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Neto2- mice have impaired GABAergic inhibition and are susceptible to seizures.

Mahadevan V, Dargaei Z, Ivakine EA, Hartmann AM, Ng D, Chevrier J, Ormond J, Nothwang HG, McInnes RR, Woodin MA - Front Cell Neurosci (2015)

Bottom Line: Using gramicidin perforated patch clamp recordings we found that the reversal potential for GABA (EGABA) was significantly depolarized.We also observed that surface levels of KCC2 and phosphorylation of KCC2 serine 940 (Ser940) were reduced in Neto2(-/-) neurons compared to wild-type controls.To examine GABAergic inhibition we recorded spontaneous inhibitory postsynaptic currents (sIPSCs) and found that Neto2(-/-) neurons had significant reductions in both their amplitude and frequency.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Systems Biology, University of Toronto Toronto, ON, Canada.

ABSTRACT
Neto2 is a transmembrane protein that interacts with the neuron-specific K(+)-Cl(-) cotransporter (KCC2) in the central nervous system (CNS). Efficient KCC2 transport is essential for setting the neuronal Cl(-) gradient, which is required for fast GABAergic inhibition. Neto2 is required to maintain the normal abundance of KCC2 in neurons, and increases KCC2 function by binding to the active oligomeric form of this cotransporter. In the present study, we characterized GABAergic inhibition and KCC2-mediated neuronal chloride homeostasis in pyramidal neurons from adult hippocampal slices. Using gramicidin perforated patch clamp recordings we found that the reversal potential for GABA (EGABA) was significantly depolarized. We also observed that surface levels of KCC2 and phosphorylation of KCC2 serine 940 (Ser940) were reduced in Neto2(-/-) neurons compared to wild-type controls. To examine GABAergic inhibition we recorded spontaneous inhibitory postsynaptic currents (sIPSCs) and found that Neto2(-/-) neurons had significant reductions in both their amplitude and frequency. Based on the critical role of Neto2 in regulating GABAergic inhibition we rationalized that Neto2- mice would be prone to seizure activity. We found that Neto2- mice demonstrated a decrease in the latency to pentylenetetrazole (PTZ)-induced seizures and an increase in seizure severity.

No MeSH data available.


Related in: MedlinePlus

Neto2- CA1 neurons have reduced spontaneous IPSCs. (A) Example of sIPSC traces obtained from CA1 pyramidal neurons in acute hippocampal slices prepared from wild-type mice (Ai), or Neto2- mice (Aii); WT: n = 7 neurons; Neto2−/−: n = 8 neurons; across three animals. (Bi) Cumulative probability plot indicating the distribution of sIPSC peak amplitude; Inset: Histograms (mean ± S.E.) showing the sIPSC peak amplitude from the two genotypes. ***p < 0.001. (Bii) Average sIPSC peak amplitude plotted against time course analyzed. (C) Histograms (mean ± S.E.) showing the sIPSC frequency of the events of different amplitudes.
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Figure 5: Neto2- CA1 neurons have reduced spontaneous IPSCs. (A) Example of sIPSC traces obtained from CA1 pyramidal neurons in acute hippocampal slices prepared from wild-type mice (Ai), or Neto2- mice (Aii); WT: n = 7 neurons; Neto2−/−: n = 8 neurons; across three animals. (Bi) Cumulative probability plot indicating the distribution of sIPSC peak amplitude; Inset: Histograms (mean ± S.E.) showing the sIPSC peak amplitude from the two genotypes. ***p < 0.001. (Bii) Average sIPSC peak amplitude plotted against time course analyzed. (C) Histograms (mean ± S.E.) showing the sIPSC frequency of the events of different amplitudes.

Mentions: Results are given as mean ± SEM. Statistical significance for Figures 1Aii,B, 2B, 5B were tested using the Student’s t test; statistical significance was determined as follows: *p < 0.05, **p < 0.01, ***p < 0.001. For Figure 4, differences between strains was analyzed using the Cochran-Armitage Trend Test (**z < 0.01).


Neto2- mice have impaired GABAergic inhibition and are susceptible to seizures.

Mahadevan V, Dargaei Z, Ivakine EA, Hartmann AM, Ng D, Chevrier J, Ormond J, Nothwang HG, McInnes RR, Woodin MA - Front Cell Neurosci (2015)

Neto2- CA1 neurons have reduced spontaneous IPSCs. (A) Example of sIPSC traces obtained from CA1 pyramidal neurons in acute hippocampal slices prepared from wild-type mice (Ai), or Neto2- mice (Aii); WT: n = 7 neurons; Neto2−/−: n = 8 neurons; across three animals. (Bi) Cumulative probability plot indicating the distribution of sIPSC peak amplitude; Inset: Histograms (mean ± S.E.) showing the sIPSC peak amplitude from the two genotypes. ***p < 0.001. (Bii) Average sIPSC peak amplitude plotted against time course analyzed. (C) Histograms (mean ± S.E.) showing the sIPSC frequency of the events of different amplitudes.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4585209&req=5

Figure 5: Neto2- CA1 neurons have reduced spontaneous IPSCs. (A) Example of sIPSC traces obtained from CA1 pyramidal neurons in acute hippocampal slices prepared from wild-type mice (Ai), or Neto2- mice (Aii); WT: n = 7 neurons; Neto2−/−: n = 8 neurons; across three animals. (Bi) Cumulative probability plot indicating the distribution of sIPSC peak amplitude; Inset: Histograms (mean ± S.E.) showing the sIPSC peak amplitude from the two genotypes. ***p < 0.001. (Bii) Average sIPSC peak amplitude plotted against time course analyzed. (C) Histograms (mean ± S.E.) showing the sIPSC frequency of the events of different amplitudes.
Mentions: Results are given as mean ± SEM. Statistical significance for Figures 1Aii,B, 2B, 5B were tested using the Student’s t test; statistical significance was determined as follows: *p < 0.05, **p < 0.01, ***p < 0.001. For Figure 4, differences between strains was analyzed using the Cochran-Armitage Trend Test (**z < 0.01).

Bottom Line: Using gramicidin perforated patch clamp recordings we found that the reversal potential for GABA (EGABA) was significantly depolarized.We also observed that surface levels of KCC2 and phosphorylation of KCC2 serine 940 (Ser940) were reduced in Neto2(-/-) neurons compared to wild-type controls.To examine GABAergic inhibition we recorded spontaneous inhibitory postsynaptic currents (sIPSCs) and found that Neto2(-/-) neurons had significant reductions in both their amplitude and frequency.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Systems Biology, University of Toronto Toronto, ON, Canada.

ABSTRACT
Neto2 is a transmembrane protein that interacts with the neuron-specific K(+)-Cl(-) cotransporter (KCC2) in the central nervous system (CNS). Efficient KCC2 transport is essential for setting the neuronal Cl(-) gradient, which is required for fast GABAergic inhibition. Neto2 is required to maintain the normal abundance of KCC2 in neurons, and increases KCC2 function by binding to the active oligomeric form of this cotransporter. In the present study, we characterized GABAergic inhibition and KCC2-mediated neuronal chloride homeostasis in pyramidal neurons from adult hippocampal slices. Using gramicidin perforated patch clamp recordings we found that the reversal potential for GABA (EGABA) was significantly depolarized. We also observed that surface levels of KCC2 and phosphorylation of KCC2 serine 940 (Ser940) were reduced in Neto2(-/-) neurons compared to wild-type controls. To examine GABAergic inhibition we recorded spontaneous inhibitory postsynaptic currents (sIPSCs) and found that Neto2(-/-) neurons had significant reductions in both their amplitude and frequency. Based on the critical role of Neto2 in regulating GABAergic inhibition we rationalized that Neto2- mice would be prone to seizure activity. We found that Neto2- mice demonstrated a decrease in the latency to pentylenetetrazole (PTZ)-induced seizures and an increase in seizure severity.

No MeSH data available.


Related in: MedlinePlus