Production of superoxide and TNF-α from alveolar macrophages is blunted by glycine

Glycine blunts lipopolysaccharide (LPS)-induced increases in intracellular calcium concentration ([Ca2+](i)) and tumor necrosis factor-alpha (TNF-alpha) production by Kupffer cells through a glycine-gated chloride channel. Alveolar macrophages, which have a similar origin as Kupffer cells, play a significant role in the pathogenesis of several lung diseases including asthma, endotoxemia, and acute inflammation due to inhaled bacterial particles and dusts. Therefore, studies were designed here to test the hypothesis that alveolar macrophages could be inactivated by glycine via a glycine-gated chloride channel: The ability of glycine to prevent endotoxin [lipopolysaccharide (LPS)]-induced increases in [Ca2+](i) and subsequent production of superoxide and TNF-alpha in alveolar macrophages was examined. LPS caused a transient increase in intracellular calcium to nearly 200 nM, with EC50 values slightly greater than 25 ng/ml. Glycine, in a dose-dependent manner, blunted the increase in [Ca2+](i), with an IC50 less than 100 mu M Like the glycine-gated chloride channel in the central nervous system, the effects of glycine on [Ca2+](i) were both strychnine sensitive and chloride dependent. Glycine also caused a dose-dependent influx of radiolabeled chloride with EC50 values near 10 10 mu M, a phenomenon which was also inhibited by strychnine (1 mu M). LPS-induced superoxide production was also blunted in a dose-dependent manner by glycine and was reduced similar to 50% with 10 10 mu M glycine. Moreover, TNF-alpha: production was also inhibited by glycine and also required nearly 10 mu M glycine for half-inhibition. These data provide strong pharmacological evidence that alveolar macrophages contain glycine-gated chloride channels and that their activation is protective against the LPS-induced increase in [Ca2+](i) and subsequent production of toxic radicals and cytokines.