p21-activated kinase 1 restricts tonic endocannabinoid signaling in the hippocampus
- Southeast University, China
- The Hospital for Sick Children, Canada
- University of Toronto, Canada
- Cumming School of Medicine, Canada
- University of Calgary, Canada
Figures
Figure 1
Genetic ablation of PAK1 enhances E/I ratio by selectively suppressing inhibitory synaptic responses.
(a) Diagram of a hippocampal slice showing the placement of stimulating and recoding electrodes. (b) A representative whole-cell recording experiment and samples traces at indicated time points …
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Figure 1—source data 1
Statistical data summary for Figure 1d: input/output curves of eIPSC using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.004
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Figure 1—source data 2
Statistical data summary for Figure 1e: IPA3 effect on eIPSC using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.005
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Figure 1—source data 3
Statistical data summary for Figure 1f: PAK1 inhibitory peptide effect on eIPSC using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.006
Figure 2 with 2 supplements
PAK1 deletion specifically reduces the frequency, but not the amplitude of inhibitory synaptic responses.
(a) Sample traces of sIPSC recordings. (b, c) Summary graphs of (a) showing normal distribution and mean value of the amplitude (b: WT = 19.07 ± 2.44 pA, n = 13 (5); KO = 16.44 ± 1.62 pA, n = 20 (6);…
Figure 2—figure supplement 1
Normal inhibitory transmission in ROCK2 KO mice.
https://doi.org/10.7554/eLife.14653.008-
Figure 2—figure supplement 1—source data 1
Statistical data summary for Figure 2—figure supplement 1: Normal inhibitory transmission in ROCK2 KO mice using one-way ANOVA.
- https://doi.org/10.7554/eLife.14653.009
Figure 2—figure supplement 2
Impaired transmitter depletion in response to sustained synaptic activation.
https://doi.org/10.7554/eLife.14653.010
Figure 3
Acute disruption of postsynaptic PAK1 also selectively impairs the frequency, but not the amplitude of inhibitory synaptic responses.
(a) Sample traces of sIPSC recordings. Scale bar: 60 pA/1 s. (b, c) Summary graphs of (a) showing normal amplitude (b: WT+DMSO = 16.14 ± 0.65 pA, n = 13 (4); WT+IPA3 = 16.83 ± 0.95 pA, n = 15 (4); …
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Figure 3—source data 1
Statistical data summary for Figure 3b,c: Effect of IPA3 on frequency and amplitude of sIPSC of WT and PAK1 KO neurons using two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.012
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Figure 3—source data 2
Statistical data summary for Figure 3e,f: Effect of IPA3 on frequency and amplitude of mIPSC of WT and PAK1 KO neurons using two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.013
Figure 4 with 1 supplement
Normal GABAergic neurons, synapses, GABA receptor function and postsynaptic actin network in PAK1 KO mice.
(a) Confocal images of hippocampal sections stained with the nucleus marker DAPI and GABA and summary graph (b) showing similar number of GABAergic neurons in PAK1 KO and WT control mice (WT = 30 ± …
Figure 4—figure supplement 1
Normal GABAergic neurons and synapses in the cortex.
https://doi.org/10.7554/eLife.14653.015
Figure 5
GABAergic transmission is independent of postsynaptic actin cytoskeleton.
(a–f) Sample traces of sIPSCs and summary graphs showing neither the actin polymerization inhibitor cytochalasin D (a–c) nor Rac1 inhibitor NSC23766 (d–f) had an effect on the amplitude (b: DMSO = …
Figure 6 with 1 supplement
PAK1 disruption enhances endocannabinoid signaling.
(a) Sample traces of eIPSC and averaged data showing that bath application of the CB1 receptor antagonist AM251 potentiated the amplitude of eIPSC significantly more in PAK1 KO compared to WT …
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Figure 6—source data 1
Statistical data summary for Figure 6a: Effect of AM251 on eIPSC in WT and PAK1 KO using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.018
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Figure 6—source data 2
Statistical data summary for Figure 6b: Effect of AM251 on eIPSC in the presence or absence of IPA3 using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.019
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Figure 6—source data 3
Statistical data summary for Figure 6c: Effect of AM251 on eIPSC in the presence or absence of PAK1 inhibitory peptide using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.020
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Figure 6—source data 4
Statistical data summary for Figure 6f: Effect of WIN on eIPSC in WT and PAK1 KO using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.021
Figure 6—figure supplement 1
The lack of effect of DMSO on eIPSCs.
https://doi.org/10.7554/eLife.14653.022-
Figure 6—figure supplement 1—source data 1
Statistical data summary for Figure 6—figure supplement 1: Lack of effect of DMSO on eIPSC using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.023
Figure 7 with 1 supplement
Elevated AEA and reduced COX-2 in PAK1 KO mice.
(a) Summary graph showing a significant increase in hippocampal tissue AEA in PAK1 KO compared to WT control (WT = 5.82 ± 0.44 pmol/g, n = 11 (11); KO = 7.64 ± 0.70 pmol/g, n = 13 (13), *p=0.046; …
Figure 7—figure supplement 1
Reduced synaptosomal COX-2 in PAK1 KO hippocampus.
https://doi.org/10.7554/eLife.14653.025
Figure 8
COX-2 inhibition recapitulates the effect of PAK1 disruption.
(a) Sample traces and averaged data of eIPSCs showing that bath application of the COX-2 inhibitor Nim depressed eIPSCs in WT, but this depression was significantly reduced in PAK1 KO neurons …
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Figure 8—source data 1
Statistical data summary for Figure 8a: Effect of Nimesulide on eIPSC in WT and PAK1 KO using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.027
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Figure 8—source data 2
Statistical data summary for Figure 8b: Effect of AM251 on eIPSC after Nimesulide treatment in WT and PAK1 KO using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.028
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Figure 8—source data 3
Statistical data summary for Figure 8c: Effect of URB597 on eIPSC in WT and PAK1 KO using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.029
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Figure 8—source data 4
Statistical data summary for Figure 8d: Effect of AM251 on eIPSC after URB597 treatment in WT and PAK1 KO using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.030
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Figure 8—source data 5
Statistical data summary for Figure 8e: Effect of JZL184 on eIPSC in WT and PAK1 KO using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.031
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Figure 8—source data 6
Statistical data summary for Figure 8f: Effect of AM251 on eIPSC after JZL treatment in WT and PAK1 KO using repeated measures two-way ANOVA.
- https://doi.org/10.7554/eLife.14653.032
Figure 9
Reduced COX-2 localization at GABAergic synapses in PAK1 KO neurons.
(a–c) Cultured hippocampal neurons costained for PAK1 and the excitatory marker PSD-95 (a), the GABAergic marker gephyrin (b) or COX-2 (c) showing PAK1 colocalization with PSD-95, gephyrin and …
Figure 10
A hypothetical model.
In wild type neurons, constitutively active PAK1 is required for maintaining a sufficient level of synaptic COX-2 to keep AEA low, thus less suppression of GABA release and normal inhibitory …
