Limits...
Neonatal seizures alter NMDA glutamate receptor GluN2A and 3A subunit expression and function in hippocampal CA1 neurons.

Zhou C, Sun H, Klein PM, Jensen FE - Front Cell Neurosci (2015)

Bottom Line: Moreover, GluN eEPSCs showed a decreased sensitivity to GluN2B selective antagonists and decreased Mg(2+) sensitivity at negative holding potentials, indicating a higher proportion of GluN2A and GluN3A subunit function, respectively.These physiological findings were accompanied by a concurrent increase in GluN2A phosphorylation and GluN3A protein.These results suggest that altered GluN function and expression could potentially contribute to future epileptogenesis following neonatal seizures, and may represent potential therapeutic targets for the blockade of future epileptogenesis in the developing brain.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, Division of Neuroscience, Boston Children's Hospital Boston, MA, USA ; Program in Neurobiology, Harvard Medical School Boston, MA, USA.

ABSTRACT
Neonatal seizures are commonly caused by hypoxic and/or ischemic injury during birth and can lead to long-term epilepsy and cognitive deficits. In a rodent hypoxic seizure (HS) model, we have previously demonstrated a critical role for seizure-induced enhancement of the AMPA subtype of glutamate receptor (GluA) in epileptogenesis and cognitive consequences, in part due to GluA maturational upregulation of expression. Similarly, as the expression and function of the N-Methyl-D-aspartate (NMDA) subtype of glutamate receptor (GluN) is also developmentally controlled, we examined how early life seizures during the critical period of synaptogenesis could modify GluN development and function. In a postnatal day (P)10 rat model of neonatal seizures, we found that seizures could alter GluN2/3 subunit composition of GluNs and physiological function of synaptic GluNs. In hippocampal slices removed from rats within 48-96 h following seizures, the amplitudes of synaptic GluN-mediated evoked excitatory postsynaptic currents (eEPSCs) were elevated in CA1 pyramidal neurons. Moreover, GluN eEPSCs showed a decreased sensitivity to GluN2B selective antagonists and decreased Mg(2+) sensitivity at negative holding potentials, indicating a higher proportion of GluN2A and GluN3A subunit function, respectively. These physiological findings were accompanied by a concurrent increase in GluN2A phosphorylation and GluN3A protein. These results suggest that altered GluN function and expression could potentially contribute to future epileptogenesis following neonatal seizures, and may represent potential therapeutic targets for the blockade of future epileptogenesis in the developing brain.

No MeSH data available.


Related in: MedlinePlus

Hypoxic seizure-induced increases in GluN2A-containing GluN eEPSCs. (A) Representative GluN eEPSC traces before (black) and after bath application of 5 μM ifenprodil (gray) in hippocampus CA1 pyranmidal neurons at +40 mV holding potential from control and 96 h post-HS rats. (B) Group data of ifenprodil sensitivity in CA1 pyramidal neurons in slices from 1–24 h (n = 11 cells), 48–96 h (n = 11 cells) and 1 week post-HS rats (n = 5 cells) and slices from littermate controls (n = 6–10 cells). *p < 0.05. Error bars indicate S.E.M. (C) Group data of absolute GluN2A-containing GluN eEPSC amplitude (ifenprodil insensitive component) in CA1 pyramidal neurons in slices from 1–24 h (n = 11 cells), 48–96 h (n = 11 cells) and 1 week post-HS rats (n = 5 cells) and slices from littermate controls (n = 6–10 cells). *p < 0.05. Error bars indicate S.E.M. (D) Group data of absolute GluN2B-containing GluN eEPSC amplitude (ifenprodil sensitive component) in CA1 pyramidal neurons in slices from 1–24 h (n = 11 cells), 48–96 h (n = 11 cells) and 1 week post-HS rats (n = 5 cells) and slices from littermate controls (n = 6–10 cells). Error bars indicate S.E.M.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4585040&req=5

Figure 2: Hypoxic seizure-induced increases in GluN2A-containing GluN eEPSCs. (A) Representative GluN eEPSC traces before (black) and after bath application of 5 μM ifenprodil (gray) in hippocampus CA1 pyranmidal neurons at +40 mV holding potential from control and 96 h post-HS rats. (B) Group data of ifenprodil sensitivity in CA1 pyramidal neurons in slices from 1–24 h (n = 11 cells), 48–96 h (n = 11 cells) and 1 week post-HS rats (n = 5 cells) and slices from littermate controls (n = 6–10 cells). *p < 0.05. Error bars indicate S.E.M. (C) Group data of absolute GluN2A-containing GluN eEPSC amplitude (ifenprodil insensitive component) in CA1 pyramidal neurons in slices from 1–24 h (n = 11 cells), 48–96 h (n = 11 cells) and 1 week post-HS rats (n = 5 cells) and slices from littermate controls (n = 6–10 cells). *p < 0.05. Error bars indicate S.E.M. (D) Group data of absolute GluN2B-containing GluN eEPSC amplitude (ifenprodil sensitive component) in CA1 pyramidal neurons in slices from 1–24 h (n = 11 cells), 48–96 h (n = 11 cells) and 1 week post-HS rats (n = 5 cells) and slices from littermate controls (n = 6–10 cells). Error bars indicate S.E.M.

Mentions: Given that HS-induced changes in GluN eEPSC amplitudes, we next determined whether this was mediated by post-HS alteration in GluN subunits. GluN2A and GluN2B-containing GluNs can be pharmacologically distinguished by sensitivity to blockade by the GluN2B specific blocker ifenprodil (Arrigoni and Greene, 2004). In CA1 pyramidal neurons from hippocampal slices from P12–14 control animals, bath application of ifenprodil (5 μM) significantly decreased GluN eEPSC amplitudes by about 50% (48.62 ± 7.67%, n = 11 cells, Figure 2). In contrast, the ifenprodil sensitivity was significantly lower in CA1 pyramidal neurons in slices from post-HS 48–96 h rats (19.74 ± 4.51% reduction, n = 10 cells, p = 0.009, Figure 2). Moreover, HS subacutely increased the absolute eEPSC amplitude mediated by ifenprodil-insensitive GluN2A-containing GluNs at 48–96 h post-HS (P12–14 control: 15.19 ± 3.18pA; post-HS 48–96 h: 39.37 ± 4.50pA, n = 10–11 cells, p = 0.0004, Figure 2C). In contrast, at this same time point, there were no changes in the amplitude of ifenprodil-sensitive GluN2B-containing GluNs at 48–96 h post-HS (P12–14 control: 12.75 ± 3.20pA; post-HS 48–96 h: 9.66 ± 3.00pA, n = 10–11 cells, p = 0.49, Figure 2D). These data suggest that the absolute increases in GluN2A-containing GluN function, without a change in GluN2B function, underlies the sub-acute post-HS enhancement in GluN function. To confirm our findings with ifenprodil, we also tested eEPSC subunit pharmacology with Ro-25–6981, another selective antagonist blocking GluN2B-containing GluNs (Kark et al., 1995; Oren et al., 1995), and similar findings were seen (at P11 normoxia, eEPSC blockade percentage 44.64 ± 7.05%, n = 5 by ifenprodil (5 μM) vs 36.6 ± 6.34%, n = 3 by Ro-25–6981 (0.8 μM); at P11 HS, eEPSC blockade percentage 20.23 ± 10.99%, n = 6 by ifenprodil vs 18.5 ± 4.36%, n = 3 by Ro-25-6981).


Neonatal seizures alter NMDA glutamate receptor GluN2A and 3A subunit expression and function in hippocampal CA1 neurons.

Zhou C, Sun H, Klein PM, Jensen FE - Front Cell Neurosci (2015)

Hypoxic seizure-induced increases in GluN2A-containing GluN eEPSCs. (A) Representative GluN eEPSC traces before (black) and after bath application of 5 μM ifenprodil (gray) in hippocampus CA1 pyranmidal neurons at +40 mV holding potential from control and 96 h post-HS rats. (B) Group data of ifenprodil sensitivity in CA1 pyramidal neurons in slices from 1–24 h (n = 11 cells), 48–96 h (n = 11 cells) and 1 week post-HS rats (n = 5 cells) and slices from littermate controls (n = 6–10 cells). *p < 0.05. Error bars indicate S.E.M. (C) Group data of absolute GluN2A-containing GluN eEPSC amplitude (ifenprodil insensitive component) in CA1 pyramidal neurons in slices from 1–24 h (n = 11 cells), 48–96 h (n = 11 cells) and 1 week post-HS rats (n = 5 cells) and slices from littermate controls (n = 6–10 cells). *p < 0.05. Error bars indicate S.E.M. (D) Group data of absolute GluN2B-containing GluN eEPSC amplitude (ifenprodil sensitive component) in CA1 pyramidal neurons in slices from 1–24 h (n = 11 cells), 48–96 h (n = 11 cells) and 1 week post-HS rats (n = 5 cells) and slices from littermate controls (n = 6–10 cells). Error bars indicate S.E.M.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4585040&req=5

Figure 2: Hypoxic seizure-induced increases in GluN2A-containing GluN eEPSCs. (A) Representative GluN eEPSC traces before (black) and after bath application of 5 μM ifenprodil (gray) in hippocampus CA1 pyranmidal neurons at +40 mV holding potential from control and 96 h post-HS rats. (B) Group data of ifenprodil sensitivity in CA1 pyramidal neurons in slices from 1–24 h (n = 11 cells), 48–96 h (n = 11 cells) and 1 week post-HS rats (n = 5 cells) and slices from littermate controls (n = 6–10 cells). *p < 0.05. Error bars indicate S.E.M. (C) Group data of absolute GluN2A-containing GluN eEPSC amplitude (ifenprodil insensitive component) in CA1 pyramidal neurons in slices from 1–24 h (n = 11 cells), 48–96 h (n = 11 cells) and 1 week post-HS rats (n = 5 cells) and slices from littermate controls (n = 6–10 cells). *p < 0.05. Error bars indicate S.E.M. (D) Group data of absolute GluN2B-containing GluN eEPSC amplitude (ifenprodil sensitive component) in CA1 pyramidal neurons in slices from 1–24 h (n = 11 cells), 48–96 h (n = 11 cells) and 1 week post-HS rats (n = 5 cells) and slices from littermate controls (n = 6–10 cells). Error bars indicate S.E.M.
Mentions: Given that HS-induced changes in GluN eEPSC amplitudes, we next determined whether this was mediated by post-HS alteration in GluN subunits. GluN2A and GluN2B-containing GluNs can be pharmacologically distinguished by sensitivity to blockade by the GluN2B specific blocker ifenprodil (Arrigoni and Greene, 2004). In CA1 pyramidal neurons from hippocampal slices from P12–14 control animals, bath application of ifenprodil (5 μM) significantly decreased GluN eEPSC amplitudes by about 50% (48.62 ± 7.67%, n = 11 cells, Figure 2). In contrast, the ifenprodil sensitivity was significantly lower in CA1 pyramidal neurons in slices from post-HS 48–96 h rats (19.74 ± 4.51% reduction, n = 10 cells, p = 0.009, Figure 2). Moreover, HS subacutely increased the absolute eEPSC amplitude mediated by ifenprodil-insensitive GluN2A-containing GluNs at 48–96 h post-HS (P12–14 control: 15.19 ± 3.18pA; post-HS 48–96 h: 39.37 ± 4.50pA, n = 10–11 cells, p = 0.0004, Figure 2C). In contrast, at this same time point, there were no changes in the amplitude of ifenprodil-sensitive GluN2B-containing GluNs at 48–96 h post-HS (P12–14 control: 12.75 ± 3.20pA; post-HS 48–96 h: 9.66 ± 3.00pA, n = 10–11 cells, p = 0.49, Figure 2D). These data suggest that the absolute increases in GluN2A-containing GluN function, without a change in GluN2B function, underlies the sub-acute post-HS enhancement in GluN function. To confirm our findings with ifenprodil, we also tested eEPSC subunit pharmacology with Ro-25–6981, another selective antagonist blocking GluN2B-containing GluNs (Kark et al., 1995; Oren et al., 1995), and similar findings were seen (at P11 normoxia, eEPSC blockade percentage 44.64 ± 7.05%, n = 5 by ifenprodil (5 μM) vs 36.6 ± 6.34%, n = 3 by Ro-25–6981 (0.8 μM); at P11 HS, eEPSC blockade percentage 20.23 ± 10.99%, n = 6 by ifenprodil vs 18.5 ± 4.36%, n = 3 by Ro-25-6981).

Bottom Line: Moreover, GluN eEPSCs showed a decreased sensitivity to GluN2B selective antagonists and decreased Mg(2+) sensitivity at negative holding potentials, indicating a higher proportion of GluN2A and GluN3A subunit function, respectively.These physiological findings were accompanied by a concurrent increase in GluN2A phosphorylation and GluN3A protein.These results suggest that altered GluN function and expression could potentially contribute to future epileptogenesis following neonatal seizures, and may represent potential therapeutic targets for the blockade of future epileptogenesis in the developing brain.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, Division of Neuroscience, Boston Children's Hospital Boston, MA, USA ; Program in Neurobiology, Harvard Medical School Boston, MA, USA.

ABSTRACT
Neonatal seizures are commonly caused by hypoxic and/or ischemic injury during birth and can lead to long-term epilepsy and cognitive deficits. In a rodent hypoxic seizure (HS) model, we have previously demonstrated a critical role for seizure-induced enhancement of the AMPA subtype of glutamate receptor (GluA) in epileptogenesis and cognitive consequences, in part due to GluA maturational upregulation of expression. Similarly, as the expression and function of the N-Methyl-D-aspartate (NMDA) subtype of glutamate receptor (GluN) is also developmentally controlled, we examined how early life seizures during the critical period of synaptogenesis could modify GluN development and function. In a postnatal day (P)10 rat model of neonatal seizures, we found that seizures could alter GluN2/3 subunit composition of GluNs and physiological function of synaptic GluNs. In hippocampal slices removed from rats within 48-96 h following seizures, the amplitudes of synaptic GluN-mediated evoked excitatory postsynaptic currents (eEPSCs) were elevated in CA1 pyramidal neurons. Moreover, GluN eEPSCs showed a decreased sensitivity to GluN2B selective antagonists and decreased Mg(2+) sensitivity at negative holding potentials, indicating a higher proportion of GluN2A and GluN3A subunit function, respectively. These physiological findings were accompanied by a concurrent increase in GluN2A phosphorylation and GluN3A protein. These results suggest that altered GluN function and expression could potentially contribute to future epileptogenesis following neonatal seizures, and may represent potential therapeutic targets for the blockade of future epileptogenesis in the developing brain.

No MeSH data available.


Related in: MedlinePlus