ABSTRACT

Both nitric oxide synthase (NOS) inhibitors and free radical scavengers have been shown to protect brain tissue in ischemia-reperfusion injury. Nitric oxide and superoxide anion act via distinct mechanisms and react together to form the highly deleterious peroxynitrite. Therefore the authors examined the effects and the interaction between the NOS inhibitor, NG nitro-L-arginine (LNA) and the antioxidant/superoxide scavenger, di-tert-butyl-hydroxybenzoic acid (DtBHB) in the rat submitted to 2 hours of middle cerebral artery occlusion. Posttreatment was initiated 4 hours after the onset of ischemia and infarct volume was measured at 48 hours. The dose-related effect of LNA resulted in a bell-shaped curve: 15, 56, 65, and 33% reduction of total infarct for 0.03, 0.1, 0.3, and 1 mg/kg (intravenously [IV]) respectively and 11% increase in infarct volume for 3 mg/kg (IV). Whereas DtBHB (20 mg/kg; intraperitoneally [IP]) was ineffective, the dose of 60 mg/kg produced 65% protection in infarct volume. The combination of a subthreshold dose of LNA (0.03 mg/kg; IV) and DtBHB (20 mg/kg; IP) resulted in significant reduction (49%) in infarct volume. These results show that LNA and DtBHB act synergistically to provide a consistent neuroprotection against ischemic injury when administered 4 hours after ischemia. This suggests that nitric oxide and free radicals are involved and interact in synergy in ischemia-reperfusion injury.

Botulinum neurotoxin (BoNT) is a major therapeutic agent. Of seven native BoNT serotypes (A to G), only A and B are currently used in the clinic. Here we compared the potency of commercially available purified native serotypes A1 to F1 across in vitro, ex vivo, and in vivo assays. BoNT potency in vitro was assessed in rat primary cells (target protein cleavage and neurotransmitter release assays) in supraspinal, spinal, and sensory systems. BoNT potency ex vivo was measured in the mouse phrenic nerve hemidiaphragm (PNHD) assay, measuring muscle contractility. In vivo, BoNT‐induced muscle relaxation in mice and rats was assessed in the Digit Abduction Score (DAS) test, while effects on body weight (BW) gain were used to assess tolerability. In all assays, all BoNT serotypes were potent toxins, except serotype D1 in vivo which failed to produce significant muscle flaccidity in mice and rats. In rats, all serotypes were well‐tolerated, whereas in mice, reductions in BW were detected at high doses. Serotype A1 was the most potent serotype across in vitro, ex vivo, and in vivo assays. The rank order of potency of the serotypes revealed differences among assays. For example, species‐specificity was seen for serotype B1, and to a lesser extent for serotype C1. Serotypes F1 and C1, not currently in the clinic, showed preference for sensory over motor models and therefore could be considered for development in conditions involving the somatosensory system.

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