Cholinergic anti-inflammatory pathway in intracerebral hemorrhage

Abstract

Stimulated vagus nerve excretes acetylcholine into the peripheral immune organs such as the spleen, reducing innate inflammation. Here, we investigated whether activation of this “cholinergic anti-inflammatory pathway” can be used to reduce cerebral inflammation in a model of hemorrhagic stroke. Experimental intracerebral hemorrhage (ICH) was induced by stereotaxic collagenase injection in rats. Muscarine, an activator of the vagus nerve, or phosphate-buffered saline (control) was injected into the lateral ventricle after induction of ICH. Intraventricular muscarine injection increased heart rate variability in the ICH model, suggesting increased vagus nerve output. Muscarine-injected ICH rats showed improved neurologic outcomes, reduced brain water content, and decreased levels of inflammatory mediators in both brain and spleen. Central muscarine injection was ineffective at reducing cerebral edema without spleen, suggesting that the effect of muscarine is mediated through the vagus nerve-spleen pathway rather than through a direct interaction with the brain. Our results suggest that the brain possesses a cholinergic anti-inflammatory pathway that counteracts the inflammatory responses after ICH, thereby limiting damage to the brain itself.

Introduction

Increased frequency of intracerebral hemorrhage (ICH) is a complication of thrombolytic treatment in ischemic stroke (Lapchak, 2002). ICH induces local tissue deformation and subsequent development of excitotoxicity, apoptosis, and inflammation (Aronowski and Hall, 2005). ICH-induced inflammation, in particular, appears to be a key factor of secondary brain damage, and anti-inflammatory approaches may lessen the outcome of hemorrhagic stroke (Aronowski and Hall, 2005, Chu et al., 2004, Wang and Dore, 2007, Lee et al., 2008). ICH initiates prominent inflammatory responses, which include activation of resident brain microglia and infiltration of circulating inflammatory cells into the brain (Del Bigio et al., 1996). This process is accompanied by the massive and rapid activation of the peripheral immune system as well as the dynamic and widespread activation of inflammatory cytokines in spleen (Offner et al., 2006). The spleen is the key organ that produces tumor necrosis factor-α (TNF-α) and high mobility group box 1 (HMGB1) in response to systemic inflammatory conditions (Huston et al., 2006, Tracey, 2007). TNF-α, interleukin (IL)-1β, and HMGB1 are the major inflammatory mediators in stroke (Zheng and Yenari, 2004, Kim et al., 2006, Faraco et al., 2007), and neutralizing antibody can protect the brain from post-stroke inflammatory injury (Hosomi et al., 2005).

The nervous system can inhibit cytokine release and prevent tissue injury via an efferent neural signaling pathway termed the cholinergic anti-inflammatory pathway (Tracey, 2007). Stimulation of the vagus nerve prevents damage caused by cytokine release in experimental sepsis, endotoxemia, hemorrhagic shock, arthritis, and other inflammatory syndromes (Tracey, 2007, Borovikova et al., 2000, Saeed et al., 2005, van Westerloo et al., 2006). The cholinergic anti-inflammatory pathway is mainly mediated by the spleen. Released acetylcholine inhibits the production of TNF-α and HMGB1 from splenic macrophages by activating α₇ nicotinic receptors, which protects target organs from inflammatory damage (Tracey, 2007). Because stimulation of the vagus nerve is commonly and safely used in the treatment of intractable epilepsy (Nadkarni et al., 2005), cholinergic anti-inflammatory pathway is a feasible candidate for anti-inflammatory therapeutics.

Cholinergic anti-inflammatory pathway can be activated by intraventricular muscarine injection, which lowers serum TNF-α levels in endotoxemia models and increases vagus nerve activity as measured by changes in heart-rate-variability; thus, it serves as a reproducible model for studies on the cholinergic anti-inflammatory pathway (Pavlov et al., 2006). Here, we investigated the presence and physiology of the cholinergic anti-inflammatory pathway after ICH.