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Astrocytes protect neurons against methylmercury via ATP/P2Y(1) receptor-mediated pathways in astrocytes.

Noguchi Y, Shinozaki Y, Fujishita K, Shibata K, Imura Y, Morizawa Y, Gachet C, Koizumi S - PLoS ONE (2013)

Bottom Line: MeHg-treated astrocyte-conditioned medium (ACM) showed neuro-protective effects against MeHg, which was blocked by anti-IL-6 antibody and was mimicked by the application of recombinant IL-6.As for the mechanism of neuro-protection by IL-6, an adenosine A1 receptor-mediated pathway in neurons seems to be involved.Taken together, when astrocytes sense MeHg, they release ATP that autostimulates P2Y1 receptors to upregulate IL-6, thereby leading to A1 receptor-mediated neuro-protection against MeHg.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuropharmacology, Interdisciplinary Graduate School of Medicine and Engineering, University of Yamanashi, Yamanashi, Japan.

ABSTRACT
Methylmercury (MeHg) is a well known environmental pollutant that induces serious neuronal damage. Although MeHg readily crosses the blood-brain barrier, and should affect both neurons and glial cells, how it affects glia or neuron-to-glia interactions has received only limited attention. Here, we report that MeHg triggers ATP/P2Y1 receptor signals in astrocytes, thereby protecting neurons against MeHg via interleukin-6 (IL-6)-mediated pathways. MeHg increased several mRNAs in astrocytes, among which IL-6 was the highest. For this, ATP/P2Y1 receptor-mediated mechanisms were required because the IL-6 production was (i) inhibited by a P2Y1 receptor antagonist, MRS2179, (ii) abolished in astrocytes obtained from P2Y1 receptor-knockout mice, and (iii) mimicked by exogenously applied ATP. In addition, (iv) MeHg released ATP by exocytosis from astrocytes. As for the intracellular mechanisms responsible for IL-6 production, p38 MAP kinase was involved. MeHg-treated astrocyte-conditioned medium (ACM) showed neuro-protective effects against MeHg, which was blocked by anti-IL-6 antibody and was mimicked by the application of recombinant IL-6. As for the mechanism of neuro-protection by IL-6, an adenosine A1 receptor-mediated pathway in neurons seems to be involved. Taken together, when astrocytes sense MeHg, they release ATP that autostimulates P2Y1 receptors to upregulate IL-6, thereby leading to A1 receptor-mediated neuro-protection against MeHg.

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MeHg-induced IL-6 mRNA upregulation and protein release from astrocytes.(A) Effect of MeHg on IL-6 mRNA expression in astrocytes. MeHg (1–3 µM) transiently increased IL-6 expression and the induction peak was observed at 2 hr exposure. Low concentration of MeHg (0.1 µM) had no effect on IL-6 expression. *P<0.05 and **P<0.01 vs. control. (B) MeHg-induced IL-6 protein production to the supernatant from astrocytes. MeHg (1 or 3 µM, 12 or 24 hr) induced IL-6 production. The 12-hr exposure exhibited a lower level of IL-6 release than that with 24-hr exposure of MeHg.
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pone-0057898-g001: MeHg-induced IL-6 mRNA upregulation and protein release from astrocytes.(A) Effect of MeHg on IL-6 mRNA expression in astrocytes. MeHg (1–3 µM) transiently increased IL-6 expression and the induction peak was observed at 2 hr exposure. Low concentration of MeHg (0.1 µM) had no effect on IL-6 expression. *P<0.05 and **P<0.01 vs. control. (B) MeHg-induced IL-6 protein production to the supernatant from astrocytes. MeHg (1 or 3 µM, 12 or 24 hr) induced IL-6 production. The 12-hr exposure exhibited a lower level of IL-6 release than that with 24-hr exposure of MeHg.

Mentions: We first performed transcriptome analysis in cultured astrocytes stimulated with MeHg (10 µM) using DNA microarray (Table 1). MeHg changed the expressions of a large number of genes including those of cytokines and chaperones in astrocytes. Among them, interleukin-6 (IL-6) mRNA showed the most remarkable increase (638 fold), and we confirmed its upregulation using quantitative RT-PCR. The increase in IL-6 mRNA expression was concentration-dependent over a concentration range of from 0.1 to 3 µM with 2 hr-exposure (0.1 µM, 2.6±0.7; 1.0 µM, 6.1±0.7; 3.0 µM, 30.0±7.2 fold increase vs. control, n = 3) (Fig. 1A). The low concentration of MeHg (0.1 µM) never increased IL-6 mRNA expression at any exposure time tested (1–12 hr) (1 hr, 1.3±0.1; 2 hr, 2.6±0.7; 6 hr, 1.7±0.2; 12 hr, 1.2±0.2 fold increase vs. control, n = 3). The increase in the IL-6 mRNA level was transient and reached the maximal level at 2 hr after the exposure with 1 µM (1 hr, 2.5±0.6; 2 hr, 6.1±0.7; 6 hr, 5.1±2.0; 12 hr, 1.2±0.1 fold increase vs. control, n = 3) and 3 µM (1 hr, 4.3±1.1; 2 hr, 29.9±7.2; 6 hr, 5.1±1.6; 12 hr, 1.9±0.2 fold increase vs. control, n = 3) of MeHg. ELISA analysis of the supernatants showed that MeHg (1 and 3 µM, 24 hr) increased IL-6 derived from astrocytes (1 µM MeHg, 51.5±11.4 pg/ml; 3 µM MeHg, 80.9±18.6 pg/ml, n = 4) (Fig. 1B). With 12-hr exposure, a lower level of IL-6 release was observed (1 µM MeHg, 8.7±5.0 pg/ml; 3 µM MeHg, 30.5±16.9 pg/ml, n = 4). Without MeHg stimulation, no detectable level of IL-6 was observed (n.d.).


Astrocytes protect neurons against methylmercury via ATP/P2Y(1) receptor-mediated pathways in astrocytes.

Noguchi Y, Shinozaki Y, Fujishita K, Shibata K, Imura Y, Morizawa Y, Gachet C, Koizumi S - PLoS ONE (2013)

MeHg-induced IL-6 mRNA upregulation and protein release from astrocytes.(A) Effect of MeHg on IL-6 mRNA expression in astrocytes. MeHg (1–3 µM) transiently increased IL-6 expression and the induction peak was observed at 2 hr exposure. Low concentration of MeHg (0.1 µM) had no effect on IL-6 expression. *P<0.05 and **P<0.01 vs. control. (B) MeHg-induced IL-6 protein production to the supernatant from astrocytes. MeHg (1 or 3 µM, 12 or 24 hr) induced IL-6 production. The 12-hr exposure exhibited a lower level of IL-6 release than that with 24-hr exposure of MeHg.
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pone-0057898-g001: MeHg-induced IL-6 mRNA upregulation and protein release from astrocytes.(A) Effect of MeHg on IL-6 mRNA expression in astrocytes. MeHg (1–3 µM) transiently increased IL-6 expression and the induction peak was observed at 2 hr exposure. Low concentration of MeHg (0.1 µM) had no effect on IL-6 expression. *P<0.05 and **P<0.01 vs. control. (B) MeHg-induced IL-6 protein production to the supernatant from astrocytes. MeHg (1 or 3 µM, 12 or 24 hr) induced IL-6 production. The 12-hr exposure exhibited a lower level of IL-6 release than that with 24-hr exposure of MeHg.
Mentions: We first performed transcriptome analysis in cultured astrocytes stimulated with MeHg (10 µM) using DNA microarray (Table 1). MeHg changed the expressions of a large number of genes including those of cytokines and chaperones in astrocytes. Among them, interleukin-6 (IL-6) mRNA showed the most remarkable increase (638 fold), and we confirmed its upregulation using quantitative RT-PCR. The increase in IL-6 mRNA expression was concentration-dependent over a concentration range of from 0.1 to 3 µM with 2 hr-exposure (0.1 µM, 2.6±0.7; 1.0 µM, 6.1±0.7; 3.0 µM, 30.0±7.2 fold increase vs. control, n = 3) (Fig. 1A). The low concentration of MeHg (0.1 µM) never increased IL-6 mRNA expression at any exposure time tested (1–12 hr) (1 hr, 1.3±0.1; 2 hr, 2.6±0.7; 6 hr, 1.7±0.2; 12 hr, 1.2±0.2 fold increase vs. control, n = 3). The increase in the IL-6 mRNA level was transient and reached the maximal level at 2 hr after the exposure with 1 µM (1 hr, 2.5±0.6; 2 hr, 6.1±0.7; 6 hr, 5.1±2.0; 12 hr, 1.2±0.1 fold increase vs. control, n = 3) and 3 µM (1 hr, 4.3±1.1; 2 hr, 29.9±7.2; 6 hr, 5.1±1.6; 12 hr, 1.9±0.2 fold increase vs. control, n = 3) of MeHg. ELISA analysis of the supernatants showed that MeHg (1 and 3 µM, 24 hr) increased IL-6 derived from astrocytes (1 µM MeHg, 51.5±11.4 pg/ml; 3 µM MeHg, 80.9±18.6 pg/ml, n = 4) (Fig. 1B). With 12-hr exposure, a lower level of IL-6 release was observed (1 µM MeHg, 8.7±5.0 pg/ml; 3 µM MeHg, 30.5±16.9 pg/ml, n = 4). Without MeHg stimulation, no detectable level of IL-6 was observed (n.d.).

Bottom Line: MeHg-treated astrocyte-conditioned medium (ACM) showed neuro-protective effects against MeHg, which was blocked by anti-IL-6 antibody and was mimicked by the application of recombinant IL-6.As for the mechanism of neuro-protection by IL-6, an adenosine A1 receptor-mediated pathway in neurons seems to be involved.Taken together, when astrocytes sense MeHg, they release ATP that autostimulates P2Y1 receptors to upregulate IL-6, thereby leading to A1 receptor-mediated neuro-protection against MeHg.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuropharmacology, Interdisciplinary Graduate School of Medicine and Engineering, University of Yamanashi, Yamanashi, Japan.

ABSTRACT
Methylmercury (MeHg) is a well known environmental pollutant that induces serious neuronal damage. Although MeHg readily crosses the blood-brain barrier, and should affect both neurons and glial cells, how it affects glia or neuron-to-glia interactions has received only limited attention. Here, we report that MeHg triggers ATP/P2Y1 receptor signals in astrocytes, thereby protecting neurons against MeHg via interleukin-6 (IL-6)-mediated pathways. MeHg increased several mRNAs in astrocytes, among which IL-6 was the highest. For this, ATP/P2Y1 receptor-mediated mechanisms were required because the IL-6 production was (i) inhibited by a P2Y1 receptor antagonist, MRS2179, (ii) abolished in astrocytes obtained from P2Y1 receptor-knockout mice, and (iii) mimicked by exogenously applied ATP. In addition, (iv) MeHg released ATP by exocytosis from astrocytes. As for the intracellular mechanisms responsible for IL-6 production, p38 MAP kinase was involved. MeHg-treated astrocyte-conditioned medium (ACM) showed neuro-protective effects against MeHg, which was blocked by anti-IL-6 antibody and was mimicked by the application of recombinant IL-6. As for the mechanism of neuro-protection by IL-6, an adenosine A1 receptor-mediated pathway in neurons seems to be involved. Taken together, when astrocytes sense MeHg, they release ATP that autostimulates P2Y1 receptors to upregulate IL-6, thereby leading to A1 receptor-mediated neuro-protection against MeHg.

Show MeSH
Related in: MedlinePlus