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A pancreas–hippocampus feedback mechanism regulates circadian changes in depression-related behaviors

Nature Neuroscience volume 28, pages 2078–2091 (2025)Cite this article

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Abstract

Individuals with neuropsychiatric disorders often show metabolic symptoms. However, the mechanisms underlying this co-occurrence remain unclear. Here we show that induced pluripotent stem cell-derived pancreatic islets from individuals with bipolar disorder have insulin secretion deficits caused by increased expression of RORβ, a susceptibility gene for bipolar disorder. Enhancing RORβ expression in mouse pancreatic β cells induced depression-related behaviors in the light phase and mania-like behaviors in the dark phase. Pancreatic RORβ overexpression in the light phase reduced insulin release from islets, inducing hippocampal hyperactivity and depression-like behaviors. Furthermore, this hippocampal hyperactivity in the light phase had the delayed effect of promoting insulin release in the dark phase, resulting in mania-like behaviors and hippocampal neuronal hypoactivity. Our results in mice point to a pancreas–hippocampus feedback mechanism by which metabolic and circadian factors cooperate to generate behavioral fluctuations and which may play a role in bipolar disorder.

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Fig. 1: Analysis of forebrain organoids derived from iPSCs of patients with BD or MDD.
Fig. 2: The iPSC-derived islet-like organoids of a patient with BD show insulin abnormalities.
Fig. 3: RORβ expression abnormalities in iPSC-derived islet-like organoids of patients with BD.
Fig. 4: Pancreatic activation of RORβ expression induces behavioral fluctuations in mice.
Fig. 5: RORβ activation reduces islet insulin release and induces depression-like behaviors.
Fig. 6: Reversed diurnal patterns of hippocampal neuronal activity in sg1/2 mice.
Fig. 7: Long-term hippocampal feedback to the pancreas induces insulin and behavioral inversion.
Fig. 8: Schematic showing the induction of diurnal fluctuation of behaviors in the sg1/sg2 mice via the pancreas–hippocampus feedback mechanism.

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Data availability

All statistical data are available in the main text or Supplementary Information. Uncropped RT–PCR and western blot gel images are available in Supplementary Information. RNA-seq data of neurons and islet-like organoids are available in the Gene Expression Omnibus database (accession number GSE283118). Source data are available from the corresponding author.

Change history

  • 28 August 2025

    In the version of Supplementary Information initially published alongside this article, due to figure preparation errors, there were duplications in Supplementary Fig. 1k, where the sample image of HCI#4 was mistakenly replaced with that of BDI#2 and in Supplementary Fig. 3b (left), where the flow cytometry image was mistakenly duplicated from Supplementary Fig. 3a (left). The figures are now updated in the Supplementary Information.

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Acknowledgements

We thank T. Xu (Institute of Biophysics, Chinese Academy of Science), C. Xu (Peking University) and Y. Chen (THU) for technical and material help. We thank all members of the laboratory for helpful discussions and technical assistance. This work was supported by the Beijing Natural Science Foundation (grant numbers Z210011 to J.Y. and Z200024), the National Natural Science Foundation of China (grant numbers 32371008 and 31830038), the Open Project of Collaborative Innovation Center for Language Ability of Jiangsu Province, China and the China Postdoctoral Science Foundation (grant number 2022TQ0182), and funding from Tsingha-Peking Center for Life Sciences.

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Author notes

  1. These authors contributed equally: Yao-Nan Liu, Qiu-Wen Wang, Xin-Yu She, Li-Jun Li.

Authors and Affiliations

  1. State Key Laboratory of Membrane Biology, IDG/McGovern Institute for Brain Research, School of Life Sciences, Tsinghua University, Beijing, China

    Yao-Nan Liu, Xin-Yu She, Bing Wang, Si-Yao Lu, Ying-Han Wang, Wei Shen, Chong-Lei Fu, Dan Li, Lan Yi & Jun Yao

  2. Laboratory of Genetics, The Salk Institute for Biological Studies, La Jolla, CA, USA

    Qiu-Wen Wang

  3. Department of Neuropsychiatry, Behavioral Neurology and Clinical Psychology, Beijing Tiantan Hospital, Capital Medical University, China National Clinical Research Centre for Neurological Diseases, Beijing, China

    Li-Jun Li & Chun-Xue Wang

  4. The Affiliated Brain Hospital, Guangzhou Medical University, Key Laboratory of Neurogenetics and Channelopathies of Guangdong Province and the Ministry of Education of China, Guangzhou Medical University, Guangzhou, China

    Ruilan Yang & Liping Cao

  5. China National Center for Bioinformation, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China

    Qian Li & Shuangli Mi

  6. University of Chinese Academy of Sciences, Beijing, China

    Qian Li & Shuangli Mi

  7. Key Laboratory for Biomechanics and Mechanobiology of Ministry of Education, School of Engineering Medicine, Beihang University, Beijing, China

    Wei Shi

  8. State Key Laboratory of Cell Biology, Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China

    Xin Cheng

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Contributions

Y.-N.L. Q.-W.W., C.-L.F., D.L. and X.C. conducted iPSC experiments. Y.-N.L., Q.-W.W. and B.W. conducted molecular biology and biochemistry experiments. Y.-N.L., B.W. and W. Shen conducted the animal behavioral experiments. Q.-W.W., S.-Y.L. and Y.-H.W. conducted electrophysiological experiments. L.-J.L., R.Y., C.-X.W. and L.C. conducted clinical investigations. X.-Y.S., Q.L. and S.M. conducted RNA-seq experiments. W. Shi and L.Y. provided technical assistance. Y.-N.L. and J.Y. analyzed the data. J.Y. conceived the project and designed the experiments. J.Y. wrote the paper with input from all the authors.

Corresponding author

Correspondence to Jun Yao.

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Extended data

Extended Data Fig. 1 Autonomous insulin signaling deficits in BDII iPSC-derived forebrain organoids.

a-c, qRT-PCR analysis showing the mRNA expression of IRS1 (a), INSR (b), and PI3K (c) in organoids before and after insulin treatment. Compared to the HCII, the BDII organoids show slightly reduced expression of these genes. In the HCII group, insulin treatment significantly decreases the PI3K expression. In the BDII group, insulin treatment increased IRS1 expression but reduced INSR expression. d, Immunoblot showing changes in InsR phosphorylation in BDII forebrain organoids. e-f, Quantitative analysis of InsR expression (e) and phosphorylation level (f). Compared to the HCII, the BDII group showed reduced levels of InsR protein expression and phosphorylation. In the HCII group, insulin treatment reduced InsR expression but enhanced InsR phosphorylation. In the BDII group, insulin treatment increased the InsR phosphorylation while the InsR protein expression remained unaffected. n = 3. One-way ANOVA with Sidak’s multiple comparisons test; error bars, s.e.m. Statistical data are listed in Supplementary Table 7.

Extended Data Fig. 2 RNA-seq and functional analysis of iPSC-derived islets.

a, Heatmap of selected differentially expressed genes between BDI and HCI islet-like organoids (left), and between HCII, BDII, and MDD islets (right). Values were shown as the Z score of log2(FPKM + 1) per gene. b, Table comparing the overlap of dysregulated KEGG pathways in BDI, BDII and MDD islet-like organoids. c, Venn plot showed enriched KEGG pathways based on differentially expressed genes in different groups. d-i, RNA-seq analysis showing the expression of islet marker genes, including SST (d), GCG (e), INS (f), INSR (g), NKX6.1 (h), and PDX1 (i), in the islet-like organoids of BDI/BDII/MDD subjects. n = 4 subjects (HCI), 6 (BDI), 5 (HCII/BDII/MDD). j,k, EM images (j) and quantification (k) of insulin-containing granules in mouse islet and HC/BD iPSC-derived islet-like organoids. Green arrowheads indicate insulin granules. n = 6 (mice), 9 (HC), 11 (BD). l, Mouse primary β-cells and HC iPSC-derived β-like cells showed similar levels of GSIS, n = 9 mice or HCI/II subjects. m, Sample Ca2+ imaging traces of mouse primary β-cells and HC/BDI/BDII iPSC-islets. (d-i) Two-tailed Student’s t-test; (k-l) One-way ANOVA with Sidak’s multiple comparisons test; *P < 0.05; error bars, s.e.m. Statistical data are listed in Supplementary Table 7.

Extended Data Fig. 3 RORβ expression abnormalities in BD patient iPSC-derived islet-like organoids.

a, b, qRT-PCR analysis showing RORβ mRNA expression in the islet-like (a) and forebrain (b) organoids of each MDD donor. n = 5 subjects. c, d, RNA-seq analysis of RORβ expression in the forebrain (c) and islet-like (d) organoids of each BDI (left) and BDII/MDD (right) donor. n = 4 (HCI), 6 (BDI), 3 (HCII/BDII/MDD) subjects. Left, two-tailed Student’s t-test; right, ANOVA test. e, Immunostaining images showing RORβ expression in insulin-expressing β-cells (upper), KRT19-expressing ductal cells (middle), and PDX1-expressing β/δ/progenitor cells (lower) in BDII islet-like organoids. Scale bar, 50 µm. f–h, Quantification of RORβ expression in insulin-expressing β-cells (f; n = 38 cells for HC, 15 cells for BD), KRT19-expressing ductal cells (g; n = 79/75), and PDX1-expressing β/δ/progenitor cells (h; n = 67/67) in BDII islet-like organoids. i, ELISA analysis of insulin secretion of HCI/II iPSC-derived islets overexpressing RORβ. Cells were challenged sequentially with 2 and 20 mM glucose for three rounds, with a 30 min incubation for each concentration, followed by a depolarization with 60 mM KCl. n = 8 subjects. j, Mean GSIS level of 7 stages in i. n = 7 stages. (a-b, f-j) Two-tailed Student’s t-test. *P < 0.05; error bars, s.e.m. Statistical data are listed in Supplementary Table 7.

Extended Data Fig. 4 RORβ shRNA reduces RORβ expression in BD patient iPSC-derived islet-like organoids.

a, b, Immunoblot (a) and quantification (b) of RORβ protein expression in BDI/BDII islet-like organoids expressing two human RORβ shRNAs, shRNA1 and shRNA2. n = 3 subjects. c, d, ELISA analysis showing glucose-stimulated insulin secretion (GSIS) in the BDI (c) and BDII (d) islet-like organoids expressing RORβ shRNA1. Left, low/high glucose and KCl stimulated insulin release. Cells were challenged sequentially with 2 and 20 mM glucose for three rounds, with a 30-minute incubation for each concentration, followed by a depolarization with 60 mM KCl. HCI/HCII, n = 4 subjects; BDI, n = 6; BDII, n = 5. Right, Mean GSIS. n = 7 stages. (b) One-way ANOVA with Sidak’s multiple comparisons test; (c, d) Two-tailed Student’s t-test; *P < 0.05; error bars, s.e.m. Statistical data are listed in Supplementary Table 7.

Extended Data Fig. 5 Expression pattern of AAV-based Cre/loxP system in Ins1CreERT mice.

a, Sample immunostaining of GFP showing the expression of rAAV-EF1a-DIO-EGFP in multiple organs of tamoxifen-activated Ins1CreERT mice. Scale bar, 100 μm. b-c, Immunoblots (b) and quantification (c) of RORβ protein expression in multiple organs of sg1/2 mice. n = 4. d-e, qRT-PCR analysis of dCas9 mRNA expression in the hippocampus (d) and cortex (e) of sg1/2 mice. PC, positive control with dCas9 expression. n = 3 (PC), 7 (Ctrl), 8 (sg1/2). (c) Two-tailed Student’s t-test; (d, e) One-way ANOVA with Sidak’s multiple comparisons test; *P < 0.05; **P < 0.001; error bars, s.e.m. Statistical data are listed in Supplementary Table 7.

Extended Data Fig. 6 Reversed diurnal patterns of insulin function in sg1/2 mice.

a, Diurnal plasma insulin changing patterns of control (upper) and sg1/2 (lower) mice. b, Diurnal pattern of GSIS in the control and sg1/2 mice. The GSIS was determined by the ratio of glucose-stimulated versus resting condition insulin concentration. (a, b) Ctrl, from ZT5 to ZT1, n = 4/4/4/4/3/3 mice; sg1/2, n = 4/3/3/4/4/4. c-f, Immunoblot (c,e) and quantification (d,f) of insulin expression and InsR phosphorylation levels in the mouse hippocampus at the insulin peak (ZT19, 14 pm) and nadir (ZT11, 6am) of the sg1/2 mice. n = 4. g, h, Total distance of sg1/2 mice in the light (g; n = 12) and dark (h; Ctrl, n = 12; sg1/2, n = 11) periods in the open field test (OFT). i, Schematic showing injection of rAAV-EF1a-DIO-CRISPRa-mRORβ into the ventricles of Ins1CreERT mice. j, k, FST (j) and SPT (k) of vsg1/vsg2 mice in the light (white) and dark (gray) phases. n = 12. l, Immunostaining of brain sections showing that in the mice receiving rAAV-EF1a-DIO-EGFP injected into the ventricles, EGFP expression was detected in the lateral ventricles (LV) but not in the hippocampus. (a, j, k) One-way ANOVA with Sidak’s multiple comparisons test; other experiments, two-tailed Student’s t-test was used; *P < 0.05; **P < 0.001; error bars, s.e.m. Statistical data are listed in Supplementary Table 7.

Extended Data Fig. 7 Circadian rhythm of the sg1/2 mice.

a, Representative single-plotted actograms of voluntary wheel-running activity in light-dark (LD) and dark-dark (DD) cycles of the control and sg1/2 mice. Mice were housed in 12 L:12D for 14 days, followed by a shift to constant darkness for 7 days. White shading indicates lights on, while gray shading represents lights off. b, c, Average counts (per 6 min) of wheel-running activity in LD (b) over 14 days or in DD (c) over 7 days. The counts of the sg1/2 mice were increased compared to the control mice (n = 5). d, Average circadian free-running periods of the control and sg1/2 mice measured by chi-square periodogram in DD over 7 days (n = 5). The control and sg1/2 mice showed a similar length of period. e, Average amplitude of locomotor activity rhythm was analyzed by fast Fourier transform (FFT) in DD over 7 days. The amplitude of the sg1/2 mice was significantly increased compared to the control mice (n = 5). (b, c) Two-way ANOVA with Sidak’s multiple comparisons test; (d, e) Two-tailed Student’s t-test; *P < 0.05; error bars, s.e.m. Statistical data are listed in Supplementary Table 7.

Extended Data Fig. 8 Effects of manipulating islet insulin function on the brain insulin signaling and behaviors of the sg1/2 mice.

a, Plasma insulin level of sg1/2 mice during the light period. Ctrl, n = 22; sg1/2, n = 24. b, Effects of hippocampal infusion of insulin on the light-phase behaviors of the sg1/2 mice. n = 13 (FST), 11 (SPT). c, Effects of hippocampal infusion of BMS-536924 (BMS), an InsR blocker, on the light-phase behaviors of the sg1/2 mice. n = 13 (FST), 11 (SPT). d, Effects of hippocampal infusion of BMS-536924 on the plasma insulin concentration of the sg1/2 mice. n = 5 (saline), 5 (15 min), 4 (30 min). e, Immunoblot (left) and quantification of InsR expression (middle) and phosphorylation (right) in the hippocampus of the sg1/2 mice treated with repaglinide (RPG; 50 μg/kg), an anti-diabetic drug, through intraperitoneal injection (sg1/2 + RPG). n = 3. (a–c,e) Two-tailed Student’s t-test; (d) One-way ANOVA with Sidak’s multiple comparisons test; *P < 0.05; **P < 0.001; error bars, s.e.m. Statistical data are listed in Supplementary Table 7.

Extended Data Fig. 9 Effects of InsR knockdown on the behaviors and peripheral insulin secretion of the sg1/2 mice.

a, sg1/2 mice with InsR knockdown (KD) in the hippocampus showed slightly reduced sucrose preference (left, n = 14 (Ctrl), 13 (InsR KD)), slightly increased FST immobility time (middle, n = 13/15), and significantly decreased LDB time-in-light (right, n = 12/15) in the dark phase. Ctrl, sg1/2 mice; InsR KD, sg1/2 mice with InsR KD. b, sg1/2 mice with InsR KD in the hippocampus showed significantly reduced sucrose preference (left, n = 13/12), increased FST immobility time (middle, n = 15/15), and decreased LDB time-in-light (right, n = 14/15) in the light phase. c, ELISA analysis of plasma insulin concentration in the sg1/2 mice with InsR KD in the hippocampus. n = 5. (a, b) Two-tailed Student’s t-test; (c) One-way ANOVA with Sidak’s multiple comparison test; *P < 0.05; **P < 0.001; error bars, s.e.m. Statistical data are listed in Supplementary Table 7.

Extended Data Fig. 10 Long-term effects of optical stimulations on InsR phosphorylation.

a, b, Long-term effects of a 6-hr ChR2 optical activation on the hippocampal InsR phosphorylation level of the sg1/2 (a) and control (b) mice at 6 hr after the 6-hr stimulation. D: dark phase; L: light phase. Left, immunoblot; right, quantification. ChR2, channelrhodopsin-2; NpHR, halorhodopsin. n = 4 mice for all groups. c-d, Long-term effects of a 6-hr NpHR optical suppression on the hippocampal InsR phosphorylation level of the control (c) and sg1/2 (d) mice at 6 hr after the 6-hr suppression. n = 4. Two-tailed Student’s t-test was used; *P < 0.05; error bars, s.e.m. Statistical data are listed in Supplementary Table 7.

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Liu, YN., Wang, QW., She, XY. et al. A pancreas–hippocampus feedback mechanism regulates circadian changes in depression-related behaviors. Nat Neurosci 28, 2078–2091 (2025). https://doi.org/10.1038/s41593-025-02040-y

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