Glycine inhibits the LPS-induced increase in cytosolic Ca^2＋ concentration and TNFα production in cardiomyocytes by activating a glycine receptor

Aim： Previous studies have demonstrated that glycine （GLY） markedly reduces lipopolysaccharide （LPS）-induced myocardial injury. However, the mechanism of this effect is still unclear. The present study investigated the effect of GLY on cytosolic calcium concentration （[Ca^2＋]c） and tumor necrosis factor-α（TNFα） production in cardiomyocytes exposed to LPS, as well as whether the glycine-gated chloride channel is involved in this process. Methods： Neonatal rat cardiomyocytes were isolated, and the [Ca^2＋]c and TNFα levels were determined by using Fura-2 and a Quan tikine enzyme-linked immunosorbent assay, respectively. The distribution of the GLY receptor and GLY-induced currents in cardiomyocytes were also investigated using immunocytochemistry and the whole-cell patch-clamp technique, respectively. Results： LPS at concentrations ranging from 10 ng/mL to 100 pg/mL significantly stimulated TNFα production. GLY did not inhibit TNFα production induced by LPS at concentrations below 10 ng/mL but did significantly decrease TNFα release stimulated by 100 pg/mL LPS and prevented an LPS-induced increase in [Ca^2＋]c, which was reversed by strychnine, a glycine receptor antagonist. GLY did not block the isoproterenol-induced increase in [Ca^2＋]c, but did prevent the potassium chloride-induced increase in [Ca2＋]cin cardiomyocytes. Strychnine reversed the inhibition of the KCl-stimulated elevation in [Ca^2＋]cby GLY. In chloride-free buffer, GLY had no effect on the dipotassium hydrogen phosphate-induced increase in [Ca^2＋]c. Furthermore, GLY receptor α1 and β subunitimmunoreactive spots were observed in cardiomyocytes, and GLY-evoked currents were blocked by strychnine. Conclusion： Cardiomyocytes possess the glycine-gated chloride channel, through which GLY prevents the increase in [Ca^2＋]c and inhibits the TNFα production induced by LPS at high doses in neonatal rat cardiomyocytes.