Effects of anesthetic regimes on inflammatory responses in a rat model of acute lung injury

Abstract

Purpose: Gamma-aminobutyric acid (GABA) is the major inhibitory neurotransmitter through activation of GABA receptors. Volatile anesthetics activate type-A (GABA(A)) receptors resulting in inhibition of synaptic transmission. Lung epithelial cells have been recently found to express GABA(A) receptors that exert anti-inflammatory properties. We hypothesized that the volatile anesthetic sevoflurane (SEVO) attenuates lung inflammation through activation of lung epithelial GABA(A) receptors.

Methods: Sprague-Dawley rats were anesthetized with SEVO or ketamine/xylazine (KX). Acute lung inflammation was induced by intratracheal instillation of endotoxin, followed by mechanical ventilation for 4 h at a tidal volume of 15 mL/kg without positive end-expiratory pressure (two-hit lung injury model). To examine the specific effects of GABA, healthy human lung epithelial cells (BEAS-2B) were challenged with endotoxin in the presence and absence of GABA with and without addition of the GABA(A) receptor antagonist picrotoxin.

Results: Anesthesia with SEVO improved oxygenation and reduced pulmonary cytokine responses compared to KX. This phenomenon was associated with increased expression of the π subunit of GABA(A) receptors and glutamic acid decarboxylase (GAD). The endotoxin-induced cytokine release from BEAS-2B cells was attenuated by the treatment with GABA, which was reversed by the administration of picrotoxin.

Conclusion: Anesthesia with SEVO suppresses pulmonary inflammation and thus protects the lung from the two-hit injury. The anti-inflammatory effect of SEVO is likely due to activation of pulmonary GABA(A) signaling pathways.

Figure 1. Mean arterial pressure, gas exchange and lung permeability

Mean arterial blood pressure (MAP); arterial partial pressure of carbon dioxide (PaCO₂) and arterial partial pressure of oxygen (PaO₂) in animals anesthetized with ketamine/xylazine (KX) or sevoflurane (SEVO) over time. Lung wet/dry weight ratio was determined at the end of 4-h mechanical ventilation started 30 min after LPS administration. N=17 per group.

Figure 2. Inflammatory mediator responses in BAL fluid

Cytokine and ICAM-1 responses in animals anesthetized with ketamine/xylazine (KX) or sevoflurane (SEVO). Bronchoalveolar lavage (BAL) fluid was obtained at the end of 4-h mechanical ventilation started 30 min after LPS intratracheal administration. Tumor necrosis factor-alpha (TNF-α); macrophage inflammatory protein-1 alpha (MIP-1α); interleukin-1 beta (IL-1β); keratinocyte growth factor (KC); monocyte chemotactic protein-1 (MCP-1) and intracellular adhesion molecule-1 (ICAM-1). N=17 per group.

Figure 3. Lung expression of GAD and GABA_AR receptor

A. Immunohistochemistry staining for the epithelial marker surfactant protein C (green), and GAD (red), GABA_AR (red) and their overlay (orange) in healthy rat lungs. B. Western blot for detection of GAD in animals anesthetized with KX or SEVO at the end of 4-h mechanical ventilation started 30 min after LPS intratracheal administration. Average expression of GAD over β-actin was from 5 experiments.

Figure 4. Cytokine responses in BEAS-2B cells

Concentration of cytokines measured in cell culture supernatants of human bronchial epithelial cells (BEAS-2B) stimulated with LPS (Control) followed by incubation with gamma amino butyric acid (GABA), picrotoxin (PTX) or GABA+PTX for 8h. N = 5 experiments per group.