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The GluN3A subunit exerts a neuroprotective effect in brain ischemia and the hypoxia process.

Wang H, Yan H, Zhang S, Wei X, Zheng J, Li J - ASN Neuro (2013)

Bottom Line: GluN3 subunits, the third member of the NMDAR family with two isoforms, GluN3A and GluN3B, have been confirmed to display an inhibitory effect on NMDAR activity.It was found that GluN3A protein expression in rat hippocampus and the prefrontal cortex was increased quickly after brain ischemia and remained at a high level for at least 24 h.Suppressing the generation of hydroxyl radicals and NO (nitric oxide) is probably also involved in the neuroprotection.

View Article: PubMed Central - PubMed

Affiliation: Beijing Institute of Pharmacology and Toxicology, 27 Taiping Road, Beijing 100850, People's Republic of China.

ABSTRACT
NMDARs (N-methyl-D-aspartate receptors) mediate the predominantly excitatory neurotransmission in the CNS (central nervous system). Excessive release of glutamate and overactivation of NMDARs during brain ischemia and the hypoxia process are causally linked to excitotoxicity and neuronal damage. GluN3 subunits, the third member of the NMDAR family with two isoforms, GluN3A and GluN3B, have been confirmed to display an inhibitory effect on NMDAR activity. However, the effect of GluN3 subunits in brain ischemia and hypoxia is not clearly understood. In the present study, the influence of ischemia and hypoxia on GluN3 subunit expression was observed by using the 2VO (two-vessel occlusion) rat brain ischemia model and cell OGD (oxygen and glucose deprivation) hypoxia model. It was found that GluN3A protein expression in rat hippocampus and the prefrontal cortex was increased quickly after brain ischemia and remained at a high level for at least 24 h. However, the expression of the GluN3B subunit was not remarkably changed in both the animal and cell models. After OGD exposure, rat hippocampal neurons with GluN3A subunit overexpression displayed more viability than the wild-type neurons. NG108-15 cells overexpressing GluN3A presented pronounced resistance to glutamate insult. Blocking the increase of intracellular Ca2+ concentration may underlie the neuroprotective mechanism of up-regulated GluN3A subunit. Suppressing the generation of hydroxyl radicals and NO (nitric oxide) is probably also involved in the neuroprotection.

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FCM results of NG108-15 cell viability after glutamate injury(A) FCM results. The lower left quadrants of each dot blot showed the viable cells (FITC−/PI−). The lower right quadrants represented the apoptotic cells (FITC+/PI−) and the upper right quadrants indicated the necrotic cell population (FITC+/PI+). Markers A1–A4 represented the WT (wild-type), WT+MK-801, GluN3A overexpressing and GluN3A+MK-801 cells, respectively. (B, C) Statistical analysis of the death ratio (B) and apoptosis ratio (C). Data were presented as means±S.E.M., **P<0.01, relative to vector alone (control), ##P<0.01, relative to the group with GluN3A overexpression and without MK-801 pretreatment.
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Figure 7: FCM results of NG108-15 cell viability after glutamate injury(A) FCM results. The lower left quadrants of each dot blot showed the viable cells (FITC−/PI−). The lower right quadrants represented the apoptotic cells (FITC+/PI−) and the upper right quadrants indicated the necrotic cell population (FITC+/PI+). Markers A1–A4 represented the WT (wild-type), WT+MK-801, GluN3A overexpressing and GluN3A+MK-801 cells, respectively. (B, C) Statistical analysis of the death ratio (B) and apoptosis ratio (C). Data were presented as means±S.E.M., **P<0.01, relative to vector alone (control), ##P<0.01, relative to the group with GluN3A overexpression and without MK-801 pretreatment.

Mentions: FCM also indicated a lower death rate among GluN3A transfected cells subjected to glutamate insult (Figure 7). Cell death attributable to glutamate insult in wild-type NG108-15 cells, wild-type NG108-15 cells pretreated with MK-801, NG108-15 cells overexpressing GluN3A, and NG108-15 cells overexpressing GluN3A pretreated with MK-801 were 28.7±1.7, 16.2±1.8, 9.7±1.2 and 8.5±0.8%, respectively. The death rates among the three test groups were all significantly lower than that of the control group (Figure 7B, F=127.4, P<0.01, n=3). The group overexpressing GluN3A with MK-801 pretreatment showed the highest viability. However, the changes of apoptosis ratio displayed a different characteristic (Figure 7C). Only NG108-15 cells overexpressing GluN3A presented an apoptosis ratio (9.3±0.7%) lower than that of control group (16.5±0.5%, P<0.01, n=3). MK-801 had no effect on the apoptosis induced by glutamate insult in NG108-15 cells without overexpressing GluN3A with the ratio of 16.6±0.6%, and even increased the apoptosis ratio of NG108-15 cells overexpressing GluN3A from 9.3±0.7 to 12.4±1.0% (P<0.01).


The GluN3A subunit exerts a neuroprotective effect in brain ischemia and the hypoxia process.

Wang H, Yan H, Zhang S, Wei X, Zheng J, Li J - ASN Neuro (2013)

FCM results of NG108-15 cell viability after glutamate injury(A) FCM results. The lower left quadrants of each dot blot showed the viable cells (FITC−/PI−). The lower right quadrants represented the apoptotic cells (FITC+/PI−) and the upper right quadrants indicated the necrotic cell population (FITC+/PI+). Markers A1–A4 represented the WT (wild-type), WT+MK-801, GluN3A overexpressing and GluN3A+MK-801 cells, respectively. (B, C) Statistical analysis of the death ratio (B) and apoptosis ratio (C). Data were presented as means±S.E.M., **P<0.01, relative to vector alone (control), ##P<0.01, relative to the group with GluN3A overexpression and without MK-801 pretreatment.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3756525&req=5

Figure 7: FCM results of NG108-15 cell viability after glutamate injury(A) FCM results. The lower left quadrants of each dot blot showed the viable cells (FITC−/PI−). The lower right quadrants represented the apoptotic cells (FITC+/PI−) and the upper right quadrants indicated the necrotic cell population (FITC+/PI+). Markers A1–A4 represented the WT (wild-type), WT+MK-801, GluN3A overexpressing and GluN3A+MK-801 cells, respectively. (B, C) Statistical analysis of the death ratio (B) and apoptosis ratio (C). Data were presented as means±S.E.M., **P<0.01, relative to vector alone (control), ##P<0.01, relative to the group with GluN3A overexpression and without MK-801 pretreatment.
Mentions: FCM also indicated a lower death rate among GluN3A transfected cells subjected to glutamate insult (Figure 7). Cell death attributable to glutamate insult in wild-type NG108-15 cells, wild-type NG108-15 cells pretreated with MK-801, NG108-15 cells overexpressing GluN3A, and NG108-15 cells overexpressing GluN3A pretreated with MK-801 were 28.7±1.7, 16.2±1.8, 9.7±1.2 and 8.5±0.8%, respectively. The death rates among the three test groups were all significantly lower than that of the control group (Figure 7B, F=127.4, P<0.01, n=3). The group overexpressing GluN3A with MK-801 pretreatment showed the highest viability. However, the changes of apoptosis ratio displayed a different characteristic (Figure 7C). Only NG108-15 cells overexpressing GluN3A presented an apoptosis ratio (9.3±0.7%) lower than that of control group (16.5±0.5%, P<0.01, n=3). MK-801 had no effect on the apoptosis induced by glutamate insult in NG108-15 cells without overexpressing GluN3A with the ratio of 16.6±0.6%, and even increased the apoptosis ratio of NG108-15 cells overexpressing GluN3A from 9.3±0.7 to 12.4±1.0% (P<0.01).

Bottom Line: GluN3 subunits, the third member of the NMDAR family with two isoforms, GluN3A and GluN3B, have been confirmed to display an inhibitory effect on NMDAR activity.It was found that GluN3A protein expression in rat hippocampus and the prefrontal cortex was increased quickly after brain ischemia and remained at a high level for at least 24 h.Suppressing the generation of hydroxyl radicals and NO (nitric oxide) is probably also involved in the neuroprotection.

View Article: PubMed Central - PubMed

Affiliation: Beijing Institute of Pharmacology and Toxicology, 27 Taiping Road, Beijing 100850, People's Republic of China.

ABSTRACT
NMDARs (N-methyl-D-aspartate receptors) mediate the predominantly excitatory neurotransmission in the CNS (central nervous system). Excessive release of glutamate and overactivation of NMDARs during brain ischemia and the hypoxia process are causally linked to excitotoxicity and neuronal damage. GluN3 subunits, the third member of the NMDAR family with two isoforms, GluN3A and GluN3B, have been confirmed to display an inhibitory effect on NMDAR activity. However, the effect of GluN3 subunits in brain ischemia and hypoxia is not clearly understood. In the present study, the influence of ischemia and hypoxia on GluN3 subunit expression was observed by using the 2VO (two-vessel occlusion) rat brain ischemia model and cell OGD (oxygen and glucose deprivation) hypoxia model. It was found that GluN3A protein expression in rat hippocampus and the prefrontal cortex was increased quickly after brain ischemia and remained at a high level for at least 24 h. However, the expression of the GluN3B subunit was not remarkably changed in both the animal and cell models. After OGD exposure, rat hippocampal neurons with GluN3A subunit overexpression displayed more viability than the wild-type neurons. NG108-15 cells overexpressing GluN3A presented pronounced resistance to glutamate insult. Blocking the increase of intracellular Ca2+ concentration may underlie the neuroprotective mechanism of up-regulated GluN3A subunit. Suppressing the generation of hydroxyl radicals and NO (nitric oxide) is probably also involved in the neuroprotection.

Show MeSH
Related in: MedlinePlus