Incorporation of sodium channel blocking and free radical scavenging activities into a single drug, AM-36, results in profound inhibition of neuronalapoptosis

1 AM-36 is a novel neuroprotective agent incorporating both antioxidant and Na+ channel blocking actions. In cerebral ischaemia, loss of cellular ion homeostasis due to Na+ channel activation, together with increased reactive oxygen species (ROS) production, are thought to contribute to neuronal dea th. Since neuronal death in the penumbra of the ischaemic lesion is suggest ed to occur by apoptosis, we investigated the ability of AM-36, antioxidant s and Na+ channel antagonists to inhibit toxicity induced by the neurotoxin , veratridine in cultured cerebellar granule cells (CGC's). 2 Veratridine (10-300 muM) concentration-dependently reduced cell viability of cultured CGC's. Under the experimental conditions employed, cell death induced by veratridine (100 muM) possessed the characteristics of apoptosis as assessed by morphology, TUNEL staining and DNA laddering on agarose gel s. 3 Neurotoxicity and apoptosis induced by veratridine (100 muM) were inhibit ed to a maximum of 50% by the antioxidants, U74500A (0.1 - 10 muM) and U838 36E (0.03 - 10 muM), and to a maximum of 30% by the Na+ channel blocker, di bucaine (0.1-100 muM). In contrast, AM-36 (0.01-10 muM) completely inhibite d veratridine-induced toxicity (IC50 1.7 (1.5-1.9) muM, 95% confidence inte rvals (CI) in parentheses) and concentration-dependently inhibited apoptosi s. 4 These findings suggest veratridine-induced toxicity and apoptosis are par tially mediated by generation of ROS. AM-36, which combines both Na+ channe l blocking and antioxidant activity, provided superior neuroprotection comp ared with agents possessing only one of these actions. This bifunctional pr ofile of activity may underlie the potent neuroprotective effects of AM-36 recently found in a stroke model in conscious rats.