Background— We investigated whether electrical stimulation (STIM) of efferent vagus nerves may suppress nuclear factor (NF)-κB activation and the inflammatory cascade in hemorrhagic (Hem) shock. Methods and Results— Rats were subjected to bilateral cervical vagotomy (VGX) or sham surgical procedures. Hem shock was induced by intermittent withdrawing of blood until mean arterial pressure stabilized within the range of 35 to 40 mm Hg. Application of constant voltage pulses to the caudal vagus ends (STIM; 5 V, 2 ms, 1 Hz for 12 minutes, 5 minutes after mean arterial pressure stabilization) increased survival time (VGX+Hem+Sham STIM=38±3 minutes; VGX+Hem+STIM >180 minutes), reverted the marked hypotension (VGX+Hem+Sham STIM=33±3 mm Hg; VGX+Hem+STIM=66±5 mm Hg), inhibited IκBα liver loss, and blunted the augmented NF-κB activity, decreased hepatic tumor necrosis factor (TNF)-α mRNA (VGX+Hem+Sham STIM=1.42±0.5 amount of TNF-α m-RNA; VGX+Hem+STIM=0.51±0.2 amount of TNF-α mRNA), and reduced plasma TNF-α (VGX+Hem+Sham STIM=190±24 pg/mL; VGX+Hem+STIM=87±15 pg/mL). Chlorisondamine, a nicotinic receptor antagonist, abated the effects of vagal stimulation. Conclusions— Our results show a parasympathetic inhibition of NF-κB by which the brain opposes NF-κB activation in the liver and modulates the inflammatory response during acute hypovolemic hemorrhagic shock.

A vagus nerve–mediated, brain cholinergic protective mechanism activated by melanocortin peptides is operative in conditions of circulatory shock; moreover, there is anatomical evidence of dual vagal-cardiac efferent pathways in rats, which could play different roles in controlling heart function. Therefore, we investigated the role and functional mechanism of such vagal efferent pathway(s) in an experimental model of ischemic heart disease. Randomized experimental study. Research laboratory. Adult Wistar rats of either sex. After bilateral cervical vagotomy (with or without pretreatment with atropine), efferent vagal fibers were electrically stimulated in rats subjected to coronary artery occlusion (5 mins) followed by reperfusion (5 mins). Other rats (intact, vagotomized, or pretreated with atropine) were treated with nanomolar doses of melanocortin peptides. Electrical stimulation of efferent vagal fibers (5 V, 2 msecs, 1–9 Hz, for the whole period of ischemia/reperfusion) strongly reduced the high incidence of severe arrhythmias and lethality, reduced the increase in free radical blood levels and left-ventricle histologic alterations, and augmented the extracellular signal–regulated kinase activation. Treatment with the melanocortin peptides adrenocorticotropin and γ2-melanocyte-stimulating hormone (162 nmol/kg intravenously or 16.2 nmol/kg intracerebroventricularly, during coronary occlusion) produced the same protective effects of electrical stimulation and with the same muscarinic acetylcholine receptor–dependent mechanism, seemingly through brain activation (mediated by melanocortin MC3 receptors, as previously described) of such efferent vagal pathway. The present results give evidence for the identification of a protective, melanocortin-activated, efferent vagal cholinergic pathway, operative in conditions of myocardial ischemia/reperfusion. These data suggest that melanocortins and pertinent compounds able to activate such a pathway could provide the potential for development of a new class of drugs for a novel approach to management of ischemic heart disease.

Several melanocortin peptides have a prompt and sustained resuscitating effect in conditions of hemorrhagic shock. The transcription nuclear factor kappaB (NF-kappaB) triggers a potentially lethal systemic inflammatory response, with marked production of tumor necrosis factor-alpha (TNF-alpha), in hemorrhagic shock. Here we investigated whether the hemorrhagic shock reversal produced by the melanocortin ACTH-(1-24) (adrenocorticotropin) depends on the activation of the recently recognized, vagus nerve-mediated, brain "cholinergic anti-inflammatory pathway". Anesthetized rats were stepwise bled until mean arterial pressure (MAP) stabilized at 20-25 mm Hg. The severe hypovolemia was incompatible with survival, and all saline-treated animals died within 30 min. In rats intravenously (i.v.) treated with ACTH-(1-24), neural efferent activity along vagus nerve (monitored by means of a standard system for extracellular recordings) was markedly increased, and the restoration of cardiovascular and respiratory functions was associated with blunted NF-kappaB activity and with decreased TNF-alpha mRNA liver content and TNF-alpha plasma levels. Bilateral cervical vagotomy, pretreatment with the melanocortin MC(4) receptor antagonist HS014, atropine sulfate or chlorisondamine, but not with atropine methylbromide, prevented the life-saving effect of ACTH-(1-24) and the associated effects on NF-kappaB activity and TNF-alpha levels. HS014 and atropine sulfate prevented, too, the ACTH-(1-24)-induced increase in neural efferent vagal activity, and accelerated the evolution of shock in saline-treated rats. The present data show, for the first time, that the melanocortin ACTH-(1-24) suppresses the NF-kappaB-dependent systemic inflammatory response triggered by hemorrhage, and reverses shock condition, by brain activation (in real-time) of the "cholinergic anti-inflammatory pathway", this pathway seeming to be melanocortin-dependent.

Ischemic stroke is one of the main causes of death and disability. We investigated whether melanocortin peptides, which have protective effects in severe hypoxic conditions, also produce neuroprotection in a gerbil model of ischemic stroke. A 10-min period of global cerebral ischemia, induced by occluding both common carotid arteries, caused impairment in spatial learning and memory that was associated with activation of inflammatory and apoptotic pathways, including severe DNA damage and delayed neuronal death, in the hippocampus. Treatment with nanomolar doses of the melanocortin analog [Nle⁴, d-Phe⁷] α-MSH [which activates the melanocortin receptor subtypes (MC) mainly expressed in central nervous system, namely MC₃ and MC₄] modulated the inflammatory and apoptotic cascades and reduced hippocampus injuries even when delayed up to 9 h after ischemia, with consequent dose-dependent improvement in subsequent functional recovery. The selective MC₃ receptor agonist γ₂-MSH had no protective effects. Pharmacological blockade of MC₄ receptors prevented the neuroprotective effects of [Nle⁴, d-Phe⁷] α-MSH and worsened some ischemia outcomes. Together, our findings suggest that MC₄ receptor-stimulating melanocortins might provide potential to develop a class of drugs with a broad treatment window for a novel approach to neuroprotection in ischemic stroke.

The cholinergic anti-inflammatory pathway has not yet been studied in splanchnic artery occlusion (SAO) shock. We investigated whether electrical stimulation (STIM) of efferent vagus nerves suppresses the inflammatory cascade in SAO shock. Animals were subjected to clamping of the splanchnic arteries for 45 min, followed by reperfusion. This surgical procedure resulted in an irreversible state of shock (SAO shock). Sham-operated animals were used as controls. Two minutes before the start of reperfusion, rats were subjected to bilateral cervical vagotomy (VGX) or sham surgical procedures. Application of constant voltage pulses to the caudal vagus ends (STIM: 5 V, 2 ms, 6 Hz for 15 min, 5 min after the beginning of reperfusion) increased survival rate (VGX + SAO + Sham STIM = 0% at 4 h of reperfusion; VGX + SAO + STIM = 90% at 4 h of reperfusion), reverted the marked hypotension, inhibited IκBα liver loss, blunted the augmented nuclear factor-κB activity, decreased hepatic tumor necrosis factor (TNF)-α mRNA (VGX + SAO + Sham STIM = 1.0 ± 1.9 TNF-α/glyceraldehyde-3-phosphate dehydrogenase ratio; VGX + SAO + STIM = 0.3 ± 0.2 TNF-α/glyceraldehyde-3-phosphate dehydrogenase ratio), reduced plasma TNF-α (VGX + SAO + Sham STIM = 118 ± 19 pg/mL; VGX + SAO + STIM = 39 ± 8 pg/mL), ameliorated leukopenia, and decreased leukocyte accumulation, as revealed by means of myeloperoxidase activity in the ileum (VGX + SAO + Sham STIM = 7.9 ± 1 U/g tissue; VGX + SAO + STIM = 3.1 ± 0.7 U/g tissue) and in the lung (VGX + SAO + Sham STIM = 8.0 ± 1.0 U/g tissue; VGX + SAO + STIM = 3.2 ± 0.6 U/g tissue). Chlorisondamine, a nicotinic receptor antagonist, abated the effects of vagal stimulation. Our results show a parasympathetic inhibition of nuclear factor-κB and TNF-α in SAO shock.