The present findings make a compelling case for the use of resveratrol as a local treatment for both incision induced pain and prevention of chronic pain induced by incision. They show that resveratrol potently and efficaciously inhibits ERK and mTOR signaling in sensory neurons in vitro. The mTOR [39–42] and ERK pathways  have been linked to pathology in multiple preclinical pain models and our previous findings strongly implicate these pathways in the induction of mechanical allodynia by IL-6 and NGF [17, 44] and the development and maintenance of nerve-injury induced allodynia . The pharmacological action of resveratrol observed in our in vitro experiments is linked to activation of AMPK and in vivo effects are consistent with engagement of AMPK. We have previously implicated AMPK activation in alleviation of neuropathic pain , hence, the findings described herein expand the potential clinical usefulness of AMPK activators into the area of post-surgical pain. We conclude that diverse pharmacological mechanisms for activation of AMPK may have utility as novel analgesics for a variety of pain conditions.
Anabolic processes, such as protein synthesis, are orchestrated by upstream kinases that signal to the translation machinery  such as mTOR and ERK. These kinases can be targeted individually by selective inhibitors or they can be negatively modulated by endogenous signaling factors that act on these pathways . A crucial kinase for negative regulation of translation is the ubiquitous, energy-sensing kinase AMPK. Activation of AMPK by depletion of cellular nutrients or through pharmacological intervention results in a dampening of signaling to the translation machinery . This is the natural cellular response to energy deprivation wherein high AMP levels signal to AMPK thereby shutting down anabolic processes when nutrient levels are low. AMPK is not solely regulated by cellular homeostatic mechanisms as it can also be targeted pharmacologically via a number of investigational compounds (e.g. AICAR and A769662 ), natural products (resveratrol [29, 30]) and by the widely clinically available and safe drug metformin [47, 48]. AMPK negatively regulates mTOR via activation of mTOR's negative regulator, TSC2 . This results in a profound inhibition of mTOR and its downstream targets involved in translation control (e.g. 4EBP and ribosomal S6 kinase and rS6p ). Activation of AMPK also negatively regulates ERK activity induced by growth factors and cytokines . This likely occurs via phosphorylation of the insulin receptor substrate 1 (IRS1) protein at Serine 789 . IRS-1 is a critical component of the signaling module of all tyrosine kinase receptors (Trks) and is linked to GP130 (the IL-6 signal transduction receptor) signaling . This interaction may explain the inhibitory effect of resveratrol on IL-6-mediated ERK/eIF4E signaling observed here. Hence, engaging AMPK with potent activators of this pathway (e.g. resveratrol) represents a unique opportunity to achieve inhibition of pain-related signaling because it harnesses the cell's natural mechanism for dampening signaling in two pathways strongly implicated in pain amplification in the periphery, ERK  and mTOR [18, 39–42].
Accordingly, we hypothesized that AMPK activators may represent a novel tool for the treatment of post-surgical pain. We chose to focus on resveratrol for these experiments because resveratrol is a potent and efficacious activator of AMPK . Our findings clearly demonstrate that local application of resveratrol to the site of incision reduces mechanical allodynia, and, importantly, prevents the transition to a chronic pain-like state as measured by PGE2 precipitated persistent nociceptive sensitization. These findings are consistent with previous experiments where we have shown that inhibition of translation regulation signaling during the initiation of allodynia induced by IL-6 or IL-6 and NGF prevents the development of persistent nociceptive sensitization, which, importantly, can be precipitated by a variety of stimuli, not solely PGE2 [44, 52]. In fact, precipitation of persistent nociceptive sensitization following incision can be induced by administration of opioid antagonists suggesting that precipitation does not even require subsequent injury . Moreover, we have recently shown that AMPK activators reduce peripheral nerve injury-induced allodynia and decrease excitability of sensory neurons in vitro . While here, and in our previous work , we have largely ascribed the effects of AMPK activators to sensory neurons, we cannot rule out potential effects on other cell types in the behavioral effects observed. These findings collectively create a compelling case for the further exploration and development of AMPK activators for the treatment of post-surgical pain.
While the pharmacological action of resveratrol has been an area of controversy, most evidence now points to AMPK as the major target of resveratrol. As described above, much attention was originally paid to resveratrol as an activator of sirtuins, in particular sirt1. However, subsequent studies have questioned these original results and recent studies in transgenic animals point to AMPK as a requisite component of resveratrol signaling . Resveratrol, stimulates AMPK in a liver kinase B1 (LKB1) -dependent fashion, similar to the upstream activation of AMPK by metformin [30, 54]. We found that resveratrol stimulates AMPK in TG neurons in a concentration- and time-dependent fashion and that this AMPK activation is correlated with decreased ERK and mTOR signaling, events that we have previously shown are stimulated by other AMPK activators in TG and DRG neurons [17, 18]. Inhibition or activation of sirt1 failed to inhibit or recapitulate the effects of resveratrol, respectively, ruling out an effect of sirtuins in our experiments. Several other mechanisms of action have been ascribed to resveratrol including inhibition of inducible cyclooxygenase  and inhibition of cyclin-dependent kinase 5 . It is unlikely that these mechanisms contribute to the inhibition of ERK and mTOR signaling that we have observed in TG neurons in vitro; however, we cannot exclude the potential contribution of these effects of resveratrol to our behavioral results. Finally, resveratrol has been shown to possess voltage gated-sodium channel inhibition properties . This effect has a slow onset (minutes of drug application is needed), which could be a result of slowly developing direct block of sodium channels, but is more consistent with AMPK activation. In support of the latter conclusion, we have shown that other AMPK activators induce a profound decrease in sensory neuron excitability via a suppression of ramp-current evoked spiking .