 10 min after xanthotoxin. To measure the consolidation of memory processes, xanthotoxin (1 and 2.5 mg/kg) was injected immediately after removing the mouse from the apparatus and 10 min after scopolamine was administered. In subchronic experiments mice were injected with xanthotoxin (1 mg/kg) or saline, 6 days, twice daily. 24 hours after last injection of the drugs, the hippocampus and the prefrontal cortex were removed for biochemical assays. The results demonstrated that either single (2.5 1 ACS Paragon Plus Environment
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and 5 mg/kg) or repeatable (1 mg/kg) administration of xanthotoxin significantly increased index of latency (IL) in both acquisition and consolidation of memory processes, showing some procognitive effects. The behavioral tests also showed that an acute (2.5 mg/kg) and subchronic (1 mg/kg) administration of xanthotoxin prevent memory impairment induced by injection of scopolamine (1 mg/kg). Observed effects could be due to the inhibition of AChE activities and amelioration of oxidative stress processes in the hippocampus and the prefrontal cortex. It was suggested that xanthotoxin could show neuroprotective effect in scopolamineinduced cognitive impairment connected to cholinergic neurotransmission and oxidative stress in the brain structures.
Keywords methoxypsoralen; memory impairment, scopolamine; Apiaceae, oxidative stress, mice, total antioxidant capacity, malondialdehyde
Introduction Xanthotoxin (8-methoxypsoralen; XT) is popularly occurring furanocoumarin, which can be found in plants from Apiaceae family. Together with psoralen and bergapten (5methoxypsoralen), XT is used in PUVA therapy in treatment of vertiligo and psoriaris (1,2). Additionally, antioxidant (3,4), antiproliferative (2) and antimicrobial (5) activity has been reported. XT, among series of tested coumarins, is the strongest inhibitor of HL60 cells proliferation (6). Activity of XT towards the central nervous system (CNS) has been investigated for a long time. Our previous study demonstrated that XT (injected 15, 30, 60, and 120 min before experiments) revealed the anticonvulsant activity in the MES-induced seizure test in mice. It was noticed that the ED50 values were in a range of 219.1-252.4 mg/kg, which suggests that 2 ACS Paragon Plus Environment
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the effect of target compound can be compared to the conventional drug e.g. valproate (7). Moreover, XT, at a doses of 50 and 100 mg/kg, intraperitoneally (i.p.) acts synergistically with the synthetic antiepileptic drugs like carbamazepine and valproate. Additionally, the total brain concentration of carbamazepine and valproate increased at about 84 and 46%, respectively (8). Further experiments using the same model showed that when XT was administered at the dose of 100 mg/kg, (i.p.) the anticonvulsant activity of other antiepileptic drugs like lacosamide, oxcarbazepine, pregabalin, and topiramate, was significantly enhanced, however their total brain concentrations were not changed (9). Antiseizure activity may be due to the ability to interfere with voltage-dependent sodium channels and blockade of highvoltage-activated calcium channels, which are widely known to play a role in therapeutic activity of many drugs, including antiepileptic medicines (8,9). It was also revealed that XT (15 mg/kg), as a CYP2A6 inhibitor, prolongs the antidepressive and procognitive effects of nicotine (10). From our previous results it was concluded that imperatorin, C-8 substituted structural analogue of XT, may act as a protective agent against either scopolamine (SC)-induced memory impairments or oxidative stress. In this model of cognitive impairment, imperatorin (acute administration, 5 and 10 mg/kg before SC injection, 1 mg/kg) enhanced memory acquisition and consolidation disrupted by SC. After chronic administration of imperatorin at the dose of 10 mg/kg, significant reduction of the influence of SC on memory acquisition was noticed. Additionally, tested compound increased antioxidant enzymes activity in brain structures like superoxide dismutase, glutathione peroxidase and glutathione reductase (11). Moreover, when imperatorin was administered at the doses of 10 and 20 mg/kg (acute administration, the impairment of the anxiogenic effect of nicotine (0.1 mg/kg) and improvement of cognitive processes were noticed (both in acquisition and consolidation trials). When imperatorin (1 mg/kg) was co-administered with non-active dose of nicotine 3 ACS Paragon Plus Environment
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(0.05 mg/kg, subcutaneously, s.c), it improved the processes of memory acquisition and enhanced both memory acquisition and consolidation after repeated administration, which may indicate the influence of both compounds on the cholinergic system (12).
As it was revealed previously, the C-8 substitution in psoralen ring may be crucial for activity toward the CNS. Both XT and imperatorin showed dose-dependent anticonvulsant activity, as well as prolonged the duration of nicotine-induced acquisition and consolidation of memory improvement, whereas bergapten (C-5 substituted structural analogue of XT) influenced only memory acquisition in the passive avoidance (PA) test and was not active in the mouse maximal electroshock-induced seizure model (10,13). To extend the current knowledge about the pharmacological and biochemical mechanisms of coumarins, we evaluated the influence of XT on cognition and memory processes. Moreover, in spite of long-term research, no effective medicine for cognitive deficits has been developed so far. Currently existed cholinesterase inhibitors (galantamine, tacrine, rivastigmine, donepezil), and N-methyl-Daspartate blockers (memantine) have some limitations as well as serious adverse effects, thus there is increasing demand to develop new drugs, preferably from natural sources (14). Thus, the behavioral interactions between XT and disturbing memory, cholinergic agents – SC, administered acutely and chronically, were determined. The effect of acute and subchronic administration of XT on SC-induced learning and memory impairments in mice was investigated in the PA test. In order to verify the involvement of cholinesterase inhibition in the molecular mechanism of XT, the activity of AChE was also examined in the prefrontal cortex and the hippocampus, the structures important for cognitive functions. Finally, to find out if antioxidant mechanism is also applied in neuroprotective action of XT, the oxidative stress markers in aforementioned brain structures were also determined.
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Results and discussion The influence of XT on SC-induced memory impairments in the PA task was evaluated. Additionally, the underlying mechanisms were examined for the first time. SC, a nonselective muscarinic antagonist, produces memory deficits by blocking muscarinic receptors in the CNS. Apart from neurochemical alterations, memory impairment induced by SC may be a result of imbalance of oxidative stress in brain (15). These effects are comparable to those occurring in older people and result in age-related dementia (16,17), so animal model where memory is impaired by injection of SC is useful for the imitation of some dysfunction of the CNS, like AD, which is characterized by hippocampal-dependent learning and memory deficits, oxidative stress, as well as behavioral disorder.
Behavioral studies Behavioral tests, including Morris water maze (MWM) test and the PA test, are often used to evaluate cognitive impairments in murine models. In our study, the PA test was applied to assess the memory and learning deficits in mice. Exposure to painful experience like electric foot shock after entering the dark chamber, stopped animal to re-enter this chamber and this is how cognitive ability can be measured. SC administration before the acquisition trial impaired the learning and memory processes and administration of any other substance, such XT in our case, can let to determine the potency of the drug on cognitive functions (18).
Single injection of XT affects memory-related processes in the PA test in mice XT (1, 2.5, 5 mg/kg, acute trial, acute administration), significantly changed index of latency (IL) values [F(2,27)=7.437; p=0.0009, one-way ANOVA]. Higher doses (2.5 and 5 mg/kg) significantly increased IL (p
