Electrophilic compounds are a newly recognized class of redox‐active neuroprotective compounds with electron deficient, electrophilic carbon centers that react with specific cysteine residues on targeted proteins via thiol (S‐)alkylation. Although plants produce a variety of physiologically active electrophilic compounds, the detailed mechanism of action of these compounds remains unknown. Catechol ring‐containing compounds have attracted attention because they become electrophilic quinones upon oxidation, although they are not themselves electrophilic. In this study, we focused on the neuroprotective effects of one such compound, carnosic acid (CA), found in the herb rosemary obtained from Rosmarinus officinalis. We found that CA activates the Keap1/Nrf2 transcriptional pathway by binding to specific Keap1 cysteine residues, thus protecting neurons from oxidative stress and excitotoxicity. In cerebrocortical cultures, CA‐biotin accumulates in non‐neuronal cells at low concentrations and in neurons at higher concentrations. We present evidence that both the neuronal and non‐neuronal distribution of CA may contribute to its neuroprotective effect. Furthermore, CA translocates into the brain, increases the level of reduced glutathione in vivo, and protects the brain against middle cerebral artery ischemia/reperfusion, suggesting that CA may represent a new type of neuroprotective electrophilic compound.

Electrophilic neurite outgrowth-promoting prostaglandin (NEPP) compounds protect neurons from oxidative insults. At least part of the neuroprotective action of NEPPs lies in induction of hemeoxygenase-1 (HO-1), which, along with other phase II enzymes, serve as a defense system against oxidative stress. Here, we found that, by using fluorescent tags and immunoprecipitation assays, NEPPs are taken up preferentially into neurons and bind in a thiol-dependent manner to Keap1, a negative regulator of the transcription factor Nrf2. By binding to Keap1, NEPPs prevent Keap1-mediated inactivation of Nrf2 and, thus, enhance Nrf2 translocation into the nucleus of cultured neuronal cells. In turn, Nrf2 binds to antioxidant͞ electrophile-responsive elements of the HO-1 promoter to induce HO-1 expression. Consistent with this notion, NEPP induction of an HO-1 reporter construct is prevented if the antioxidant-responsive elements are mutated. We show that NEPPs are neuroprotective both in vitro from glutamate-related excitotoxicity and in vivo in a model of cerebral ischemia͞reperfusion injury (stroke). Our results suggest that NEPPs prevent excitotoxicity by activating the Keap1͞ Nrf2͞HO-1 pathway. Because NEPPs accumulate preferentially in neurons, they may provide a category of neuroprotective compounds, distinct from other electrophilic compounds such as tertbutylhydroquinone, which activates the antioxidant-responsive element in astrocytes. NEPPs thus represent a therapeutic approach for stroke and neurodegenerative disorders.hemeoxygenase-1 ͉ middle cerebral artery occlusion ͉ neurite outgrowth-promoting prostaglandin ͉ stroke ͉ neurodegenerative diseases