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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

APS-MEA extracellular field recordings show decreased activity in the hilus upon stimulation of the perforant path. (A) Color-coded fPSP activity in the entire APS-MEA chip (each pixel represents one electrode) in WT and pre-symptomatic Syn II−/− slices with arrows showing the activated areas (GL, granule layer; H, hilus) upon perforant path stimulation. (B,C) Mean (±s.e.m.) amplitude of the entire response area from WT and pre-symptomatic Syn II−/− in the granule layer (B) and hilus (C); *p < 0.05, two-tailed unpaired Student's t-test. (D) Representative traces from one APS-MEA electrode from both genotypes located in the granule layer and hilus, respectively. (E,F) Stimulation-response curves representing the mean (±s.e.m.) peak amplitudes of three randomly selected electrodes from the granule layer (E) or hilus (F) at increasing stimulation intensities; ***p < 0.001, Two-Way ANOVA. (G) Mean (±s.e.m.) stimulus propagation time from GL to H at a stimulation intensity of 500 μA; *p < 0.05, two-tailed unpaired Student's t-test.
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Figure 1: APS-MEA extracellular field recordings show decreased activity in the hilus upon stimulation of the perforant path. (A) Color-coded fPSP activity in the entire APS-MEA chip (each pixel represents one electrode) in WT and pre-symptomatic Syn II−/− slices with arrows showing the activated areas (GL, granule layer; H, hilus) upon perforant path stimulation. (B,C) Mean (±s.e.m.) amplitude of the entire response area from WT and pre-symptomatic Syn II−/− in the granule layer (B) and hilus (C); *p < 0.05, two-tailed unpaired Student's t-test. (D) Representative traces from one APS-MEA electrode from both genotypes located in the granule layer and hilus, respectively. (E,F) Stimulation-response curves representing the mean (±s.e.m.) peak amplitudes of three randomly selected electrodes from the granule layer (E) or hilus (F) at increasing stimulation intensities; ***p < 0.001, Two-Way ANOVA. (G) Mean (±s.e.m.) stimulus propagation time from GL to H at a stimulation intensity of 500 μA; *p < 0.05, two-tailed unpaired Student's t-test.

Mentions: Our first step was to investigate the response of different areas of the DG to the stimulation of the perforant path in horizontal brain slices containing the hippocampus and rhinal cortices from pre-symptomatic (3–6 weeks) Syn II−/− mice. To this aim, we employed a high-resolution Active Pixel Sensor microelectrode array system (APS-MEA, 4096 electrodes: see Materials and Methods) (Ferrea et al., 2012). Field postsynaptic potentials (fPSPs), evoked by the stimulation of the perforant path with an extracellular electrode, were detected in the granule layer of the DG and they further propagated to the hilus (Figure 1A). The mean amplitude of the response, calculated over the entire activated region in the granule cell layer, was similar in WT and Syn II−/− slices (153.7 ± 15.2 μV for WT vs. 135.6 ± 10.6 μV for Syn II−/−, n = 13 slices for each genotype; two-tailed unpaired Student's t-test, p = 0.342) (Figures 1B,D). On the other hand, the mean amplitude of the response in the hilar region was significantly reduced in Syn II−/− slices (143.0 ± 19.4 μV for WT vs. 87.13 ± 10.7 μV for Syn II−/−, n = 12 slices for each genotype; two-tailed unpaired Student's t-test, p = 0.013) (Figures 1C,D). The stimulus amplitude was set to 400–500 μA based on previous input-output curves performed for 3 selected electrodes in each of the two regions. At all stimulation intensities, a reduced amplitude of the responses in the Syn II−/− hilar region with respect to the WT was observed (Figures 1E,F). The recorded signal on several neighboring electrodes on the APS-MEA correlated with the fine anatomy of the DG and its polarity corresponded to current sinks in the dendritic-granule layer (negative) and to current sources in the hilus (positive) (Figure 1D). Moreover, the propagation time of the evoked fPSP, measured between the peaks of the response from one representative electrode in the granule and hilar regions, was significantly longer in Syn II−/− than in WT slices (2.8 ± 0.78 ms for WT vs. 5.6 ± 0.59 ms for Syn II−/−, n = 6/5 slices; two-tailed unpaired Student's t-test, p = 0.032) (Figure 1G), suggesting the presence of functional impairments of the hilar region.


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)

APS-MEA extracellular field recordings show decreased activity in the hilus upon stimulation of the perforant path. (A) Color-coded fPSP activity in the entire APS-MEA chip (each pixel represents one electrode) in WT and pre-symptomatic Syn II−/− slices with arrows showing the activated areas (GL, granule layer; H, hilus) upon perforant path stimulation. (B,C) Mean (±s.e.m.) amplitude of the entire response area from WT and pre-symptomatic Syn II−/− in the granule layer (B) and hilus (C); *p < 0.05, two-tailed unpaired Student's t-test. (D) Representative traces from one APS-MEA electrode from both genotypes located in the granule layer and hilus, respectively. (E,F) Stimulation-response curves representing the mean (±s.e.m.) peak amplitudes of three randomly selected electrodes from the granule layer (E) or hilus (F) at increasing stimulation intensities; ***p < 0.001, Two-Way ANOVA. (G) Mean (±s.e.m.) stimulus propagation time from GL to H at a stimulation intensity of 500 μA; *p < 0.05, two-tailed unpaired Student's t-test.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3757301&req=5

Figure 1: APS-MEA extracellular field recordings show decreased activity in the hilus upon stimulation of the perforant path. (A) Color-coded fPSP activity in the entire APS-MEA chip (each pixel represents one electrode) in WT and pre-symptomatic Syn II−/− slices with arrows showing the activated areas (GL, granule layer; H, hilus) upon perforant path stimulation. (B,C) Mean (±s.e.m.) amplitude of the entire response area from WT and pre-symptomatic Syn II−/− in the granule layer (B) and hilus (C); *p < 0.05, two-tailed unpaired Student's t-test. (D) Representative traces from one APS-MEA electrode from both genotypes located in the granule layer and hilus, respectively. (E,F) Stimulation-response curves representing the mean (±s.e.m.) peak amplitudes of three randomly selected electrodes from the granule layer (E) or hilus (F) at increasing stimulation intensities; ***p < 0.001, Two-Way ANOVA. (G) Mean (±s.e.m.) stimulus propagation time from GL to H at a stimulation intensity of 500 μA; *p < 0.05, two-tailed unpaired Student's t-test.
Mentions: Our first step was to investigate the response of different areas of the DG to the stimulation of the perforant path in horizontal brain slices containing the hippocampus and rhinal cortices from pre-symptomatic (3–6 weeks) Syn II−/− mice. To this aim, we employed a high-resolution Active Pixel Sensor microelectrode array system (APS-MEA, 4096 electrodes: see Materials and Methods) (Ferrea et al., 2012). Field postsynaptic potentials (fPSPs), evoked by the stimulation of the perforant path with an extracellular electrode, were detected in the granule layer of the DG and they further propagated to the hilus (Figure 1A). The mean amplitude of the response, calculated over the entire activated region in the granule cell layer, was similar in WT and Syn II−/− slices (153.7 ± 15.2 μV for WT vs. 135.6 ± 10.6 μV for Syn II−/−, n = 13 slices for each genotype; two-tailed unpaired Student's t-test, p = 0.342) (Figures 1B,D). On the other hand, the mean amplitude of the response in the hilar region was significantly reduced in Syn II−/− slices (143.0 ± 19.4 μV for WT vs. 87.13 ± 10.7 μV for Syn II−/−, n = 12 slices for each genotype; two-tailed unpaired Student's t-test, p = 0.013) (Figures 1C,D). The stimulus amplitude was set to 400–500 μA based on previous input-output curves performed for 3 selected electrodes in each of the two regions. At all stimulation intensities, a reduced amplitude of the responses in the Syn II−/− hilar region with respect to the WT was observed (Figures 1E,F). The recorded signal on several neighboring electrodes on the APS-MEA correlated with the fine anatomy of the DG and its polarity corresponded to current sinks in the dendritic-granule layer (negative) and to current sources in the hilus (positive) (Figure 1D). Moreover, the propagation time of the evoked fPSP, measured between the peaks of the response from one representative electrode in the granule and hilar regions, was significantly longer in Syn II−/− than in WT slices (2.8 ± 0.78 ms for WT vs. 5.6 ± 0.59 ms for Syn II−/−, n = 6/5 slices; two-tailed unpaired Student's t-test, p = 0.032) (Figure 1G), suggesting the presence of functional impairments of the hilar region.

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