The flavonoid, 2′-methoxy-6-methylflavone, affords neuroprotection following focal cerebral ischaemia

Abstract

Tonic inhibitory currents, mediated by extrasynaptic GABA receptors, are elevated at a delay following stroke. Flavonoids minimise the extent of cellular damage following stroke, but little is known about their mode of action. We demonstrate that the flavonoid, 2'-methoxy-6-methylflavone (0.1-10 µM; 2'MeO6MF), increases GABA receptor tonic currents presumably via δ-containing GABA receptors. Treatment with 2'MeO6MF 1-6 h post focal ischaemia dose dependently decreases infarct volume and improves functional recovery. The effect of 2'MeO6MF was attenuated in δ mice, indicating that the effects of the flavonoid were mediated via δ-containing GABA receptors. Further, as flavonoids have been shown to have multiple modes of action, we investigated the anti-inflammatory effects of 2'MeO6MF. Using a macrophage cell line, we show that 2'MeO6MF can dampen an LPS-induced elevation in NFkB activity. Assessment of vehicle-treated stroke animals revealed a significant increase in circulating IL1β, TNFα and IFγ levels. Treatment with 2'MeO6MF dampened the stroke-induced increase in circulating cytokines, which was blocked in the presence of the pan-AKT inhibitor, GSK690693. These studies support the hypothesis that compounds that potentiate tonic inhibition via δ-containing GABA receptors soon after stroke can afford neuroprotection.

Stroke Induces a BDNF-Dependent Improvement in Cognitive Flexibility in Aged Mice

“…The brains were cut coronally with a section thickness of 40 μ m on a sliding microtome with a freezing stage, with all sections stored in cryoprotectant at -20°C. Infarct volume was determined by histological assessment using a previously published cresyl violet staining protocol [36]. Infarct volume was quantified using ImageJ (National Institutes of Health, USA) by an observer blind as to the treatment groups and was based on obtaining measurements from every 6th section through the entire infarct (area in mm 2 ), with infarct volume being quantified as follows: trueinfarct volume mm3=area mm2×section thickness×section interval.…”

Inhibition of GABA transporters fails to afford significant protection following focal cerebral ischemia

“…7 Prior studies have reported that GABA uptake in the sensorimotor cortex is synergistically regulated by both GAT1 and GAT2/3 function. 7,40 Since tonic inhibition is increased already after one day 41 and GAT3 is impaired from quite early on after a PT stroke, 7 one likely explanation for the strokeinduced seizures observed, is that blocking GAT1, even with low doses of tiagabine, could enhance tonic inhibition to detrimental levels. As not all animals in our study and not all studies have reported increased seizure activity with tiagabine post-stroke, 16,19 it is likely that in animals that showed no seizure activity upon tiagabine treatment that GAT3 is still functional at the time of compound administration.…”

The gliopeptide ODN, a ligand for the benzodiazepine site of GABAA receptors, boosts functional recovery after stroke

“…For recordings from layer 2/3 pyramidal neurons in sham or stroke mice, strokes were induced in 2-3 month-old-mice. The mice were anesthetized with isoflurane and brains isolated at 3-7 days post-stroke or sham surgery and prepared for recordings of tonic currents (Clarkson et al, 2010;Clarkson et al, 2019). All recordings were made from peri-infarct cortical layer 2/3 pyramidal neurons within the primary motor cortex, as previously described (Clarkson et al, 2010;Clarkson et al, 2019).…”

“…The animals were then allowed to recover and were killed 48 hours or at day 28 after MCAO.Cortical photothrombosis. Focal stroke was induced in the left hemisphere using the photothrombosis method in aged (20 ± 2 month old) female C57BL/6J mice weighing 35.8 ± 5.6 g(Clarkson et al, 2010;Clarkson et al, 2011;Clarkson et al, 2019).…”