Tanshinone IIA Exhibits Anticonvulsant Activity in Zebrafish and Mouse Seizure Models

Abstract

Danshen or Chinese red sage (Salvia miltiorrhiza, Bunge) is used by traditional Chinese medicine (TCM) practitioners to treat neurological, cardiovascular, and cerebrovascular disorders and is included in some TCM formulations to control epileptic seizures. In this study, acetonic crude extracts of danshen inhibited pentylenetetrazol (PTZ)-induced seizure activity in zebrafish larvae. Subsequent zebrafish bioassay-guided fractionation of the extract resulted in the isolation of four major tanshinones, which suppressed PTZ-induced activity to varying degrees. One of the active tanshinones, tanshinone IIA, also reduced c-fos expression in the brains of PTZ-exposed zebrafish larvae. In rodent seizure models, tanshinone IIA showed anticonvulsive activity in the mouse 6-Hz psychomotor seizure test in a biphasic manner and modified seizure thresholds in a complex manner for the mouse i.v. PTZ seizure assay. Interestingly, tanshinone IIA is used as a prescription drug in China to address cerebral ischemia in patients. Here, we provide the first in vivo evidence demonstrating that tanshinone IIA has anticonvulsant properties as well. KEYWORDS: Tanshinone IIA, Salvia miltiorrhiza, zebrafish PTZ model, mouse seizure models, pentylenetetrazol E pilepsy affects approximately 60 million people worldwide, of whom 30% suffer from pharmacoresistant seizures. The economics of maintaining a cost-effective therapeutic regimen hinge on its long-term efficacy and the probability of patients developing treatment-resistant seizures or possible side-effects ranging from sedation to Steven−Johnson syndrome.1 To address such issues, there is an ongoing hunt for new antiepileptic drugs (AEDs) with novel mechanisms of action and with minimal or no side-effects.There is a resurgence of interest in exploring plant, microbial, and marine resources used in traditional medicine for potential drug leads. According to the World Health Organization (WHO), 70−80% of the population in developing and developed countries have used, or depend on such therapies, which became the impetus for efforts in bioprospecting and drug development.2 Despite prevalent skepticism, as many as 25% of pharmaceutical drugs on the market are plant-based and were discovered through investigations on traditional or folk medicine practiced by different cultures.1,2 The rationale behind this approach in drug discovery is that small molecules isolated or derived from natural sources offer a more diverse set of structures compared with compounds synthesized through medicinal chemistry or combinatorial techniques. 2Each plant or marine extract can be treated as a library potential of hits, which can be screened using an appropriate medium-to high-throughput in vivo model such as zebrafish (Danio rerio), a freshwater teleost of the Cyprinidae family.3,4 In screening for potential hits and leads for AED development, we have previously described the use of larval zebrafish as a platform for pinpointing AED-like activity of small molecules isolated from plant sources.3,4 ...

Cross-species pharmacological characterization of the allylglycine seizure model in mice and larval zebrafish

“…In the latter instance, we were able to circumvent this solubility problem by improving the low bioactivity of a compound through microinjection into the blood circulation after solubilization in 50% DMSO. Surprisingly, zebrafish embryos were able to survive such high concentrations of DMSO solvent quite well [69].…”

The value of zebrafish as an integrative model in effect-directed analysis - a review

“…Natural product studies aimed to identify bioactive compounds for pharmacological applications, investigating mostly plant extracts [19,[34][35][36][37][38][39][40][41][42][43][44][45] but also extracts of bacteria [46], cyanobacteria and algae [47], seaweed [48] and marine organisms [49]. Environmental toxicology studies aimed to identify the toxic compounds in various environmental samples, including marine and fluvial sediments [50][51][52], soil [53], cyanobacteria and algae [54,55], industrial effluent [33], rubber tyre leachates [32], oil sand process waters [56,57] and river pore water [58].…”

“…Zebrafish up to 5 days post fertilization (dpf ) were the life stages mostly applied, except for experiments that extended the assays up to 6 to 7 dpf [34,49,53,56,57] or a few studies with adults [33,47,59]. Environmental toxicology studies for the most part performed exposure not only in 24-well plates (200 μL to 2 mL per embryo or larva) but also in crystallization dishes, scintillation vials or beakers (450 μL to 5 mL per embryo or larva, 40 to 300 mL per adult), while natural product studies were performed exclusively in multiwell-plate setup (<100 to 250 μL per embryo or larva).…”

“…For that, there was exposure of the zebrafish to compounds known to cause specific effects such as anti-convulsant activity [39], glucose uptake [45], pro-angiogenesis [36,37], antiangiogenesis [19,44], or estrogenic effects [52,56]. Regarding negative control conditions, most of the studies reported the testing of solvent controls in the same concentration as for the respective sample testing [19,32,34,35,[38][39][40]43,44,47,[49][50][51]53,55]. Some studies have additionally evaluated a medium only condition in addition to the solvent control [36,37,42,54,56,57].…”

“…[49] Antioxidant effects ROS generation, cell death Natural products Plant extracts [35] Estrogenicity GFP induction in tg(cyp19a1b-GFP) Environmental toxicology Sediment extracts [52] vtg1 gene expession by qPCR Environmental toxicology Oil sand process waters [56] Anti-convulsant Locomotor activity, electrographic activity and epileptiform discharges Natural products Plant extracts [39] Inhibition of pentylenetetrazol-induced seizure activity, WISH for brain c-fos expression Natural products Plant extracts [34] Fear behaviour Alarm response, olfactory bulb activation in Tα1:GCaMP2…”