Limits...
Dentate gyrus network dysfunctions precede the symptomatic phase in a genetic mouse model of seizures.

Toader O, Forte N, Orlando M, Ferrea E, Raimondi A, Baldelli P, Benfenati F, Medrihan L - Front Cell Neurosci (2013)

Bottom Line: Neuronal circuit disturbances that lead to hyperexcitability in the cortico-hippocampal network are one of the landmarks of temporal lobe epilepsy.We made use of a high-resolution microelectrode array (4096 electrodes) and patch-clamp recordings, and found that in acute hippocampal slices of young pre-symptomatic (3-6 week-old) Syn II(-/-) mice excitatory synaptic output of the mossy fibers is reduced.Moreover, we showed that the main excitatory neurons present in the polymorphic layer of the DG, hilar mossy cells, display a reduced excitability.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience and Brain Technologies, Fondazione Istituto Italiano di Tecnologia Genoa, Italy ; International Max-Planck Research School for Neurosciences Göttingen, Germany.

ABSTRACT
Neuronal circuit disturbances that lead to hyperexcitability in the cortico-hippocampal network are one of the landmarks of temporal lobe epilepsy. The dentate gyrus (DG) network plays an important role in regulating the excitability of the entire hippocampus by filtering and integrating information received via the perforant path. Here, we investigated possible epileptogenic abnormalities in the function of the DG neuronal network in the Synapsin II (Syn II) knockout mouse (Syn II(-/-)), a genetic mouse model of epilepsy. Syn II is a presynaptic protein whose deletion in mice reproducibly leads to generalized seizures starting at the age of 2 months. We made use of a high-resolution microelectrode array (4096 electrodes) and patch-clamp recordings, and found that in acute hippocampal slices of young pre-symptomatic (3-6 week-old) Syn II(-/-) mice excitatory synaptic output of the mossy fibers is reduced. Moreover, we showed that the main excitatory neurons present in the polymorphic layer of the DG, hilar mossy cells, display a reduced excitability. We also provide evidence of a predominantly inhibitory regulatory output from mossy cells to granule cells, through feed-forward inhibition, and show that the excitatory-inhibitory ratio is increased in both pre-symptomatic and symptomatic Syn II(-/-) mice. These results support the key role of the hilar mossy neurons in maintaining the normal excitability of the hippocampal network and show that the late epileptic phenotype of the Syn II(-/-) mice is preceded by neuronal circuitry dysfunctions. Our data provide new insights into the mechanisms of epileptogenesis in the Syn II(-/-) mice and open the possibility for early diagnosis and therapeutic interventions.

No MeSH data available.


Related in: MedlinePlus

A decreased number of mossy fiber synaptic vesicles is associated with reduced mEPSC frequency and amplitude in mossy cells from pre-symptomatic Syn II−/− mice. (A) Transmission electron microscopy images of mossy fiber terminals in the DG hilus of brain slices from WT (black bars) and pre-symptomatic Syn II−/− (red bars) mice (scale bar 200 nm). (B) Mean (±s.e.m.) density of total SVs and number of docked SVs in presynaptic terminals of WT and Syn II−/− neurons; ***p < 0.001, two-tailed unpaired Student's t-test. (C) Representative Syn II−/− hilar mossy cell patch-clamped in an acute brain slice and filled with AlexaFluor568. Yellow arrows point toward thorny excrescences that are visible in the focal plane. (D) Representative mEPSC traces recorded in mossy cells from WT (black lines) and pre-symptomatic Syn II−/− (red lines) mice in the presence of GABA receptor and Na+ channel blockers. (E) Cumulative distributions of the amplitudes and frequencies of mEPSCs in WT and Syn II−/− neurons; ***p < 0.001, Kolmogorov–Smirnov test. (F) Mean (±s.e.m.) rise-time (10–90%) and mono-exponential τ of decay of mEPSCs from WT (black bars) and Syn II−/− (red bars) neurons.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3757301&req=5

Figure 4: A decreased number of mossy fiber synaptic vesicles is associated with reduced mEPSC frequency and amplitude in mossy cells from pre-symptomatic Syn II−/− mice. (A) Transmission electron microscopy images of mossy fiber terminals in the DG hilus of brain slices from WT (black bars) and pre-symptomatic Syn II−/− (red bars) mice (scale bar 200 nm). (B) Mean (±s.e.m.) density of total SVs and number of docked SVs in presynaptic terminals of WT and Syn II−/− neurons; ***p < 0.001, two-tailed unpaired Student's t-test. (C) Representative Syn II−/− hilar mossy cell patch-clamped in an acute brain slice and filled with AlexaFluor568. Yellow arrows point toward thorny excrescences that are visible in the focal plane. (D) Representative mEPSC traces recorded in mossy cells from WT (black lines) and pre-symptomatic Syn II−/− (red lines) mice in the presence of GABA receptor and Na+ channel blockers. (E) Cumulative distributions of the amplitudes and frequencies of mEPSCs in WT and Syn II−/− neurons; ***p < 0.001, Kolmogorov–Smirnov test. (F) Mean (±s.e.m.) rise-time (10–90%) and mono-exponential τ of decay of mEPSCs from WT (black bars) and Syn II−/− (red bars) neurons.

Mentions: Mossy cells were identified by their shape, which has a triangular appearance in an infrared differential interference contrast image, size (mossy cells are clearly larger that surrounding interneurons) and location (deep hilus). Mossy cells were also filled with AlexaFluor 568 (40 μM in the pipette solution) to make the specific complex spines (named thorny excrescences) on the proximal dendrites visible in epifluorescence imaging (see Figure 4C). For cells that could not be clearly visualized, electrophysiological features, such as firing frequency adaptation during positive current injection, reduced afterhyperpolarization, broad action potentials, and high frequency/large amplitude spontaneous excitatory postsynaptic currents (sEPSPs) in the presence of bicuculline/CGP 55845, were used to identify them. Intrinsic cell properties were calculated from recordings performed in the presence of 50 μM D-APV, 10 μM CNQX, 5 μM CGP 55845, and 30 μM bicuculline. For the recording of miniature excitatory postsynaptic currents (mEPSCs), aCSF cointaining 5 μM CGP 55845, 30 μM bicuculline, and 0.3 μM TTX was used. All experiments were performed at a holding potential (Vh) of -70 mV in the presence of 30 μM bicuculine and 5 μM CGP 55845 (all from Tocris Bioscience, Ellisville, MO).


Dentate gyrus network dysfunctions precede the symptomatic phase in a genetic mouse model of seizures.

Toader O, Forte N, Orlando M, Ferrea E, Raimondi A, Baldelli P, Benfenati F, Medrihan L - Front Cell Neurosci (2013)

A decreased number of mossy fiber synaptic vesicles is associated with reduced mEPSC frequency and amplitude in mossy cells from pre-symptomatic Syn II−/− mice. (A) Transmission electron microscopy images of mossy fiber terminals in the DG hilus of brain slices from WT (black bars) and pre-symptomatic Syn II−/− (red bars) mice (scale bar 200 nm). (B) Mean (±s.e.m.) density of total SVs and number of docked SVs in presynaptic terminals of WT and Syn II−/− neurons; ***p < 0.001, two-tailed unpaired Student's t-test. (C) Representative Syn II−/− hilar mossy cell patch-clamped in an acute brain slice and filled with AlexaFluor568. Yellow arrows point toward thorny excrescences that are visible in the focal plane. (D) Representative mEPSC traces recorded in mossy cells from WT (black lines) and pre-symptomatic Syn II−/− (red lines) mice in the presence of GABA receptor and Na+ channel blockers. (E) Cumulative distributions of the amplitudes and frequencies of mEPSCs in WT and Syn II−/− neurons; ***p < 0.001, Kolmogorov–Smirnov test. (F) Mean (±s.e.m.) rise-time (10–90%) and mono-exponential τ of decay of mEPSCs from WT (black bars) and Syn II−/− (red bars) neurons.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3757301&req=5

Figure 4: A decreased number of mossy fiber synaptic vesicles is associated with reduced mEPSC frequency and amplitude in mossy cells from pre-symptomatic Syn II−/− mice. (A) Transmission electron microscopy images of mossy fiber terminals in the DG hilus of brain slices from WT (black bars) and pre-symptomatic Syn II−/− (red bars) mice (scale bar 200 nm). (B) Mean (±s.e.m.) density of total SVs and number of docked SVs in presynaptic terminals of WT and Syn II−/− neurons; ***p < 0.001, two-tailed unpaired Student's t-test. (C) Representative Syn II−/− hilar mossy cell patch-clamped in an acute brain slice and filled with AlexaFluor568. Yellow arrows point toward thorny excrescences that are visible in the focal plane. (D) Representative mEPSC traces recorded in mossy cells from WT (black lines) and pre-symptomatic Syn II−/− (red lines) mice in the presence of GABA receptor and Na+ channel blockers. (E) Cumulative distributions of the amplitudes and frequencies of mEPSCs in WT and Syn II−/− neurons; ***p < 0.001, Kolmogorov–Smirnov test. (F) Mean (±s.e.m.) rise-time (10–90%) and mono-exponential τ of decay of mEPSCs from WT (black bars) and Syn II−/− (red bars) neurons.
Mentions: Mossy cells were identified by their shape, which has a triangular appearance in an infrared differential interference contrast image, size (mossy cells are clearly larger that surrounding interneurons) and location (deep hilus). Mossy cells were also filled with AlexaFluor 568 (40 μM in the pipette solution) to make the specific complex spines (named thorny excrescences) on the proximal dendrites visible in epifluorescence imaging (see Figure 4C). For cells that could not be clearly visualized, electrophysiological features, such as firing frequency adaptation during positive current injection, reduced afterhyperpolarization, broad action potentials, and high frequency/large amplitude spontaneous excitatory postsynaptic currents (sEPSPs) in the presence of bicuculline/CGP 55845, were used to identify them. Intrinsic cell properties were calculated from recordings performed in the presence of 50 μM D-APV, 10 μM CNQX, 5 μM CGP 55845, and 30 μM bicuculline. For the recording of miniature excitatory postsynaptic currents (mEPSCs), aCSF cointaining 5 μM CGP 55845, 30 μM bicuculline, and 0.3 μM TTX was used. All experiments were performed at a holding potential (Vh) of -70 mV in the presence of 30 μM bicuculine and 5 μM CGP 55845 (all from Tocris Bioscience, Ellisville, MO).

Bottom Line: Neuronal circuit disturbances that lead to hyperexcitability in the cortico-hippocampal network are one of the landmarks of temporal lobe epilepsy.We made use of a high-resolution microelectrode array (4096 electrodes) and patch-clamp recordings, and found that in acute hippocampal slices of young pre-symptomatic (3-6 week-old) Syn II(-/-) mice excitatory synaptic output of the mossy fibers is reduced.Moreover, we showed that the main excitatory neurons present in the polymorphic layer of the DG, hilar mossy cells, display a reduced excitability.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience and Brain Technologies, Fondazione Istituto Italiano di Tecnologia Genoa, Italy ; International Max-Planck Research School for Neurosciences Göttingen, Germany.

ABSTRACT
Neuronal circuit disturbances that lead to hyperexcitability in the cortico-hippocampal network are one of the landmarks of temporal lobe epilepsy. The dentate gyrus (DG) network plays an important role in regulating the excitability of the entire hippocampus by filtering and integrating information received via the perforant path. Here, we investigated possible epileptogenic abnormalities in the function of the DG neuronal network in the Synapsin II (Syn II) knockout mouse (Syn II(-/-)), a genetic mouse model of epilepsy. Syn II is a presynaptic protein whose deletion in mice reproducibly leads to generalized seizures starting at the age of 2 months. We made use of a high-resolution microelectrode array (4096 electrodes) and patch-clamp recordings, and found that in acute hippocampal slices of young pre-symptomatic (3-6 week-old) Syn II(-/-) mice excitatory synaptic output of the mossy fibers is reduced. Moreover, we showed that the main excitatory neurons present in the polymorphic layer of the DG, hilar mossy cells, display a reduced excitability. We also provide evidence of a predominantly inhibitory regulatory output from mossy cells to granule cells, through feed-forward inhibition, and show that the excitatory-inhibitory ratio is increased in both pre-symptomatic and symptomatic Syn II(-/-) mice. These results support the key role of the hilar mossy neurons in maintaining the normal excitability of the hippocampal network and show that the late epileptic phenotype of the Syn II(-/-) mice is preceded by neuronal circuitry dysfunctions. Our data provide new insights into the mechanisms of epileptogenesis in the Syn II(-/-) mice and open the possibility for early diagnosis and therapeutic interventions.

No MeSH data available.


Related in: MedlinePlus