The innate immune system protects against infection and tissue injury through the specialized organs of the reticuloendothelial system, including the lungs, liver, and spleen. The central nervous system regulates innate immune responses via the vagus nerve, a mechanism termed the cholinergic antiinflammatory pathway. Vagus nerve stimulation inhibits proinflammatory cytokine production by signaling through the α7 nicotinic acetylcholine receptor subunit. Previously, the functional relationship between the cholinergic antiinflammatory pathway and the reticuloendothelial system was unknown. Here we show that vagus nerve stimulation fails to inhibit tumor necrosis factor (TNF) production in splenectomized animals during lethal endotoxemia. Selective lesioning of the common celiac nerve abolishes TNF suppression by vagus nerve stimulation, suggesting that the cholinergic pathway is functionally hard wired to the spleen via this branch of the vagus nerve. Administration of nicotine, an α7 agonist that mimics vagus nerve stimulation, increases proinflammatory cytokine production and lethality from polymicrobial sepsis in splenectomized mice, indicating that the spleen is critical to the protective response of the cholinergic pathway. These results reveal a specific, physiological connection between the nervous and innate immune systems that may be exploited through either electrical vagus nerve stimulation or administration of α7 agonists to inhibit proinflammatory cytokine production during infection and tissue injury.

The excessive release of cytokines by the immune system contributes importantly to the pathogenesis of inflammatory diseases. Recent advances in understanding the biology of cytokine toxicity led to the discovery of the "cholinergic anti-inflammatory pathway," defined as neural signals transmitted via the vagus nerve that inhibit cytokine release through a mechanism that requires the alpha7 subunit-containing nicotinic acetylcholine receptor (α7nAChR). Vagus nerve regulation of peripheral functions is controlled by brain nuclei and neural networks, but despite considerable importance, little is known about the molecular basis for central regulation of the vagus nerve-based cholinergic anti-inflammatory pathway. Here we report that brain acetylcholinesterase activity controls systemic and organ specific TNF production during endotoxemia. Peripheral administration of the acetylcholinesterase inhibitor galantamine significantly reduced serum TNF levels through vagus nerve signaling, and protected against lethality during murine endotoxemia. Administration of a centrally-acting muscarinic receptor antagonist abolished the suppression of TNF by galantamine, indicating that suppressing acetylcholinesterase activity, coupled with central muscarinic receptors, controls peripheral cytokine responses. Administration of galantamine to α7nAChR knockout mice failed to suppress TNF levels, indicating that the α7nAChR-mediated cholinergic anti-inflammatory pathway is required for the anti-inflammatory effect of galantamine. These findings show that inhibition of brain acetylcholinesterase suppresses systemic inflammation through a central muscarinic receptor-mediated and vagal-and α7nAChR-dependent mechanism. Our data also indicate that a clinically used centrally-acting acetylcholinesterase inhibitor can be utilized to suppress abnormal inflammation to therapeutic advantage.

These findings are of interest for the development of alpha7-nicotinic acetylcholine receptor agonists as a new class of anti-inflammatory therapeutics.

TNF has a critical mediator role in inflammation and is an important therapeutic target. We recently discovered that TNF production is regulated by neural signals through the vagus nerve. Activation of this ''cholinergic antiinflammatory pathway'' inhibits the production of TNF and other cytokines and protects animals from the inflammatory damage caused by endotoxemia and severe sepsis. Here, we describe a role for central muscarinic acetylcholine receptors in the activation of the cholinergic antiinflammatory pathway. Central muscarinic cholinergic activation by muscarine, the M1 receptor agonist McN-A-343, and the M2 receptor antagonist methoctramine inhibited serum TNF levels significantly during endotoxemia. Centrally administered methoctramine stimulated vagus-nerve activity measured by changes in instantaneous heart-rate variability. Blockade of peripheral muscarinic receptors did not abolish antiinflammatory signaling through the vagus nerve, indicating that peripheral muscarinic receptors on immune cells are not required for the cytokine-regulating activities of the cholinergic antiinflammatory pathway. The role of central muscarinic receptors in activating the cholinergic antiinflammatory pathway is of interest for the use of centrally acting muscarinic cholinergic enhancers as antiinflammatory agents. muscarinic receptors ͉ systemic inflammation ͉ TNF ͉ vagus nerve T he overproduction of cytokines, including TNF, mediates tissue damage during the response to injury or pathogenic invasion (1). Physiological and molecular mechanisms that control TNF production maintain cytokine homeostasis and minimize or prevent damage during the host response to infection, injury, arthritis, and other disorders (1-3). Recently, we found (4-6) that electrical or pharmacological activation of the efferent vagus nerve inhibits the release of TNF and attenuates the development of endotoxin-induced shock in rodents. Stimulation of the efferent vagus nerve activates the release of acetylcholine. Although the vagus nerve is a ''classical'' cholinergic regulator of visceral functions in which peripheral muscarinic acetylcholine receptors have a major mediating role, the vagusnerve cytokine-inhibiting activity (which is termed ''the cholinergic antiinflammatory pathway'') requires signaling through nicotinic ␣ 7 subunit-containing receptors (4, 5).Studies have implicated muscarinic receptor-dependent mechanisms in the central modulation and integration of vagal regulation of visceral functions. For example, vagus nerve control of glycogen synthesis in the liver, the Bezold-Jarish cardiovascular reflex and the regulation of exocrine pancreatic secretion are modulated centrally by brain muscarinic receptor mechanisms (7-10). Thus, we reasoned that brain muscarinic receptors could be involved in the central regulation of the vagus-nerve immunomodulatory function. Our results indicate that central cholinergic activation by selective muscarinic receptor ligands significantly inhibits systemic TNF in endotoxemic rats and activates th...

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