Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2014 Jul 29;111(30):E3139-48.
doi: 10.1073/pnas.1411131111. Epub 2014 Jul 14.

Impaired excitability of somatostatin- and parvalbumin-expressing cortical interneurons in a mouse model of Dravet syndrome

Chao Tai  1 , Yasuyuki Abe  2 , Ruth E Westenbroek  1 , Todd Scheuer  1 , William A Catterall  3
Affiliations

Impaired excitability of somatostatin- and parvalbumin-expressing cortical interneurons in a mouse model of Dravet syndrome

Chao Tai et al. Proc Natl Acad Sci U S A. .

Abstract

Haploinsufficiency of the voltage-gated sodium channel NaV1.1 causes Dravet syndrome, an intractable developmental epilepsy syndrome with seizure onset in the first year of life. Specific heterozygous deletion of NaV1.1 in forebrain GABAergic-inhibitory neurons is sufficient to cause all the manifestations of Dravet syndrome in mice, but the physiological roles of specific subtypes of GABAergic interneurons in the cerebral cortex in this disease are unknown. Voltage-clamp studies of dissociated interneurons from cerebral cortex did not detect a significant effect of the Dravet syndrome mutation on sodium currents in cell bodies. However, current-clamp recordings of intact interneurons in layer V of neocortical slices from mice with haploinsufficiency in the gene encoding the NaV1.1 sodium channel, Scn1a, revealed substantial reduction of excitability in fast-spiking, parvalbumin-expressing interneurons and somatostatin-expressing interneurons. The threshold and rheobase for action potential generation were increased, the frequency of action potentials within trains was decreased, and action-potential firing within trains failed more frequently. Furthermore, the deficit in excitability of somatostatin-expressing interneurons caused significant reduction in frequency-dependent disynaptic inhibition between neighboring layer V pyramidal neurons mediated by somatostatin-expressing Martinotti cells, which would lead to substantial disinhibition of the output of cortical circuits. In contrast to these deficits in interneurons, pyramidal cells showed no differences in excitability. These results reveal that the two major subtypes of interneurons in layer V of the neocortex, parvalbumin-expressing and somatostatin-expressing, both have impaired excitability, resulting in disinhibition of the cortical network. These major functional deficits are likely to contribute synergistically to the pathophysiology of Dravet syndrome.

PubMed Disclaimer

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Excitability of cortical layer V PV interneurons. (A) Double labeling of cortical layer V PV interneurons from WT animals labeled with anti-NaV1.1 antibody (green) and anti-PV antibody (red). The merged image shows double-labeled PV interneurons in yellow to illustrate the presence of NaV1.1 channels in PV interneurons in cortical layer V. (Scale bar: 100 µm.) (B) Sample whole-cell current-clamp recordings in response to incremental steps of current (3 s, ranging from 140–380 pA), in a WT and a HET PV interneuron. (Calibration: 1 s, 40 mV.) (C and D) The mean number (C) and frequency (D) of APs in response to each step for WT (n = 19) and HET (n = 18). (E–I) Properties of individual WT and HET APs. (E) Expanded and superimposed individual APs from the recordings of WT and HET PV interneurons in B. (Calibration: 2 ms, 15 mV.) (F) Mean AP threshold. (G) Mean AP amplitude. (H) Mean rheobase. (I) Mean AP half width. Significant differences between WT and HET are expressed as *P < 0.05; **P < 0.01.
Fig. 2.
Fig. 2.
Ability of cortical layer V PV interneurons to sustain repetitive firing. Whole-cell current-clamp recordings in response to trains of 100 10-ms pulses. Pulse amplitude was the minimum current required to trigger an AP. Frequency was 1, 2, 5, 10, 20, or 50 Hz. (A) Sample APs of the 1-Hz train for WT and HET PV interneurons. (Calibration: 5 ms, 20 mV.) (B) Sample traces at 5 (calibration: 2 s, 25 mV), 10 (calibration: 1 s, 25 mV), and 20 (calibration: 0.5 s, 25 mV) Hz. (C) Percent of stimuli that failed to produce an AP as a function of frequency. *P < 0.05; **P < 0.01.
Fig. 3.
Fig. 3.
Monosynaptic connection between layer V pyramidal neurons and fast-spiking PV interneurons. (A) Cartoon showing the dual patch clamp of a layer V pyramidal neuron (PY) and a neighboring layer V PV interneuron. (B and C) Excitatory monosynaptic connection from layer V pyramidal neurons to PV interneurons. (B) Sample traces (black for WT, gray for HET) of evoked EPSPs in postsynaptic PV interneurons (Post PV) in response to a single AP in a presynaptic pyramidal neuron (Pre PY). (C) The amplitudes of postsynaptic EPSPs in PV interneurons were reduced significantly in HET (n = 8) compared with WT (n = 10) (4.29 ± 0.52 mV in WT vs. 2.26 ± 0.57 mV in HET; **P = 0.009) in response to APs in presynaptic pyramidal neurons. (D and E) Inhibitory monosynaptic connection from layer V PV interneurons to pyramidal neurons. (D) The amplitudes of inhibitory postsynaptic potentials (IPSPs) in layer V pyramidal neurons as a function of number of presynaptic APs in PV interneurons (WT: n = 7; HET: n = 7). (E) Sample traces (black for WT, gray for HET) of postsynaptic IPSP in layer V pyramidal neurons in response to different numbers of APs in presynaptic PV interneurons.
Fig. 4.
Fig. 4.
Excitability of cortical layer V SST interneurons. (A) Cortical layer V SST interneurons were double labeled with anti-NaV1.1 (green) and anti-SST antibody (red) to show the presence of anti-NaV1.1 channels in SST interneurons. The merged image shows double-labeled SST interneurons in yellow. (Scale bar: 100 µm.) (B) Sample traces of whole-cell current-clamp recordings in response to incremental steps of current (1 s, ranging from 40–200 pA) in a WT and a HET SST interneuron. (Calibration: 0.6 s, 60 mV.) (C and D) The mean number (C) and frequency (D) of APs in response to each step for WT (n = 21) and HET (n = 23). Note that HET SST interneurons require larger current injections to trigger spikes. (E–I) Properties of individual WT and HET APs. (E) Expanded and superimposed individual APs from recordings of the WT and HET SST interneurons in B. (Calibration: 5 ms, 20 mV.) (F) Mean AP threshold. (G) Mean AP rheobase. (H) Mean AP amplitude. (I) Mean AP half width. *P < 0.05.
Fig. 5.
Fig. 5.
Ability of cortical layer V SST interneurons to sustain repetitive firing. Whole-cell current-clamp recordings in response to trains of 100 10-ms pulses. Pulse amplitude was the minimum current to trigger an AP. Frequency was 1, 2, 5, 10, or 20 Hz. (A) Sample APs of the 1-Hz train for WT and HET SST interneurons. (Calibration: 4 ms, 25 mV.) (B) Sample traces at 5 (calibration: 2 s, 40 mV), 10 (calibration: 1 s, 40 mV), and 20 (calibration: 0.5 s, 40 mV) Hz. (C) Percent of stimuli that failed to produce an AP as a function of frequency. *P < 0.05; **P < 0.01; ***P < 0.001.
Fig. 6.
Fig. 6.
Martinotti cell-mediated FDDI. (A) Cartoon showing the dual patch clamp of a pair of neighboring layer V pyramidal neurons with an intermediate layer V Martinotti cell. (B) A train of 15 APs at 70 Hz in one pyramidal neuron (Upper) triggered a delayed hyperpolarization (Lower) in the second pyramidal neuron. Ten representative traces (gray) and their average (black) are superimposed. (Calibration: 0.2 s, 0.5 mV.) (C) Fraction of dual patch-clamp recordings producing measurable disynaptic inhibition using this protocol. WT: 48%, 32/67; HET: 26%, 19/73; Fisher’s exact test, *P = 0.009. (D) Amplitude of disynaptic inhibition in successful pairs. WT: 1.63 ± 0.22 mV (n = 35), HET: 0.89 ± 0.15 mV (n = 19); **P = 0.0035.
Fig. 7.
Fig. 7.
Frequency dependence of FDDI. (A) Examples of traces of FDDI in response to 15 presynaptic APs at the indicated frequencies. (Calibration: 0.2 s, 1 mV.) (B) Fraction of recordings giving measurable FDDI at lower frequency for pairs with measurable Martinotti cell-mediated FDDI in response to 15 presynaptic APs at 70 Hz and 100 Hz. (C and D) Peak FDDI amplitude (C) and area (D) as a function of frequency. Fifteen presynaptic APs were generated at the indicated frequencies (WT, n = 16; HET, n = 12) *P < 0.05; **P < 0.01. (E) The peak latency of the Martinotti cell-mediated FDDI decreases as a function of the frequency of the presynaptic AP train in both WT and HET animals at 20 Hz (0.68 ± 0.03 ms in WT vs. 0.77 ± 0.05 ms in HET; P = 0.17), 30 Hz (0.51 ± 0.03 ms in WT vs. 0.57 ± 0.05 ms in HET; P = 0.17), 50 Hz (0.39 ± 0.03 ms in WT vs. 0.41 ± 0.03 ms in HET; P = 0.31), 70 Hz (0.30 ± 0.01 ms in WT vs. 0.34 ± 0.02 ms in HET; P = 0.09), and 100 Hz (0.29 ± 0.01 ms in WT vs. 0.29 ± 0.01 ms in HET; P = 0.48).
Fig. 8.
Fig. 8.
Dependence of FDDI on the number of presynaptic APs. (A) Examples of FDDI in response to 5, 10, 15, 20, and 30 presynaptic APs at 70 Hz. (Calibration: 0.2 s, 1 mV.) (B) Fraction of recordings giving measurable FDDI in response to fewer stimuli for pairs with measurable Martinotti cell-mediated FDDI in response to 15 presynaptic APs at 70 Hz. (C and D) Peak FDDI amplitude (C) and area (D) as a function of number of presynaptic APs (WT, n = 16; HET, n = 12). (E) The peak latency of the Martinotti cell-mediated FDDI did not change in response to presynaptic stimulation with five APs (0.31 ± 0.01 ms in WT vs. 0.34 ± 0.03 ms in HET; P = 0.38), eight APs (0.31 ± 0.01 ms in WT vs. 0.35 ± 0.02 ms in HET; P = 0.16), 10 APs (0.32 ± 0.01 ms in WT vs. 0.34 ± 0.02 ms in HET; P = 0.25), 12 APs (0.31 ± 0.01 ms in WT vs. 0.34 ± 0.02 ms in HET; P = 0.13), 15 APs (0.31 ± 0.01 ms in WT vs. 0.34 ± 0.03 ms in HET; P = 0.38), 20 APs (0.31 ± 0.01 ms in WT vs. 0.34 ± 0.02 ms in HET; P = 0.14), or 30 APs (0.31 ± 0.01 ms in WT vs. 0.33 ± 0.03 ms in HET; P = 0.29).
Fig. 9.
Fig. 9.
Monosynaptic connection between layer V pyramidal neurons and Martinotti cells. (A) Cartoon showing the dual patch clamp of a layer V pyramidal neuron (PY) and a neighboring layer V Martinotti cell (SST). (B–D) Excitatory monosynaptic connection from layer V pyramidal neurons to Martinotti cells. (B) The amplitudes of postsynaptic EPSPs in Martinotti cells were reduced in HET compared with WT cells when evoked by one (1.21 ± 0.09 mV in WT vs. 0.95 ± 0.14 mV in HET cells; P = 0.06), three (2.38 ± 0.26 mV in WT vs. 1.57 ± 0.18 mV in HET cells; *P = 0.01), and five (3.60 ± 0.50 mV in WT vs. 2.13 ± 0.29 mV in HET cells; *P = 0.01) APs in presynaptic pyramidal neurons. (C) The number of postsynaptic APs in Martinotti cells evoked by presynaptic stimulations was impaired dramatically in HET as compared with WT cells when evoked by 10 (0.31 ± 0.13 mV in WT vs. 0 ± 0 mV in HET cells; *P = 0.02), 15 (0.69 ± 0.23 mV in WT vs. 0.13 ± 0.09 mV in HET cells; *P = 0.02), 20 (1.00 ± 0.31 mV in WT vs. 0.21 ± 0.10 mV in HET cells; *P = 0.02), and 30 (1.46 ± 0.36 mV in WT vs. 0.36 ± 0.15 mV in HET; *P = 0.01) APs in presynaptic pyramidal neurons. (D) Sample traces (black for WT, gray for HET cells) of EPSPs and evoked APs in Martinotti cells in response to different numbers of APs in presynaptic pyramidal neurons. (E and F) Inhibitory monosynaptic connection from layer V Martinotti cells to pyramidal neurons. (E) The amplitudes of postsynaptic IPSPs in layer V pyramidal neurons as a function of number of presynaptic APs in Martinotti cells. (F) Sample traces (black for WT, gray for HET cells) of Martinotti cell-induced postsynaptic IPSPs in layer V pyramidal neurons in response to different numbers of APs in presynaptic pyramidal neurons.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Wolff M, Cassé-Perrot C, Dravet C. Severe myoclonic epilepsy of infants (Dravet syndrome): Natural history and neuropsychological findings. Epilepsia. 2006;47(Suppl 2):45–48. - PubMed
    1. Genton P, Velizarova R, Dravet C. Dravet syndrome: The long-term outcome. Epilepsia. 2011;52(Suppl 2):44–49. - PubMed
    1. Brunklaus A, Dorris L, Zuberi SM. Comorbidities and predictors of health-related quality of life in Dravet syndrome. Epilepsia. 2011;52(8):1476–1482. - PubMed
    1. Claes L, et al. De novo mutations in the sodium-channel gene SCN1A cause severe myoclonic epilepsy of infancy. Am J Hum Genet. 2001;68(6):1327–1332. - PMC - PubMed
    1. Claes LR, et al. The SCN1A variant database: A novel research and diagnostic tool. Hum Mutat. 2009;30(10):E904–E920. - PubMed

Publication types

MeSH terms