IL-17a promotes sociability in mouse models of neurodevelopmental disorders - PubMed (original) (raw)
. 2020 Jan;577(7789):249-253.
doi: 10.1038/s41586-019-1843-6. Epub 2019 Dec 18.
Yeong Shin Yim # 1 2, Ralf D Wimmer 1 2 3, Hyunju Kim 4, Changhyeon Ryu 1 2, Gwyneth Margaret Welch 1 2, Matias Andina 1 2, Hunter Oren King 2, Ari Waisman 5, Michael M Halassa 1 2 3, Jun R Huh 6 7, Gloria B Choi 8 9
Affiliations
- PMID: 31853066
- PMCID: PMC8112727
- DOI: 10.1038/s41586-019-1843-6
IL-17a promotes sociability in mouse models of neurodevelopmental disorders
Michael Douglas Reed et al. Nature. 2020 Jan.
Abstract
A subset of children with autism spectrum disorder appear to show an improvement in their behavioural symptoms during the course of a fever, a sign of systemic inflammation1,2. Here we elucidate the molecular and neural mechanisms that underlie the beneficial effects of inflammation on social behaviour deficits in mice. We compared an environmental model of neurodevelopmental disorders in which mice were exposed to maternal immune activation (MIA) during embryogenesis3,4 with mouse models that are genetically deficient for contactin-associated protein-like 2 (Cntnap2)5, fragile X mental retardation-1 (Fmr1)6 or Sh3 and multiple ankyrin repeat domains 3 (Shank3)7. We establish that the social behaviour deficits in offspring exposed to MIA can be temporarily rescued by the inflammatory response elicited by the administration of lipopolysaccharide (LPS). This behavioural rescue was accompanied by a reduction in neuronal activity in the primary somatosensory cortex dysgranular zone (S1DZ), the hyperactivity of which was previously implicated in the manifestation of behavioural phenotypes associated with offspring exposed to MIA8. By contrast, we did not observe an LPS-induced rescue of social deficits in the monogenic models. We demonstrate that the differences in responsiveness to the LPS treatment between the MIA and the monogenic models emerge from differences in the levels of cytokine production. LPS treatment in monogenic mutant mice did not induce amounts of interleukin-17a (IL-17a) comparable to those induced in MIA offspring; bypassing this difference by directly delivering IL-17a into S1DZ was sufficient to promote sociability in monogenic mutant mice as well as in MIA offspring. Conversely, abrogating the expression of IL-17 receptor subunit a (IL-17Ra) in the neurons of the S1DZ eliminated the ability of LPS to reverse the sociability phenotypes in MIA offspring. Our data support a neuroimmune mechanism that underlies neurodevelopmental disorders in which the production of IL-17a during inflammation can ameliorate the expression of social behaviour deficits by directly affecting neuronal activity in the central nervous system.
Conflict of interest statement
The authors declare no competing financial interests.
Figures
Extended Data Figure 1.. Cntnap2, Fmr1, and Shank3 mutant mice show variable sociability performance.
a, Sociability performance (MIA n=13, WT n=22, Cntnap2 n=71, Fmr1 n=165, Shank3 n=50; from 30 independent experiments). b, Time spent investigating social (S) versus inanimate (I) objects for mice described in (a). c,d, total interaction time (c) and distance traveled (d) during three-chambered sociability experiments described in (a). Statistics calculated by one-way ANOVA with Dunnett’s post-hoc test (a,c,d) and two-way ANOVA with Dunnett’s post-hoc test (b). Graphs indicate mean ± s.e.m.
Extended Data Figure 2.. Further behavioral analyses for sociability performance following LPS treatment in PBS and MIA offspring, and monogenic mutant mice.
a, Time spent investigating social (S) versus inanimate (I) objects (a), total interaction time (b), time spent in social (S), center (C) or inanimate (I) chamber (c), and distance traveled (d) for sociability experiments in Fig 1c (PBS-Veh n=8, PBS-LPS n=9, MIA-Veh n=13, MIA-LPS n=11; from 3 independent experiments). Statistics calculated by two-way ANOVA with Sidak’s (a) Dunnett’s (c) post-hoc test and two-way repeated measures ANOVA with Sidak’s post-hoc test (b,d). Graphs indicate mean ± s.e.m.
Extended Data Figure 3.. LPS-induced rescue of MIA behavioral phenotypes is transient, effective in aged mice, and extends beyond three-chambered sociability.
a-e, Sociability measured 72hrs following Veh or LPS injection in PBS and MIA offspring from Fig. 1c. Data expressed as percent sociability (a), time spent investigating social (S) versus inanimate (I) objects (b), total interaction time (c), time spent in social (S), center (C) or inanimate (I) chamber (d), and distance traveled (e) during three-chambered sociability experiments (PBS-Veh n=7, PBS-LPS n=7, MIA-Veh n=8, MIA-LPS n=6; from 2 independent experiments). f-j, Sociability measured before and 4hr after Veh or LPS injection in aged MIA mice (9–12 months). Data expressed as percent sociability (f), time spent investigating social (S) versus inanimate (I) objects (g), total interaction time (h), time spent in social (S), center (C) or inanimate (I) chamber (i), and distance traveled (j) during three-chambered sociability experiments (MIA-Veh n=6, MIA-LPS n=7: from 2 independent experiments). k, Reciprocal social interactions measured upon Veh or LPS treatment in PBS or MIA offspring (PBS-Veh n=9, PBS-LPS n=9, MIA-Veh n=11, MIA-LPS n=11; from 4 independent experiments). l, Marble burying index (% of buried marbles) measured before and 4hr after Veh or LPS treatment in PBS or MIA offspring (PBS-Veh n=12, PBS-LPS n=12, MIA-Veh n=12, MIA-LPS n=11; from 5 independent experiments). Statistics calculated by two-way ANOVA with Sidak’s (a,b,c,e,g), Dunnett’s (d,i), and Tukey’s (k) post-hoc tests, and two-way repeated measures ANOVA with Sidak’s post-hoc test (f,h,j,l). Graphs indicate mean ± s.e.m.
Extended Data Figure 4.. Acute increase in body temperature is insufficient to promote sociability.
a, Body temperature profile following Veh or LPS injection in MIA offspring (Veh n=10, LPS n=10; from 4 independent experiments). Initial spike in body temperature is due to handling stress. b-e, Data expressed as time spent investigating social (S) versus inanimate (I) objects (b), total interaction time (c), time spent in social (S), center (C) or inanimate (I) chamber (d), and distance traveled (e) during three-chambered sociability experiments described in Fig. 1g (n=9 for all groups; from 2 independent experiments). f-j, Sociability performance in Vgat-Cre PBS and MIA offspring following Veh, CNO, or LPS treatment. Data expressed as percent sociability (f), time spent investigating social (S) versus inanimate (I) objects (g), total interaction time (h), time spent in social (S), center (C) or inanimate (I) chamber (i), and distance traveled (j) during three-chambered sociability experiments (PBS n=11, MIA n=7; from 2 independent experiments). *P<0.05, **P<0.01 calculated by two-way repeated measures ANOVA with Bonferroni’s (a) and Dunnett’s (f,h,j) post-hoc tests, and two-way ANOVA with Sidak’s (b,g) and Dunnett’s (d,i) post-hoc tests, and one-way repeated measures ANOVA with Tukey’s post-hoc test (c,e). Graphs indicate mean ± s.e.m.
Extended Data Figure 5.. Histological identification of S1DZ.
a, Coronal section of the cortex counterstained with DAPI to highlight the abrupt reduction in cell density in layer 4 between the S1DZ and the S1BF at AP −0.46mm. (n=5, from 1 independent experiment). b, Coronal section of the cortex imaged with differential interference contrast further highlighting the reduced layer 4 in the S1DZ at AP −0.46mm (n=3, from 2 independent experiments). White arrows indicate borders of S1DZ. Scare bar represents 500μm (a) and 300μm (b). D: dorsal, V: ventral.
Extended Data Figure 6.. LPS treatment in MIA offspring does not have a distinguishable effect on c-Fos expression in other cortical regions analyzed.
a, Full cortical depth of S1DZ c-Fos staining as shown in Fig. 2a for PBS and MIA offspring after Veh or LPS administration **(**PBS-Veh n=8, PBS-LPS n=9, MIA-Veh n=13, MIA-LPS n=11; from 3 independent experiments). Scare bar represents 200μm. b,c, Representative images (b) and quantification (c) of c-Fos(Green)/NeuN(Red) co-labeled cells within the S1DZ of PBS and MIA offspring (PBS n=4, MIA n=3; from 1 independent experiment). Scale bar represents 50μm. d,e, Representative images (d) and quantification (e) of c-Fos expression in a series of cortical regions and following Veh or LPS injection in PBS or MIA offspring. Sections are stained for c-Fos (Green) and DAPI (Blue). Scale bar represents 200μm. S1BF, Primary somatosensory cortex, barrel field; M1, Primary motor cortex; M2, Secondary motor cortex; mPFC, medial prefrontal cortex (prelimbic and infralimbic cortex); AuD, Primary auditory cortex; V1, Primary visual cortex (For S1BF, M2, M1, and AuD; PBS-Veh n=8, PBS-LPS n=9, MIA-Veh n=13, MIA-LPS n=11. For mPFC; PBS-Veh n=8, PBS-LPS n=9, MIA-Veh n=12, MIA-LPS n=11. For V1; PBS-Veh n=7, PBS-LPS n=8, MIA-Veh n=12, MIA-LPS n=9; from 4 independent experiments). Statistics calculated by unpaired two-tailed t-test (c) and two-way ANOVA with Tukey’s post-hoc test (e). Graphs indicate mean ± s.e.m.
Extended Data Figure 7.. Further behavioral analyses of S1DZ optical inhibition-mediated rescue of sociability in monogenic mutant mice.
a, Quantification of c-Fos expressing cells in the S1DZ of monogenic mutant mice (WT n=6, Cntnap2 n=21, Fmr1 n=17, Shank3 n=15: from 5 independent experiments). b, Correlation of c-Fos expression in the S1DZ with severity of sociability deficits across monogenic mutant mice (Cntnap2 n=21, Fmr1 n=17, Shank3 n=15; from 4 independent experiments). Black solid lines represent regression line; grey lines indicate 90% confidence intervals. c, Individual data for experiments in Fig. 2f. d-f, Data expressed as time spent investigating social (S) versus inanimate (I) objects (d), total interaction time (e) and distance traveled (f) during three-chambered sociability experiments described in Fig. 2f (WT-EYFP n=7, WT-eNpHR n=8, Cntnap2-EYFP n=11, Cntnap2-eNpHR n=9, Fmr1-EYFP n=8, Fmr1-eNpHR n=12, Shank3-EYFP n=8, Shank3-eNpHR n=10; from 6 independent experiments). Statistics calculated by one-way ANOVA with Dunnett’s post-hoc test (a), linear regression (b), one-way repeated measures ANOVA with Dunnet’s post-hoc test (c), two-way ANOVA with Sidak’s post-hoc test (d), and two-way repeated measures ANOVA with Sidak’s post-hoc test (e,f). Graphs indicate mean ± s.e.m.
Extended Data Figure 8.. Il17ra expression in the S1DZ of PBS and MIA offspring. Further behavioral analyses of S1DZ IL-17a rescue of sociability in MIA offspring and monogenic mutant mice.
a, Representative images of Il17ra expression in the S1DZ of PBS and MIA offspring. Scare bar represents 1mm. b, Quantification of Il17ra expression within the S1DZ of MIA offspring according to cortical layer (n=6; from 2 independent experiments). c, Quantification of overall Il17ra expression in the S1DZ of PBS and MIA offspring (PBS n=8, MIA n=6; from 2 independent experiments). d-h, Further behavioral analyses of experiments described in Fig. 3f; Time spent investigating social (S) versus inanimate (I) objects (e), total interaction time (f), time spent in social (S), center (C) or inanimate (I) chamber (g), and distance traveled (h) (PBS-Veh n=11, PBS-IL-17a n=12, MIA-Veh n=14, MIA-IL-17a n=10, WT-Veh n=11, WT-IL-17a n=11, Cntnap2-Veh n=8, Cntnap2-IL-17a n=10, Fmr1-Veh n=9, Fmr1-IL-17a n=11; from 6 independent experiments). Statistics calculated by unpaired two-tailed t test (c), two-way ANOVA with Sidak’s (e) and Dunnett’s (g) post-hoc tests, and two-way repeated measures ANOVA with Sidak’s post-hoc test (f,h). Graphs indicate mean ± s.e.m.
Extended Data Figure 9.. IL-17a is necessary for LPS-induced behavioral rescue and reduction of c-Fos expression in MIA offspring.
a-e, Further behavioral analyses of experiments described in Fig. 4a; Time spent investigating social (S) versus inanimate (I) objects (b), total interaction time (c), time spent in social (S), center (C) or inanimate (I) chamber (d), and distance traveled (e) (PBS-Veh-Isotype n=9, PBS-LPS-Isotype n=11, MIA-Veh-Isotype n=10, MIA-LPS-Isotype n=10, MIA-LPS-αIL-17a n=10; from 7 independent experiments). f, Quantification of c-Fos expressing cells in the S1DZ and CeA following Veh or LPS injection in MIA offspring pretreated i.c.v. with isotype control antibody or blocking antibody against IL-17a (αIL-17a). Statistics calculated by two-way ANOVA with Sidak’s (b) and Dunnett’s (d) post-hoc tests, two-way repeated measures ANOVA with Sidak’s post-hoc tests (c,e), and one-way ANOVA with Dunnett’s post-hoc test (f). Graphs indicate mean ± s.e.m.
Extended Data Figure 10.. Further analyses of the necessity of IL-17a for the LPS-induced reduction of firing rate in the S1DZ, and the necessity of S1DZ IL-17Ra expression for the LPS-induced rescue of sociability deficits in MIA offspring.
a-c, Further analyses for experiments described in Fig 4d. a, Example of a head-fixed mouse on the running wheel used during single-unit recording. b, Representative image of a tetrode placement in the S1DZ. Scale bar represents 500μm. c, Firing rate for individual cells before and 4hrs after vehicle or LPS injection in PBS and MIA offspring pretreated with isotype control antibody or blocking antibody against IL-17a (αIL-17a). d-h, Further analyses for experiments described in Fig. 4e–f. Time spent investigating social (S) versus inanimate (I) objects (e), total interaction time (f), time spent in social (S), center (C) or inanimate (I) chambers (g), and distance traveled (h) (IL-17Rafl/fl;EYFP n=9, IL-17Rafl/fl;EGFP:nCre n=10; from 5 independent experiments). i-j, Representative images (i) and corresponding quantification (j) of Il17ra and Gapdh amplicon following PCR using cDNA derived from cells isolated from the cortical region centered on S1DZ of IL-17Rafl/fl;EYFP and IL-17Rafl/fl;EGFP:nCre mice (n=3 for all groups; from 1 experiment). Statistics calculated by two-way ANOVA with Sidak’s post-hoc test (e,g), two-way repeated measures ANOVA with Sidak’s post-hoc tests, (f,h) and unpaired two-tailed t test (j). Graphs indicate mean ± s.e.m.
Figure 1.. Immune stimulation rescues sociability deficits in MIA offspring.
a, Body temperature profile following Veh or LPS injection in PBS-offspring (Veh n=11, LPS n=11; from 5 independent experiments). Initial spike in body temperature is due to handling stress. Veh,Vehicle; LPS, lipopolysaccharide. b,c, Mice were tested for sociability (% time investigating social object / total time investigating both social and inanimate objects) one day prior to LPS injection (Pre). Mice were then tested for sociability four hours after either Veh or LPS injection (Test) (PBS-Veh n=10, PBS-LPS n=9, MIA-Veh n=10, MIA-LPS n=12, WT-Veh n=8, WT-LPS n=11, Cntnap2-Veh n=11, Cntnap2-LPS n=11, Fmr1-Veh n=11, Fmr1-LPS n=15, Shank3-Veh n=8, Shank3-LPS n=10; from 3 independent experiments). d, Virus encoding inhibitory DREADD (AAV2-hSyn-DIO-hM4D(Gi)-mCherry) was targeted to the vLPO of Vgat-Cre MIA mice. Scale bar represents 2mm. vLPO, ventral part of the lateral preoptic nucleus. e, Body temperature profile following Veh or CNO injection. f,g, Mice were tested for sociability one day prior to injection (Pre). The following two days, mice received counterbalanced injections of either Veh or CNO. Sociability was assessed two hours post-injection. For experiments d-g, n=9 for all groups; from 2 independent experiments. *P<0.05, **P<0.01 calculated by two-way repeated measures ANOVA with Bonferroni’s (a,e) and Sidak’s (c) post-hoc tests, and one-way repeated measures ANOVA with Tukey’s post-hoc test (g). Graphs indicate mean ± s.e.m.
Figure 2.. Immune stimulation reduces hyperactivation in the S1DZ of MIA offspring.
a, Representative images illustrating c-Fos (green) expression in the S1DZ and CeA following Veh or LPS injection. Scale bar represents 200μm. Numerals indicate cortical layers. S1DZ, Primary somatosensory cortex, dysgranular zone; CeA, Central amygdala. b,c, Quantification of c-Fos expressing cells in the S1DZ (b) and CeA (c) following Veh or LPS injection in the S1DZ (b) and CeA (c). For experiments a-c, PBS-Veh n=8, PBS-LPS n=9, MIA-Veh n=13, MIA-LPS n=11; from 3 independent experiments. d,e, AAV encoding either EYFP or EYFP fused to eNpHR was bilaterally injected into the S1DZ of monogenic mutant animals. Scale bar represents 500μm. f, Performance on sociability was assessed in the presence and absence of optical inhibition. For experiments e,f, WT-EYFP n=7, WT-eNpHR n=8, Cntnap2-EYFP n=11, Cntnap2-eNpHR n=9, Fmr1-EYFP n=8, Fmr1-eNpHR n=12, Shank3-EYFP n=8, Shank3-eNpHR n=10; from 6 independent experiments. g, Representative images illustrating c-Fos expression in the S1DZ and CeA following Veh or LPS injection in monogenic mutant mice. Scale bar represents 200μm. h,i, Quantification of c-Fos expressing cells following Veh or LPS injection in the S1DZ (h) and CeA (i). For experiments g-i, Cntnap2-Veh n=9, Cntnap2-LPS n=10, Fmr1-Veh n=7, Fmr1-LPS n=9, Shank3-Veh n=6, Shank3-LPS n=8; from 3 independent experiments. Statistics calculated by two-way ANOVA with Tukey’s post-hoc test (b,c) and Sidak’s post-hoc test (h,i), and two-way repeated measures ANOVA with Sidak’s post-hoc test (f). Graphs indicate mean ± s.e.m.
Figure 3.. IL-17a rescues sociability deficits in both MIA offspring and monogenic mutant mice.
a, IFN-γ, IL-6, IL-17a, and TNF-α levels in plasma following Veh or LPS injection (PBS-Veh n=6, PBS-LPS n=8, MIA-Veh n=6, MIA-LPS n=8, WT-Veh n=6, WT-LPS n=7, Cntnap2-Veh n=4, Cntnap2-LPS n=5, Fmr1-Veh n=5, Fmr1-LPS n=6, Shank3-Veh n=7, Shank3-LPS n=7; from 3 independent experiments). b, Il17ra expression in WT and IL-17Ra KO animals at AP −0.58mm from 1 independent experiment. Scale bar represents 1mm. c, Co-labeling of Il17ra and NeuN in the S1DZ from 2 independent experiments. Scale bar represents 100μm. d, Quantification of Il17ra expression within the S1DZ according to cortical layer in PBS offspring (n=8; from 2 independent experiments). e,f, Mice were tested for sociability one day prior to injection (Pre) and following bilateral administration of Veh or IL-17a into the S1DZ (Test). (PBS-Veh n=11, PBS-IL-17a n=12, MIA-Veh n=14, MIA-IL-17a n=10, WT-Veh n=11, WT-IL-17a n=11, Cntnap2-Veh n=8, Cntnap2-IL-17a n=10, Fmr1-Veh n=9, Fmr1-IL-17a n=11; from 6 independent experiments). Statistics calculated by two-way ANOVA with Dunnett’s post-hoc test (a) and two-way repeated measures ANOVA with Sidak’s post-hoc test (f). Graphs indicate mean ± s.e.m.
Figure 4.. IL-17a is necessary for LPS-induced rescue of sociability deficits and reduction of S1DZ neural activity in MIA offspring.
a, Mice were tested for sociability one day prior to injection (Pre). The following day blocking antibody against IL-17a (αIL-17a) or control isotype antibody was administered i.c.v. 30 minutes prior to Veh or LPS injection. Sociability was assessed four hours following Veh or LPS injection (Test) (PBS-Veh-Isotype n=9, PBS-LPS-Isotype n=11, MIA-Veh-Isotype n=10, MIA-LPS-Isotype n=10, MIA-LPS-αIL-17a n=10; from 7 independent experiments). b-d, Firing rate of S1DZ neurons before and four hours after Veh or LPS injection in PBS and MIA offspring pretreated with isotype control antibody or blocking antibody against IL-17a. c, Example raster plot with firing rate profile before (Baseline) and after (Test) LPS treatment from an PBS and MIA mouse pretreated with isotype control antibody. At time 0 mice began walking on the wheel rotating at 7.5cm/s. Data collected between 1–2 seconds were included in analysis. d, Normalized firing rate change following treatment represented as box-whisker plots indicating median, interquartile range, and data limits as defined by Tukey (PBS-Veh-Isotype n=65 cells, PBS-LPS-Isotype n=42 cells, PBS-LPS-αIL-17a n=40 cells, MIA-Veh-Isotype n=75 cells, MIA-LPS-Isotype n=48 cells, MIA-LPS-αIL-17a n=43 cells; from 2 MIA offspring and 2 PBS offspring in 12 independent experiments). e, Lentivirus encoding either EYFP or EGFP fused to nuclear Cre (nCre) was bilaterally injected into the S1DZ of IL-17Rafl/fl MIA offspring. Scale bar represents 200μm. f,g, Mice were tested for sociability one day prior to injection (Pre). The following day, mice were tested for sociability four hours after LPS injection (Test). For experiments e,f, IL-17Rafl/fl;EYFP n=9, IL-17Rafl/fl;EGFP:nCre n=10; from 5 independent experiments. Statistics calculated by two-way repeated measures ANOVA with Bonferroni’s post-hoc test (a,f) and two-way ANOVA with Tukey’s post-hoc test (d). Graphs indicate mean ± s.e.m.
Comment in
- Interleukin-17: A Social Cytokine.
Hoogenraad CC, Riol-Blanco L. Hoogenraad CC, et al. Cell. 2020 Apr 30;181(3):517-519. doi: 10.1016/j.cell.2020.03.060. Cell. 2020. PMID: 32359435
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