Article Figures & Data
- Figure 1.
Ablation of Kcnq2 from neonatal excitatory neurons leads to increased excitability in the hippocampus and posterior cortex. All recordings were in the presence of 2.5 mm Ko. A, top panels, Examples of acute slices from control and Pyr:Kcnq2 mice with one hemisphere segmented into ROIs. Bottom panels, Representative 2D plots showing calcium activity (ΔF/F) as a function of time for the different ROIs. The numbering corresponds to the segmented area shown on the top panels with lower values toward the anterior cortex (AC) and higher values toward the CA3 area of the hippocampus and posterior cortex (PC). B, Summary graphs show the effect of Kcnq2 ablation on the calcium event duration and frequency for the CA3 area of the hippocampus and the posterior cortex. Data in the box plots show the median and the interquartile range. (*p < 0.05, **p < 0.01, ****p < 0.0001). C, Whole-cell recordings from CA3 pyramidal neurons show that ablation of Kcnq2 increases neuronal excitability. Left panels, Family of depolarizing 1-s current steps in either control or Pyr:Kcnq2 slices. Right, Summary graph shows the number of recorded action potentials in control and Pyr:Kcnq2 CA3 pyramidal neurons. *p < 0.05 compared with control group. For C, data are represented as mean ± SEM. Additional details on the statistical analysis for this figure are found in Table 1 under the Figure 1 section.
- Figure 2.
Deletion of Kcnq2 from excitatory neurons leads to elevated calcium activity across the forebrain in 8 mm Ko. A, top panels, Examples of acute slices from control and Pyr:Kcnq2 mice with one hemisphere segmented into ROIs. Bottom panels, 2D plots show the calcium activity across the different ROIs. The numbering corresponds to the segmented area shown on the top panels with lower values toward the AC and higher values toward the PC. Note that in the absence of Kcnq2, substantial calcium activity is measured across all regions of the forebrain. B, top two panels, Temporal evolution of the LFPs and ΔF/F recorded in parallel in the CA3 region of the hippocampus. Note that in contrast to the LFPs in slices from Pyr:Kcnq2 mice, the calcium responses are large and long lasting. Middle and bottom panels, Temporal evolution of the ΔF/F across multiple ROIs. C, Violin plots show the effect of Kcnq2 deletion on the amplitude, duration, and frequency of the calcium events for different anatomic regions. MC refers to the medial cortex. Note that ablation of Kcnq2 led to a large and uniform increase of the calcium response amplitude and duration. ****p < 0.0001. Additional details on the statistical analysis and number of replicates for this figure are found in Table 1 under the Figure 2 section.
- Figure 3.
Deletion of Kcnq2 leads to LF calcium oscillations. Deletion of Kcnq2 increased the likelihood of LF oscillations across a large number of ROIs; oscillations were sustained throughout the duration of the recording period. All recordings were in the presence of 8 mm Ko. A, Wavelets from control and Pyr:Kcnq2 hemispheres. The wavelets were generated from the examples shown in Figure 2A. Note the large increase in the power for the oscillations occurring at the LF range in the hemisphere from Pyr:Kcnq2 animals. Although in this example some increases in the power were also observed at the higher-frequency values, this was not seen across all slices, as shown in panel C, right panels. B, Comparison of the probability density of the frequency f of transient oscillations (left panels) and the number of ROIs undergoing sustained oscillations (right panels) for the examples depicted in panel A. C, left, Comparison of the probability density of the transient oscillations for control (n = 43) and Pyr:Kcnq2 hemispheres (n = 47) across multiple slices. Data are represented as mean ± SEM. Middle and right panels, Summary graphs quantifying the power measured for the two frequency domains, 0.02–0.2 and 0.2–2 Hz. Note that loss of Kcnq2 increased the likelihood of observing LF calcium oscillations ranging primarily from 0.03 to 0.05 Hz. Data are represented as mean ± SEM (****p < 0.0001). D, left, Comparison of the ROIs undergoing sustained oscillations for control (n = 43) and Pyr:Kcnq2 (n = 47) hemispheres. Results are reported for sustained oscillations whose frequency
- Figure 4.
GABAA receptors limit activity in neonatal control and Pyr:Kcnq2 brain slices. All recordings were in the presence of 8 mm Ko. A, Panels show examples of acute slices from control and Pyr:Kcnq2 mice before and after application of 50 μm PTX to block GABAA receptors. Top panels, Hemispheres segmented into ROIs. Bottom panels, 2D plots representing the changes in the calcium activity (ΔF/F) across the different ROIs on application of PTX. The numbering corresponds to the segmented area shown on top with lower values toward the AC and higher values toward the PC. Note that inhibiting GABAA receptors led to widespread calcium activity in the Pyr:Kcnq2 hemisphere. In contrast, the activity in the control hemisphere was primarily confined to the posterior cortex and the hippocampal formation following application of PTX. B, Calcium responses from the different anatomic regions before and after application of PTX. Note that in Pyr:Kcnq2 slices application of PTX led to the appearance of a barrage of activity across all regions. C, Scatter plots show the effect of PTX on the ΔF/F amplitude and duration. Note that application of PTX increased the number of larger and faster calcium signals. Additional details on the statistical analysis and number of replicates for this figure are found Table 1 under the Figure 4 section.
- Figure 5.
GABAA receptor activity limits the power but not the occurrence of LF calcium oscillations in Pyr:Kcnq2 slices. All recordings were in the presence of 8 mm Ko. A, Representative examples show calcium activity (ΔF/F) over time and the corresponding wavelets in the presence and absence of 50 μm PTX for control and Pyr:Kcnq2 slices. Note the large increase in power at the LF range in Pyr:Kcnq2 slices, further quantified across multiple slices in panel C. B, Comparison of the probability density of the frequency f for transient oscillations before and after PTX in control (n = 20) and Pyr:Kcnq2 (n = 14) hemispheres. Data are represented as mean ± SEM. The absence of change in the probability density on blocking GABAA receptor activity suggests that GABAA receptors alter the peak oscillation frequency at the different ROIs. C, Summary graphs show that application of PTX increased the power for the LF and HF domains in control and Pyr:Kcnq2 slices. D, Summary graphs show that on application of PTX the number of ROIs undergoing sustained oscillations in control and Pyr:Kcnq2 slices did not change. Together, C, D suggest that blocking GABAA receptors primarily increases the activity within each ROI. Data are presented as mean ± SEM (***p < 0.001, ****p < 0.0001). Additional details on the statistical analysis for panels C, D are found in Table 1 under the Figure 5 section. nd, not determined.
- Figure 6.
Kcnq2 deficient slices acquired NMDA receptor independent calcium activities. All recordings were in the presence of 8 mm Ko. A, Panels show examples of acute slices from control and Pyr:Kcnq2 mice before and after application of 25 μm D-APV (APV). Top panels, Hemispheres segmented into ROIs. Bottom panels, 2D plots representing the ΔF/F as a function of time for the different ROIs. The numbering corresponds to the segmented area shown on top with lower values toward the anterior cortex (AC) and higher values toward the posterior cortex (PC). B, Temporal evolution of calcium activity (ΔF/F) for the different anatomic regions before and after application of APV. Note that application of APV did not prevent the occurrence of large slow calcium events. C, Scatter plots show the effect of APV on the amplitude and duration of the calcium events across the different anatomic regions. Note that blocking NMDA receptors primarily targeted calcium events with faster durations (i.e., <10 s). In the medial cortex (MC) from control slices, application of APV eliminated all activity. The ΔF/F amplitude at the PC in the presence of APV went below the cutoff threshold of 0.01 ΔF/F; thus, these data points are not shown. Additional details on the statistical analysis are found in Table 1 under the Figure 6 section.
- Figure 7.
Blocking NMDA receptors does not prevent the emergence of slow oscillatory calcium activity in Pyr:Kcnq2 slices. All recordings were in the presence of 8 mm Ko. A, Representative examples of control and Pyr:Kcnq2 2D plots show calcium activity across ROIs and the corresponding wavelets. 2D plots and wavelets before and after application of 25 μm D-APV (APV) are shown. Note that APV reduced the power of the slow oscillatory activity in Pyr:Kcnq2 slices. B, Comparison of the probability density for the transient oscillation frequency ( f) in the presence and absence of APV in control (n = 20) and Pyr:Kcnq2 (n = 14) hemispheres. Note that APV did not change the likelihood of the emergence of a slow oscillatory activity at the 0.03- to 0.05-Hz frequency range. C, Summary graphs show the effect of APV in control and Pyr:Kcnq2 hemispheres on the power at the LF (0.02–0.2 Hz) and HF (0.2–2 Hz) domains. Note that APV reduced the power across all oscillatory frequencies, suggesting that NMDA receptors promote the calcium activity. D, Summary graphs show the effect of APV on the number of ROIs in control and Pyr:Kcnq2 slices, demonstrating sustained oscillations at frequency f*. Note that blocking NMDA receptors led to a decrease in the number of ROIs undergoing sustained oscillations in Pyr:Kcnq2 hemispheres, consistent with our observed reduction in power shown in C. Data are represented as mean ± SEM (**p < 0.01, ***p < 0.001, ****p < 0.0001). Additional details on the statistical analysis for panel C are found in Table 1 under the Figure 7 section. nd, not determined.
- Figure 8.
Calcium activity is maintained in the presence of fast excitatory transmission blockers, but eliminated by TTX in slices from Pyr:Kcnq2 mice. All recordings were in the presence of 8 mm Ko. A, top panels, Representative examples of control and Pyr:Kcnq2 slices demonstrating calcium activity across all ROIs in the presence and absence of 50 μm PTX followed by 25 μm D-APV and 25 μm NBQX. The numbering corresponds to the segmented area shown on the left with lower values toward the anterior cortex (AC) and higher values toward the posterior cortex (PC). B, Calcium activity (ΔF/F) across multiple ROIs representing different forebrain anatomic areas in the presence and absence of the GABAA receptor blocker PTX (middle panels) and glutamatergic transmission blockers APV and NBQX (right panels). Note that APV/NBQX primarily inhibited the smaller and faster calcium events that had emerged in the presence of PTX in Pyr:Kcnq2 slices. Summary information regarding the amplitude, duration, and event frequency for multiple slices is found in Table 1 under the Figure 8 section. C, 1 μm TTX (n = 6 slices) abolishes the calcium responses in slices preincubated with PTX/APV/NBQX.
- Figure 9.
Differential effects of fast synaptic receptor blockers on the oscillation frequencies in control and Pyr:Kcnq2 slices. All recordings were in the presence of 8 mm Ko. A, Representative examples of control and Pyr:Kcnq2 2D plots and their corresponding wavelets in the presence and absence of 50 μm PTX, 25 μm D-APV, and 25 μm NBQX (PTX/APV/NBQX). Note that application of all synaptic blockers reduced the power at low frequencies, but did not prevent the emergence of slow oscillatory activity in the Pyr:Kcnq2 hemisphere. B, Comparison of the probability density of the frequency f of transient oscillations before and after application of PTX/APV/NBQX in control (n = 18) and Pyr:Kcnq2 (n = 14) hemispheres. C, Summary graphs show the effect of PTX/APV/NBQX on the power in control and Pyr:Kcnq2 hemispheres for the LF (0.02–0.2 Hz) and HF (0.2–2 Hz) domains. D, Summary graphs show the effect of PTX/APV/NBQX on the number of ROIs undergoing sustained oscillations in control and Pyr:Kcnq2 hemispheres. Data are presented as mean ± SEM (**p < 0.01, ***p < 0.001, ****p < 0.0001). Additional details on the statistical analysis for panels C, D are found in Table 1 under the Figure 9 section.
- Figure 10.
Ablation of Kcnq3 leads to hyperexcitability across the forebrain in the presence of 8 mm Ko. A, top panels, Two examples of acute slices from Kcnq3+/+ and Kcnq3−/− mice with one hemisphere segmented into ROIs. Below, 2D plots show the changes in the ΔF/F as a function of time for the different ROIs. Note that calcium activity is recorded across all regions in the Kcnq3-null slice. B, Raw traces show calcium activity (ΔF/F) across the different anatomic areas. Top two panels, Temporal evolution of LFPs and ΔF/F recorded in parallel from the CA3 region of the hippocampus. C, Violin plots show the effect of Kcnq3 deletion on the ΔF/F amplitude, duration, and event frequency (events/s) for the different anatomic regions. The median and the interquartile ranges are also shown (*p < 0.05, **p < 0.01, ****p < 0.0001). Additional details on the statistical analysis for panel C are found in Table 1 under the Figure 10 section.
- Figure 11.
Loss of Kcnq3 leads to LF calcium oscillations in multiple ROIs. All recordings were in the presence of 8 mm Ko. A, Representative examples of Kcnq3+/+ and Kcnq3−/− ΔF/F 2D plots and wavelets. Top panels, ROIs used to generate the ΔF/F 2D plots and the wavelets. B, Comparison of the probability density of the frequency f of transient oscillations and number of ROIs undergoing sustained oscillations for the examples shown in panel A. C, left, Comparison of the probability density of the frequency f of transient oscillations for Kcnq3+/+ (n = 24) and Kcnq3−/− (n = 24) across multiple hemispheres. Right, Comparison of ROIs undergoing sustained oscillations for Kcnq3+/+ (n = 24) and Kcnq3−/− (n = 24) hemispheres at different frequencies. D, left panels, Summary graphs of the power at two frequency domains, 0.02–0.2 and 0.2–2 Hz. Right panels, Summary graphs of the ROIs exhibiting sustained oscillations at frequency
- Figure 12.
Synaptic blockers inhibit calcium activity in a region-specific manner in Kcnq3−/− slices. All recordings were in the presence of 8 mm Ko. A, top two panels, Recorded LFP and ΔF/F activity from the CA3 region of the hippocampus. Middle and bottom panels, Temporal evolution of the ΔF/F across different anatomic areas. Note that synaptic blockers (PTX/APV/NBQX) abolish the calcium activity only in the medial cortex (MC) and anterior cortex (AC). B, Box plots show the effect of Kcnq3−/− ablation on the ΔF/F amplitude, and event frequency for the different anatomic regions (*p < 0.05, ****p < 0.0001). Note that although application of synaptic blockers did not inhibit the amplitude of calcium events equally across the different regions, it did lead to a decrease to the number of calcium events across all regions. Additional details on the statistical analysis and number of replicates for panel B are found in Table 1 under the Figure 12 section.
- Figure 13.
In Kcnq3−/− slices synaptic blockers substantially reduce slow calcium oscillations in the neocortex. All recordings were in the presence of 8 mm Ko. A, Representative examples of control and Kcnq3−/− 2D plots along with their corresponding wavelets in the presence and absence of 50 μm PTX, 25 μm D-APV, and 25 μm NBQX (PTX/APV/NBQX). B, Comparison of the probability density of the frequency f of transient oscillations before and after addition of PTX/APV/NBQX in Kcnq3+/+ (n = 18) and Kcnq3−/− (n = 18) hemispheres. Note that application of the synaptic blockers did not prevent the occurrence of the slow calcium oscillations in Kcnq3-null slices. C, Summary graphs show the effect of PTX/APV/NBQX on the power for the LF (0.02–0.2 Hz) and HF (0.2–2 Hz) domains in Kcnq3+/+ and Kcnq3−/− hemispheres. Note that synaptic blockers reduce the power across the board, consistent with the large reduction of the activity in the neocortex. D, Summary graphs show the effect of PTX/APV/NBQX on the number of ROIs in Kcnq3+/+ and Kcnq3−/− hemispheres that showed sustained oscillations at different frequencies. Data are presented as mean ± SEM (***p < 0.01, ****p < 0.001). Additional details on the statistical analysis for panels C, D are found in Table 1 under the Figure 13 section.
Figures Values Statistical tests Figure 1 Control Pyr:Kcnq2 1B CA3 Duration 1.6 ± 0.46 s 2.2 ± 0.35 s Mann–Whitney U test p = 0.26 Control: n = 10; Pyr:Kcnq2: n = 18 hemispheres Events/s 0.0054 ± 0.0015 0.013 ± 0.0025 Mann–Whitney U test p = 0.0143 Control: n = 22; Pyr:Kcnq2: n = 21 hemispheres Posterior cortex Duration 1.15 ± 0.24 s 5.3 ± 0.61 s Mann–Whitney U test p < 0.0001 Control: n = 15; Pyr:Kcnq2: n = 20 hemispheres Events/s 0.008 ± 0.0018 Hz 0.015 ± 0.0018 Hz Mann–Whitney U test p = 0.0039 Control: n = 22; Pyr:Kcnq2: n = 20 hemispheres
Animals: Control: n = 6; Pyr:Kcnq2: n = 6.1C CA3 APs vs current injections Two-way ANOVA (p = 0.0025;
F(9,33) = 3.745)Control: n = 5; pPyr:Kcnq2: n = 6 cells Figure 2 Control Pyr:Kcnq2 Statistical tests and values 2C CA3 Amplitude 0.064 ± 0.007 △F/F 0.14 ± 0.014 △F/F Mann–Whitney U test p < 0.0001 Control: n = 46; Pyr:Kcnq2: n = 56 hemispheres Duration 1.4 ± 0.18 s 8.5 ± 0.68 s Mann–Whitney U test p < 0.0001 Events/s 0.26 ± 0.023 Hz 0.043 ± 0.0047 Hz Mann–Whitney U test p < 0.0001 Posterior cortex (PC) Amplitude 0.08 ± 0.006 △F/F 0.29 ± 0.023 △F/F Mann–Whitney U test p < 0.0001 Control: n = 44; Pyr:Kcnq2: n = 56 hemispheres Duration 2.2 ± 0.19 s 6.5 ± 0.37 s Mann–Whitney U test p < 0.0001 Events/s 0.14 ± 0.0072 Hz 0.048 ± 0.0032 Hz Mann–Whitney U test p < 0.0001 Medial cortex (MC) Amplitude 0.021 ± 0.01 △F/F 0.33 ± 0.029 △F/F Mann–Whitney U test p < 0.0001 Control, n = 43; Pyr:Kcnq2: n = 55 hemispheres Duration 1.4 ± 0.58 s 9.5 ± 0.56 s Mann–Whitney U test p < 0.0001 Events/s 0.0065 ± 0.0024 Hz 0.018 ± 0.0023 Hz Mann–Whitney U test Anterior cortex (AC) Amplitude 0.02 ± 0.003 △F/F 0.36 ± 0.03 △F/F Mann–Whitney U test p < 0.0001 Control: n = 44; Pyr:Kcnq2: n = 54 hemispheres Duration 1.5 ± 0.3 s 11.2 ± 0.8 s Mann–Whitney U test p < 0.0001 Events/s 0.012 ± 0.0021 Hz 0.021 ± 0.0025 Hz Mann–Whitney U test p = 0.0005 Data were obtained from 8 control and 9 Pyr:Kcnq2 mice. Figure 3 Control Pyr:Kcnq2 Statistical tests and values 3C Power 0.02–0.2 Hz 0.022 ± 0.0015 A.U. 0.11 ± 0.0046 A.U. Mann–Whitney U test p < 0.0001 Control, n = 43; Pyr:Kcnq2:, n = 47 hemispheres Power 0.2–2 Hz 0.026 ± 0.011 A.U. 0.38 ± 0.012A.U. Mann–Whitney U test p < 0.0001 3D ROI 0.02–0.2 Hz 10.1 ± 0.7 ROIs 19.9 ± 0.7 ROIs Mann–Whitney U test p < 0.0001 Control, n = 43; Pyr:Kcnq2:, n = 47 hemispheres ROI 0.2–2 Hz 6.6 ± 0.5 ROIs 0.5 ± 0.2 ROIs Mann–Whitney U test p < 0.0001 Data were obtained from 8 control and 9 Pyr:Kcnq2 mice. Figure 4 Control Pyr:Kcnq2 Statistical tests and values 4C CA3 Amplitude –PTX 0.057 △F/F ± 0.055 SD –PTX 0.08 △F/F ± 0.09 SD Mann–Whitney U test Control
p < 0.0001Control: –PTX: n = 1547; +PTX: n = 1322 events
Pyr:Kcnq2: –PTX: n = 224 events ; +PTX: n = 523 events+PTX 0.27 △F/F ± 0.12 SD +PTX 0.19 △F/F ± 0.09 SD Mann–Whitney U test Pyr:Kcnq2
p < 0.0001Duration –PTX 0.8 s ± 1.5 SD –PTX 4.3 s ± 5.4 SD Mann–Whitney U test Control
p < 0.0001+PTX 0.7 ± 1.7 SD +PTX 1.8 ± 3.8 SD Mann–Whitney U test Pyr:Kcnq2
p < 0.0001Posterior cortex Amplitude –PTX 0.08 △F/F ± 0.058 SD –PTX 0.21 △F/F ± 0.26 SD Mann–Whitney U test Control
p < 0.0001Control: –PTX: n = 931; +PTX: n = 777 events
Pyr:Kcnq2: –PTX: n = 223; +PTX: n = 444 events+PTX 0.43 △F/F ± 0.20 SD +PTX 0.38 △F/F ± 0.24 SD Mann–Whitney U test Pyr:Kcnq2
p < 0.0001Duration –PTX 1.9 s ± 2.4 SD –PTX 5.1 s ± 4.5 SD Mann–Whitney U test Control
p < 0.0001+PTX 0.58 s ± 1.0 SD +PTX 1.5 ± 1.9 SD Mann–Whitney U test Pyr:Kcnq2
p < 0.0001Medial cortex Amplitude –PTX 0.053 △F/F ± 0.066 SD –PTX 0.21 △F/F ± 0.25 SD Mann–Whitney U test Control
p < 0.0001Control: –PTX: n = 94; +PTX: n = 270 events
Pyr:Kcnq2: –PTX: n = 62; +PTX: n = 267 events+PTX 0.13 △F/F ± 0.22 SD +PTX 0.27 △F/F ± 0.24 SD Mann–Whitney U test Pyr:Kcnq2
p = 0.0003Duration –PTX 0.9 s ± 0.74 SD –PTX 7.8 s ± 6.7 SD Mann–Whitney U test Control
p < 0.0034+PTX 1.3 s ± 2.3 SD +PTX 2.1 ± 3.3 SD Mann–Whitney U test Pyr:Kcnq2
p < 0.0001Anterior cortex Amplitude –PTX 0.048 △F/F ± 0.038 SD –PTX 0.16 △F/F ± 0.26 SD Mann–Whitney U test Control: p < 0.0001 Control: –PTX: n = 125; +PTX: n = 310 events
Pyr:Kcnq2: –PTX: n = 150; +PTX: n = 66 events+PTX 0.14 △F/F ± 0.1 SD +PTX 0.37 △F/F ± 0.31 SD Mann–Whitney U test Pyr:Kcnq2
p < 0.0001Duration –PTX 1.0 s ± 1.3 SD –PTX 4.8 s ± 6.1 SD Mann–Whitney U test Control
p = 0.11+PTX 0.8 s ± 0.9 SD +PTX 5.3 ± 7.1 SD Mann–Whitney U test Pyr:Kcnq2
p = 0.87Data were obtained from 3 control and 3 Pyr:Kcnq2 mice. Figure 5 Control Pyr:Kcnq2 Statistical tests and values 5C Power 0.02–0.2 Hz –PTX 0.02 ± 0.003 A.U. –PTX 0.11 ± 0.01 A.U. Control
Wilcoxon paired testControl
p < 0.0001Control: n = 20; Pyr:Kcnq2: n = 14 hemispheres +PTX 0.06 ± 0.007 A.U. +PTX 0.18 ± 0.01 A.U. Pyr:Kcnq2
paired Student’s t testp < 0.0001
t = 6.233, df = 13Power 0.2–2 Hz –PTX 0.28 ± 0.02 A.U. –PTX 0.37 ± 0.02 A.U. Control
Wilcoxon pairedControl
p < 0.0001Control: n = 20;
Pyr:Kcnq2: n = 14 hemispheres+PTX 0.58 ± 0.03 A.U. +PTX 0.73 ± 0.0 A.U.6 Pyr:Kcnq2
Wilcoxon paired testPyr:Kcnq2
p < 0.00015D ROI 0.02–0.2 Hz –PTX 8.8 ± 0.7 ROIs –PTX 23.0 ± 1.4 ROIs Control
paired Student’s t testControl
p = 0.84 t = 0.2042,
df = 19Control: n = 20; Pyr:Kcnq2: n = 14 hemispheres +PTX 9.1 ± 1.1 ROIs +PTX 25.2 ± 1.4 ROIs Pyr:Kcnq2
paired Student’s t testPyr:Kcnq2
p = 0.19 t = 1.377,
df = 13ROI 0.2–2 Hz –PTX 7.6 ± 0.6 ROIs –PTX 0 ROI Control
Wilcoxon paired testControl
p = 0.72Control: n = 20; Pyr:Kcnq2: n = 14 hemispheres +PTX 8.7 ± 1.6 ROIs +PTX 0 ROI Pyr:Kcnq2
N/AN/A Data were obtained from 4 control and 3 Pyr:Kcnq2 mice. Figure 6 Control Pyr:Kcnq2 Statistical tests and values 6C CA3 Amplitude –D-APV 0.070 △F/F ± 0.069 SD –D-APV 0.13 △F/F ± 0.14 SD Mann–Whitney U test Control: p < 0.0001 Control: –D-APV n = 1265; +D-APV: n = 571 events
Pyr:Kcnq2: –D-APV n = 193; +D-APV: n = 156 events+D-APV 0.036 △F/F ± 0.014 SD +D-APV 0.09 △F/F ± 0.1 SD Mann–Whitney U test Pyr:Kcnq2: p < 0.0001 Duration –D-APV 0.88 s ± 2.0 SD –D-APV 5.4 s ± 5.8 SD Mann–Whitney U test Control: p < 0.0001 +D-APV 0.78 ± 1.2 SD +D-APV 5.4 ± 7.1 SD Mann–Whitney U test Pyr:Kcnq2: p = 0.0055 Posterior cortex Amplitude –D-APV 0.07 △F/F ± 0.049 SD –D-APV 0.26 △F/F ± 0.36 SD Mann–Whitney U test Control: p < 0.0001 Control: –D-APV n = 543; +D-APV n = 201 events
Pyr:Kcnq2: –D-APV n = 243; +D-APV n = 170 events+D-APV 0.02 △F/F ± 0.01 SD +D-APV 0.13 △F/F ± 0.14 SD Mann–Whitney U test Pyr:Kcnq2: p < 0.0001 Duration –D-APV 1.99 s ± 2.1 SD –D-APV 5.1 s ± 4.2 SD Mann–Whitney U test Control: p < 0.0001 +D-APV 2.5s ± 2.7 SD +D-APV 10.8 ± 6.6 SD Mann–Whitney U test Pyr:Kcnq2: p < 0.0001 Medial cortex Amplitude –D-APV 0.11 △F/F ± 0.16 SD –D-APV 0.18 △F/F ± 0.2 SD Control: N/A Control: N/A Control –D-APV n = 10; +D-APV n = 0 events
Pyr:Kcnq2 –D-APV n = 158; +D-APV, n = 50events;+D-APV N/A +D-APV 0.14 △F/F ± 0.16 SD Pyr:Kcnq2: Mann–Whitney U test Pyr:Kcnq2: p = 0.22 Duration –D-APV 7.8s ± 8.4 SD –D-APV 6.3 ± 6.2 SD Control: N/A Control: N/A +D-APV N/A +D-APV 12.9 ± 8.4 SD Pyr:Kcnq2: Mann–Whitney U test Pyr:Kcnq2: p < 0.0001 Anterior cortex Amplitude –D-APV 0.017 △F/F ± 0.006 SD –D-APV 0.31 △F/F ± 0.4 SD Control: N/A Control: N/A Control –D-APV n = 54; +D-APV n = 0 events
Pyr:Kcnq2: –D-APV n = 71; +D-APV n = 29 events+D-APV N/A +D-APV 0.15 △F/F ± 0.19 SD Pyr:Kcnq2: Mann–Whitney U test Pyr:Kcnq2: p = 0.0055 Duration –D-APV 2.5 s ± 3.2 SD –D-APV 8.9 s ± 7.1 SD Control: N/A Control: N/A +D-APV N/A +D-APV 13.3 ± 11 SD Pyr:Kcnq2: Mann–Whitney U test Pyr:Kcnq2: p = 0.04 Data were obtained from 3 control and 3 Pyr:Kcnq2 mice. Figure 7 Control Pyr:Kcnq2 Statistical tests and values 7C Power 0.02–0.2 Hz –D-APV 0.019 ± 0.0015 A.U. –D-APV 0.11 ± 0.0085 A.U. Control: Wilcoxon paired Control: p < 0.0001 Control: n = 20; Pyr:Kcnq2: n = 14 hemispheres +D-APV 0.014 ± 0.0015 A.U. +D-APV 0.069 ± 0.0057 A.U. Pyr:Kcnq2: paired Student’s t test Pyr:Kcnq2: p < 0.0001, t = 12.64, df = 13 Power 0.2–2 Hz –D-APV 0.24 ± 0.012 A.U. –D-APV 0.39 ± 0.022 A.U. Control: Wilcoxon paired Control: p = 0.0007 Control: n = 20; Pyr:Kcnq2: n = 14 hemispheres +D-APV 0.18 ± 0.017 A.U. +D-APV 0.26 ± 0.0080 A.U. Pyr:Kcnq2: Wilcoxon paired Pyr:Kcnq2: p = 0.0001 7D ROI 0.02–0.2 Hz –D-APV 8.9 ± 0.65 ROIs –D-APV 25.21 ± 1.3 ROIs Control: paired Student’s t test Control: p = 0.20 t = 1.32, df = 19 Control: n = 20; Pyr:Kcnq2: n = 14 hemispheres +D-APV 7.5 ± 0.92 ROIs +D-APV 20.71 ± 1.0 ROIs Pyr:Kcnq2: paired Student’s t test Pyr:Kcnq2: p = 0.0002 t = 5.22, df = 13 Power 0.2–2 Hz –D-APV 7.05 ± 0.54 ROIs –D-APV 0 ROIs Control: paired Student’s t test Control: p = 0.003 t = 3.472, df = 19 Control: n = 20; Pyr:Kcnq2: n = 14 hemispheres +D-APV 4.5 ± 0.54 ROIs +D-APV 0.43 ± 0.2 ROIs Pyr:Kcnq2: N/A N/A Data were obtained from 3 control and 3 Pyr:Kcnq2 mice. Figure 8 P/N/A refers to Picrotoxin/APV/NBQX Control Pyr:Kcnq2 Statistical tests and values 8B CA3 Amplitude –P/N/A: 0.046 Δ F/F ± 0.14 SD –P/N/A:0.105 ΔF/F ± 0.14 SD Mann–Whitney U test Control: N/A Control –P/N/A: n = 444; +P/N/A: n = 0 events
Pyr:Kcnq2: –P/N/A: n = 246; +P/N/A: n = 38 eventsN/A +P/N/A: ΔF/F 0.14 ± 0.13 SD Pyr:Kcnq2
p = 0.0001Duration –P/N/A: 0.59 s ± 1.3 SD –P/N/A: 4.5 s ± 6.7 SD Mann–Whitney U test Control: N/A N/A +P/N/A: 13.5 s ± 7.5 SD Pyr:Kcnq2: p < 0.0001 Posterior cortex Amplitude 0.059 Δ F/F ± 0.03 SD –P/N/A:0.48 Δ F/F ± 0.63 SD Mann–Whitney U test Control: N/A Control –P/N/A: n = 228; +P/N/A: n = 0 events
Pyr:Kcnq2: –P/N/A: n = 194; +P/N/A: n = 36 eventsN/A +P/N/A: Δ 0.71 F/F ± 0.47 SD Pyr:Kcnq2: p = 0.009 Duration –P/N/A: 1.66 s ± 1.7 SD –P/N/A: 4.6 s ± 3.9 SD Mann–Whitney U test Control: N/A N/A +P/N/A: 10.2 s ± 3.6 SD Pyr:Kcnq2: p < 0.0001 Medial cortex Amplitude 0.028 Δ F/F ± 0.018 SD –P/N/A:0.38 Δ F/F ± 0.31 SD Mann–Whitney U test Control: N/A Control –P/N/A: n = 22; +P/N/A: n = 0 events
Pyr:Kcnq2: –P/N/A: n = 64; +P/N/A: n = 56 eventsN/A +P/N/A: Δ 0.19 F/F ± 0.17 SD Pyr:Kcnq2: p = 0.005 Duration –P/N/A: 0.65 s ± 0.17 SD –P/N/A: 7.6 s ± 4.7 SD Mann–Whitney U test Control: N/A N/A +P/N/A: 8.9 s ± 7.5 SD Pyr:Kcnq2: p = 0.087 Anterior cortex Amplitude 0.029 Δ F/F ± 0.026 SD –P/N/A:0.47 Δ F/F ± 0.48 SD Mann–Whitney U test Control: N/A Control –P/N/A: n = 22; +P/N/A: n = 0 events
Pyr:Kcnq2: –P/N/A: n = 82; +P/N/A: n = 43 eventsN/A +P/N/A: Δ 0.35 F/F ± 0.38 SD Pyr:Kcnq2: p = 0.087 Duration –P/N/A: 0.77 s ± 0.63 SD –P/N/A: 7.7 s ± 5.8 SD Mann–Whitney U test Control: N/A N/A +P/N/A: 14.1 s ± 8.7 SD Pyr:Kcnq2: p = 0.016 Data were obtained from 3 control and 3 Pyr:Kcnq2 mice. Figure 9 P/N/A refers to Picrotoxin/APV/NBQX Control Pyr:Kcnq2 Statistical tests and values 9C,D Power 0.02–0.2 Hz –P/A/N 0.018 ± 0.0007 A.U. –P/A/N 0.13 ± 0.0094 A.U. Control: Wilcoxon paired Control: p < 0.0001 Control: n = 18; Pyr:Kcnq2: n = 14 hemispheres +P/A/N 0.01 ± 0.0006 A.U. +P/A/N 0.068 ± 0.0083 A.U. Pyr:Kcnq2: Wilcoxon paired Pyr:Kcnq2: p = 0.0001 Power 0.2–2 Hz –P/A/N 0.22 ± 0.007 A.U. –P/A/N 0.45 ± 0.03 A.U. Control: paired Student’s t test Control: p < 0.0001 t = 9.106, df = 17 Control: n = 18; Pyr:Kcnq2: n = 14 hemispheres +P/A/N 0.13 ± 0.002 A.U. +P/A/N 0.26 ± 0.01 A.U. Pyr:Kcnq2: paired Student’s t test Pyr:Kcnq2: p < 0.0001 t = 9.055, df = 13 ROI 0.02–0.2 Hz –P/A/N 10.17 ± 0.93 ROIs –P/A/N 19.86 ± 1.2 ROIs Control: Wilcoxon paired Control: p = 0.0001 Control: n = 18; Pyr:Kcnq2: n = 14 hemispheres +P/A/N 3.5 ± 0.85 ROIs +P/A/N 15.14 ± 1.3 ROIs Pyr:Kcnq2: paired Student’s t test Pyr:Kcnq2: p = 0.0092 t = 3.057, df = 13 ROI 0.2–2 Hz –P/A/N 8.7 ± 0.7 ROIs –P/A/N 0 ROI Control: Wilcoxon paired Control: p < 0.0001 Control: n = 18; Pyr:Kcnq2: n = 14 hemispheres +P/A/N 0.72 ± 0.18 ROIs +P/A/N 0 ROI Pyr:Kcnq2: N/A Data were obtained from 3 control and 3 Pyr:Kcnq2 mice. Figure 10 Kcnq3+/+ Kcnq3−/− Statistical tests 10C CA3 Amplitude 0.08 ± 0.005△F/F 0.08 ± 0.007△F/F Welch’s t test p = 0.95, t = 0.07052, df = 42 Kcnq3+/+, n = 23; Kcnq3−/−: n = 21 hemispheres Duration 2.4 ± 0.8 s 1.2 ± 0.2 s Mann–Whitney U test p = 0.93 Events/s 0.22 ± 0.03 Hz 0.19 ± 0.3 Hz Mann–Whitney U test p = 0.05 Kcnq3+/+, n = 24; Kcnq3−/−: n = 24 hemispheres Posterior cortex Amplitude 0.073 ± 0.006△F/F 0.12 ± 0.01△F/F Mann–Whitney U test p < 0.0001 Kcnq3+/+, n = 23; Kcnq3−/−, n = 24 hemispheres Duration 1.4 ± 0.3 s 4.5 ± 0.9 s Mann–Whitney U test p < 0.0001 Events/s 0.21 ± 0.025 Hz 0.12 ± 0.023 Hz Mann–Whitney U test p = 0.013 Kcnq3+/+, n = 24; Kcnq3−/−: n = 24 hemispheres Medial cortex Amplitude 0.028 ± 0.008△F/F 0.09 ± 0.02△F/F Mann–Whitney U test p = 0.08 Kcnq3+/+, n = 13; Kcnq3−/−: n = 21 hemispheres Duration 3.2 ± 1.1 s 4.3 ± 1.2 s Mann–Whitney U test p = 0.004 Events/s 0.02 ± 0.12 Hz 0.017 ± 0.003 Hz Mann–Whitney U test p = 0.009 Kcnq3+/+, n = 24; Kcnq3−/−: n = 24 hemispheres Anterior cortex Amplitude 0.03 ± 0.003△F/F 0.10 ± 0.13△F/F Mann–Whitney U test p < 0.0001 Kcnq3+/+, n = 20; Kcnq3−/−: n = 22 hemispheres Duration 3.1 ± 1.0 s 5.2 ± 1.3 s Mann–Whitney U test p = 0.72 Events/s 0.032 ± 0.007 Hz 0.05 ± 0.007 Hz Mann–Whitney U test p = 0.06 Kcnq3+/+, n = 24; Kcnq3−/−: n = 24 hemispheres Data were obtained from 4 Kcnq3+/+ and 4 Kcnq3−/− mice. Figure 11 Kcnq3+/+ Kcnq3−/− Statistical tests 11D Power 0.02–0.2 Hz 0.035 ± 0.005 A.U. 0.06 ± 0.005 A.U. Mann–Whitney U test p < 0.0001 Kcnq3+/+, n = 24; Kcnq3−/−: n = 24 hemispheres Power 0.2–2 Hz 0.57 ± 0.1 A.U. 0.53 ± 0.08 A.U. Mann–Whitney U test p = 0.67 ROI 0.02–0.2 Hz 12 ± 1 ROIs 25 ± 1 ROIs Mann–Whitney U test p < 0.0001 Kcnq3+/+, n = 24; Kcnq3−/−: n = 24 hemispheres ROI 0.2–2 Hz 6.2 ± 0.8 ROIs 0.6 ± 0.2 ROIs Mann–Whitney U test p < 0.0001 Data were obtained from 4 Kcnq3+/+ and 4 Kcnq3−/− mice. Figure 12 P/N/A refers to Picrotoxin/APV/NBQX Kcnq3−/−
–blockersKcnq3−/−
+blockersStatistical tests 12B CA3 Amplitude 0.09 ± 0.009 △F/F 0.064 ± 0.008 △F/F Student’s t test p = 0.054, t = 2.007, df = 29 Kcnq3−/−: n = 16; +P/A/N: n = 15 hemispheres Duration (not illustrated) 1.1 ± 0.2 s 7.7 ± 1.6 s Wilcoxon paired p = 0.0006 Kcnq3-/: n = 16; P/A/N: n = 15 hemispheres Events/s 0.22 ± 0.03 Hz 0.019 ± 0.3 Hz Wilcoxon paired test p = 0.0002 Kcnq3−/− n = 18; P/A/N: n = 18 hemispheres Posterior cortex Amplitude 0.13 ± 0.01 △F/F 0.17 ± 0.02 △F/F Wilcoxon paired p = 0.15 Kcnq3−/− n = 18; P/A/N: n = 15 hemispheres Duration (not illustrated) 3.1 ± 0.8 s 9.0 ± 0.9 s Wilcoxon paired p = 0.0006 Kcnq3−/− n = 18; P/A/N: n = 15 hemispheres Events/s 0.16 ± 0.03 Hz 0.01 ± 0.003 Hz Wilcoxon paired p < 0.0001 Kcnq3−/− n = 18; P/A/N: n = 18 hemispheres Medial cortex Amplitude 0.12 ± 0.02△F/F ND N/D N/D Kcnq3−/− n = 18; +P/A/N: n = 0 hemispheres Duration (not illustrated) 5.2 ± 1.6 s ND N/D N/D Kcnq3−/− n = 18; P/A/N: n = 0 hemispheres Events/s 0.02 ± 0.004 Hz ND N/D N/D Kcnq3−/− n = 18; P/A/N: n = 18 hemispheres Anterior cortex Amplitude 0.1 ± 0.1△F/F 0.07 ± 0.02△F/F Wilcoxon paired p = 0.88 Kcnq3−/− n = 18; P/A/N: n = 4 hemispheres Duration (not illustrated) 4.0 ± 0.7 s 9.6 ± 3.4 s Wilcoxon paired p = 0.25 Kcnq3−/− n = 18; P/A/N: n = 4 hemispheres Events/s 0.06 ± 0.007 Hz 0.001 ± 0.0006 Hz Wilcoxon paired p < 0.0001 Kcnq3−/− n = 18; P/A/N: n = 18 hemispheres Data were obtained from 3 Kcnq3+/+ and 3 Kcnq3−/− mice. Figure 13 P/N/A refers to Picrotoxin/APV/NBQX Kcnq3+/+ Kcnq3−/− Statistical tests 13C Power 0.02–0.2 Hz –P/A/N 0.025 ± 0.0019 –P/A/N 0.05 ± 0.003 Kcnq3+/+ Wilcoxon paired p < 0.0001 Kcnq3+/+ n = 17; Kcnq3−/− n = 16 hemispheres +P/A/N 0.01 ± 0.0009 +P/A/N 0.02 ± 0.0018 Kcnq3−/− paired Student’s t test p < 0.0001 t = 9.531, df = 15 Power 0.2–2 Hz –P/A/N 0.33 ± 0.012 –P/A/N 0.33 ± 0.019 Kcnq3+/+ paired Student’s t test p < 0.0001 t = 11.60, df = 16 +P/A/N 0.19 ± 0.006 +P/A/N 0.19 ± 0.011 Kcnq3−/− paired Student’s t test p < 0.0001 t = 8.787, df = 15 13D ROI 0.02–0.2 Hz –P/A/N 12.5 ± 0.96 –P/A/N 25 ± 1.3 Kcnq3+/+ Wilcoxon paired p = 0.0004 t = 4.477, df = 16 Kcnq3+/+ n = 17; Kcnq3−/−
n = 16 hemispheres+P/A/N 6.0 ± 1.1 +P/A/N 17.7 ± 1.6 Kcnq3−/− paired Student’s t test p = 0.0006 t = 4.321, df = 15 ROI 0.2–2 Hz –P/A/N 7.1 ± 0.92 –P/A/N 0.75 ± 0.25 Kcnq3+/+ Wilcoxon paired p < 0.0001 +P/A/N 0.76 ± 0.22 +P/A/N 0.81 ± 0.26 Kcnq3−/−Wilcoxon paired p = 0.83 Data were obtained from 3 Kcnq3+/+ and 3 Kcnq3−/− mice
- Movie 1.
Representative imaging showing the forebrain calcium activity in control slices in the presence of 8 mM Ko. Note the activity in the hippocampal formation. Movie played at 5× speed.
- Movie 2.
Representative imaging showing the forebrain calcium activity in Pyr:Kcnq2 slices in the presence of 8 mm Ko. Note the migrating activity across the neocortex. Movie played at 5× speed.
- Movie 3.
Representative imaging showing the forebrain calcium activity in the absence of Kcnq2 in excitatory neurons and in the presence of synaptic blockers PTX, APV, and NBQX. Movie played at 5× speed.
- Movie 4.
Representative imaging showing the forebrain calcium activity in the absence of Kcnq3. Movie played at 5× speed.
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