Endogenous CGRP activates NRF2 signaling via non-electrophilic mechanisms

The transcription factor nuclear factor erythroid 2-related factor 2 (NRF2) is crucial for regulating cellular responses to oxidative stress, making it a significant target for therapeutic interventions. While exogenous NRF2 activators offer significant therapeutic potential, their predominantly electrophilic nature poses considerable challenges for clinical use; the heightened electrophilic reactivity required to achieve therapeutic efficacy raises potential safety concerns. Calcitonin gene-related peptide (CGRP) has shown protective effects against oxidative stress and is involved in NRF2 activation; however, the underlying mechanisms are not fully understood. This study explores the mechanisms underlying endogenous CGRP-mediated NRF2 upregulation by inducing acute or chronic CGRP release through diving reflex (DR) in male Sprague-Dawley rats. Brain tissue proteomics confirmed the upregulation of NRF2-dependent antioxidant transcripts-- predominantly glutathione-related genes--without concurrent elevation of oxidative stress markers in both acute and chronic CGRP exposure paradigms. CGRP potently activated NRF2 in brain and peripheral tissues, evidenced by elevated nuclear and phosphorylated NRF2, increased nuclear:cytosolic NRF2 ratios, and enhanced antioxidant gene transcription--effects substantially attenuated by CGRP antagonism. Reduced glutathione levels increased without concurrent elevations in lipid peroxidation, protein oxidation, or evidence of tissue damage, suggesting CGRP avoids side effects characteristic of electrophilic NRF2 activators. Furthermore, our findings suggest that CGRP-mediated NRF2 activation primarily occurs via non-electrophilic mechanisms, with the p62-KEAP1-NRF2 pathway predominantly active in peripheral organs (lung and kidney), and the AMPK-NRF2 pathway more pronounced in the brain, highlighting the organ-specific nature of the response. Time-dependent variations in CGRP-mediated NRF2 activation were also observed, influencing both the response to CGRP and its impact on oxidative stress resistance. These results suggest that targeting NRF2 with endogenous CGRP may offer a promising therapeutic approach for managing oxidative stress-related diseases, both acute and chronic, across multiple organs, by avoiding electrophilic stress. HighlightsO_LIEndogenous CGRP triggers a potent and non-electrophilic activation of NRF2 signaling. C_LIO_LICGRP increases reduced glutathione levels following both acute and chronic exposures, in contrast to the effects of exogenous electrophilic NRF2 activators. C_LIO_LIIn peripheral organs, CGRP predominantly activates the KEAP1-dependent p62-KEAP1-NRF2 pathway. C_LIO_LIIn the brain, CGRP primarily activates the KEAP1-independent AMPK-NRF2 pathway. C_LIO_LICGRP exhibits time-dependent patterns, where acute exposure leads to a more significant upregulation of NRF2-targeted antioxidative gene expression and chronic exposure confers increased resistance to oxidative stress. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=166 SRC="FIGDIR/small/650677v1_ufig8.gif" ALT="Figure 8"> View larger version (44K): org.highwire.dtl.DTLVardef@ae52a3org.highwire.dtl.DTLVardef@b35d1eorg.highwire.dtl.DTLVardef@1a68f31org.highwire.dtl.DTLVardef@31234_HPS_FORMAT_FIGEXP M_FIG C_FIG