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Non-cell autonomous influence of the astrocyte system xc- on hypoglycaemic neuronal cell death.

Jackman NA, Melchior SE, Hewett JA, Hewett SJ - ASN Neuro (2012)

Bottom Line: Further, a dramatic reduction in neuronal death is observed in chimaeric cultures consisting of neurons derived from WT (wild-type) mice plated on top of astrocytes derived from sut mice, which harbour a naturally occurring mutation in the gene (Slc7a11) that encodes the substrate-specific light chain of system xc- (xCT).Finally, enhancement of astrocytic system xc- expression and function via IL-1β (interleukin-1β) exposure potentiates hypoglycaemic neuronal death, the process of which is prevented by removal of l-cystine and/or addition of system xc- inhibitors.Thus, under the conditions of GD, our studies demonstrate that astrocytes, via system xc-, have a direct, non-cell autonomous effect on cortical neuron survival.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, University of Connecticut Health Center, Farmington, CT 06030, USA.

ABSTRACT
Despite longstanding evidence that hypoglycaemic neuronal injury is mediated by glutamate excitotoxicity, the cellular and molecular mechanisms involved remain incompletely defined. Here, we demonstrate that the excitotoxic neuronal death that follows GD (glucose deprivation) is initiated by glutamate extruded from astrocytes via system xc---an amino acid transporter that imports L-cystine and exports L-glutamate. Specifically, we find that depriving mixed cortical cell cultures of glucose for up to 8 h injures neurons, but not astrocytes. Neuronal death is prevented by ionotropic glutamate receptor antagonism and is partially sensitive to tetanus toxin. Removal of amino acids during the deprivation period prevents--whereas addition of L-cystine restores--GD-induced neuronal death, implicating the cystine/glutamate antiporter, system xc-. Indeed, drugs known to inhibit system xc- ameliorate GD-induced neuronal death. Further, a dramatic reduction in neuronal death is observed in chimaeric cultures consisting of neurons derived from WT (wild-type) mice plated on top of astrocytes derived from sut mice, which harbour a naturally occurring mutation in the gene (Slc7a11) that encodes the substrate-specific light chain of system xc- (xCT). Finally, enhancement of astrocytic system xc- expression and function via IL-1β (interleukin-1β) exposure potentiates hypoglycaemic neuronal death, the process of which is prevented by removal of l-cystine and/or addition of system xc- inhibitors. Thus, under the conditions of GD, our studies demonstrate that astrocytes, via system xc-, have a direct, non-cell autonomous effect on cortical neuron survival.

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Hypoglycaemic neuronal cell death is attenuated by glutamate receptor antagonismMixed cortical cultures were washed into a BSS0 containing vehicle, MK-801 (10 μM) or MK-801 plus CNQX (6-cyano-7-nitroquinoxaline-2,3-dione; 30 μM) for 8 h (GD). Neuronal cell death was determined 20–24 h later. (*) Indicates values significantly different from control conditions ( = 10.56±3.02%) determined 24 h following wash of cells into BSS0 followed by immediate addition of glucose. (#) Represents a significant diminution of GD-induced cell death as determined by one-way ANOVA followed by Student–Newman–Keul's post-hoc test (n = 11 cultures from three experiments).
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Figure 2: Hypoglycaemic neuronal cell death is attenuated by glutamate receptor antagonismMixed cortical cultures were washed into a BSS0 containing vehicle, MK-801 (10 μM) or MK-801 plus CNQX (6-cyano-7-nitroquinoxaline-2,3-dione; 30 μM) for 8 h (GD). Neuronal cell death was determined 20–24 h later. (*) Indicates values significantly different from control conditions ( = 10.56±3.02%) determined 24 h following wash of cells into BSS0 followed by immediate addition of glucose. (#) Represents a significant diminution of GD-induced cell death as determined by one-way ANOVA followed by Student–Newman–Keul's post-hoc test (n = 11 cultures from three experiments).

Mentions: As reported previously, selective neuronal degeneration occurred in a time-dependent manner in mixed cortical cell culture following GD (Figure 1), whereas purified astrocytes (Figure 1, inset) and astrocytes in mixed cultures (Figures 4b and 4d) were resistant for up to 8 h, the longest time-point assessed (Monyer and Choi, 1988; Monyer et al., 1989; Goldberg and Choi, 1993). Also in agreement with previous studies (Monyer and Choi, 1988; Monyer et al., 1989), neuronal injury was prevented by ionotropic glutamate receptor antagonism (Figure 2). The small amount of death that is found in control cultures (1–10%) reflects LDH release that results from the extensive washing (i.e. wash damage) that is used to render the cultures aglycaemic.


Non-cell autonomous influence of the astrocyte system xc- on hypoglycaemic neuronal cell death.

Jackman NA, Melchior SE, Hewett JA, Hewett SJ - ASN Neuro (2012)

Hypoglycaemic neuronal cell death is attenuated by glutamate receptor antagonismMixed cortical cultures were washed into a BSS0 containing vehicle, MK-801 (10 μM) or MK-801 plus CNQX (6-cyano-7-nitroquinoxaline-2,3-dione; 30 μM) for 8 h (GD). Neuronal cell death was determined 20–24 h later. (*) Indicates values significantly different from control conditions ( = 10.56±3.02%) determined 24 h following wash of cells into BSS0 followed by immediate addition of glucose. (#) Represents a significant diminution of GD-induced cell death as determined by one-way ANOVA followed by Student–Newman–Keul's post-hoc test (n = 11 cultures from three experiments).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3275339&req=5

Figure 2: Hypoglycaemic neuronal cell death is attenuated by glutamate receptor antagonismMixed cortical cultures were washed into a BSS0 containing vehicle, MK-801 (10 μM) or MK-801 plus CNQX (6-cyano-7-nitroquinoxaline-2,3-dione; 30 μM) for 8 h (GD). Neuronal cell death was determined 20–24 h later. (*) Indicates values significantly different from control conditions ( = 10.56±3.02%) determined 24 h following wash of cells into BSS0 followed by immediate addition of glucose. (#) Represents a significant diminution of GD-induced cell death as determined by one-way ANOVA followed by Student–Newman–Keul's post-hoc test (n = 11 cultures from three experiments).
Mentions: As reported previously, selective neuronal degeneration occurred in a time-dependent manner in mixed cortical cell culture following GD (Figure 1), whereas purified astrocytes (Figure 1, inset) and astrocytes in mixed cultures (Figures 4b and 4d) were resistant for up to 8 h, the longest time-point assessed (Monyer and Choi, 1988; Monyer et al., 1989; Goldberg and Choi, 1993). Also in agreement with previous studies (Monyer and Choi, 1988; Monyer et al., 1989), neuronal injury was prevented by ionotropic glutamate receptor antagonism (Figure 2). The small amount of death that is found in control cultures (1–10%) reflects LDH release that results from the extensive washing (i.e. wash damage) that is used to render the cultures aglycaemic.

Bottom Line: Further, a dramatic reduction in neuronal death is observed in chimaeric cultures consisting of neurons derived from WT (wild-type) mice plated on top of astrocytes derived from sut mice, which harbour a naturally occurring mutation in the gene (Slc7a11) that encodes the substrate-specific light chain of system xc- (xCT).Finally, enhancement of astrocytic system xc- expression and function via IL-1β (interleukin-1β) exposure potentiates hypoglycaemic neuronal death, the process of which is prevented by removal of l-cystine and/or addition of system xc- inhibitors.Thus, under the conditions of GD, our studies demonstrate that astrocytes, via system xc-, have a direct, non-cell autonomous effect on cortical neuron survival.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, University of Connecticut Health Center, Farmington, CT 06030, USA.

ABSTRACT
Despite longstanding evidence that hypoglycaemic neuronal injury is mediated by glutamate excitotoxicity, the cellular and molecular mechanisms involved remain incompletely defined. Here, we demonstrate that the excitotoxic neuronal death that follows GD (glucose deprivation) is initiated by glutamate extruded from astrocytes via system xc---an amino acid transporter that imports L-cystine and exports L-glutamate. Specifically, we find that depriving mixed cortical cell cultures of glucose for up to 8 h injures neurons, but not astrocytes. Neuronal death is prevented by ionotropic glutamate receptor antagonism and is partially sensitive to tetanus toxin. Removal of amino acids during the deprivation period prevents--whereas addition of L-cystine restores--GD-induced neuronal death, implicating the cystine/glutamate antiporter, system xc-. Indeed, drugs known to inhibit system xc- ameliorate GD-induced neuronal death. Further, a dramatic reduction in neuronal death is observed in chimaeric cultures consisting of neurons derived from WT (wild-type) mice plated on top of astrocytes derived from sut mice, which harbour a naturally occurring mutation in the gene (Slc7a11) that encodes the substrate-specific light chain of system xc- (xCT). Finally, enhancement of astrocytic system xc- expression and function via IL-1β (interleukin-1β) exposure potentiates hypoglycaemic neuronal death, the process of which is prevented by removal of l-cystine and/or addition of system xc- inhibitors. Thus, under the conditions of GD, our studies demonstrate that astrocytes, via system xc-, have a direct, non-cell autonomous effect on cortical neuron survival.

Show MeSH
Related in: MedlinePlus