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Gut Serotonergic and GABA Signaling in Mouse Model of Maternally Inherited UBE3A Impairment

Altered serotonergic and GABA signaling in the gut were observed in a mouse model of impaired maternally inherited UBE3A, mimicking features of Angelman Syndrome. The study highlighted the connection between gut-brain impairments in AS-modeled mice, implicating changes in gastrointestinal motility, serotonin, and GABA signaling. These findings shed light on potential mechanisms contributing to the gastrointestinal issues observed in Angelman Syndrome.

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Gut Serotonergic and GABA Signaling in Mouse Model of Maternally Inherited UBE3A Impairment

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  1. Altered serotonergic and GABA signaling in the gut in a mouse model of impaired maternally inherited UBE3A Hyun Min Shin, Jelissa Reynoso-Garcia, and Melanie G Gareau University of California, Davis

  2. Overview I- Angelman Syndrome - Loss of function mutation of maternal Ubiquitin Ligase E3A (UBE3A) - Genomic imprinting of paternal chromosome in neurons - Deficient neuronal UBE3A - UBE3A codes for ubiquitin ligase E6- Associated Protein (E6-AP) critical for ubiquitin-proteasome pathway function - Impaired degradation of proteins

  3. Overview II - AS - Neurological impairments - Neurodevelopmental delays - Deficits in movement and coordination - Seizures - Concurrent Gastrointestinal issues - Constipation - Poor feeding due to hypotonia of throat - Reflux

  4. Gut microbiota dysbiosis

  5. Hypothesis: Gut-brain impairments, including gastrointestinal motility and altered serotonergic and GABA signaling, will be present in AS- modeled mice

  6. Serotonin and GABA Serotonin ● Most of 5-hydroxytryptamine (5-HT) released by enterochromaffin cells in the gut ● 5-HT is important in gut propulsive and segmentation motility & epithelial secretion ● Synthesized by Tph1 & Tph2 (tryptophan hydroxylases) and recycled by serotonin reuptake transporter (SERT) ● 5-HT producing enteric bacteria GABA ● Main inhibitory neurotransmitter in the body ● Widespread distribution of GABA receptors throughout gut ● Role in gastrointestinal motility, gastric emptying, & gastric acid secretion ● GABA producing enteric bacteria

  7. Mouse model of AS: Ube3am-/p+ Parents F1 F2 ♀︎ ♀︎ F1- HET: Ube3am+/p- WT: Ube3am+/p+ F2- AS: Ube3am-/p+ ♂︎ ♂︎ WT: Ube3am+/p+ AS: Ube3am-/p+ WT: Ube3am+/p+

  8. Methods Ussing Chamber - Gut tissues (ileum & distal colon) of AS & WT mice analyzed for - Isc- active ion transport - G- tight junction permeability - FITC- macromolecular permeability qPCR - Ileum, distal colon, HC, PFC tissues of AS & WT mice analyzed for - Serotonergic signaling - GABA signaling

  9. Decreased active ion transport in AS mice in the colon 80 lsc - ileum 150 Isc - colon ** 60 Isc (mS/cm2) Isc (mS/cm2) 100 40 50 20 0 0 WT HET (ube3a m+/p-) AS WT HET (ube3a m+/p-) AS (ube3a m+/p+) (ube3a m-/p+) (ube3a m+/p+) (ube3a m-/p+) Jelissa Reynoso-Garcia

  10. Increased tight junction permeability in AS mice in the ileum 100 40 G ileum G - colon * 30 G (mS/cm2) G (mS/cm2) 20 50 10 0 0 WT HET (ube3a m+/p-) AS WT HET (ube3a m+/p-) AS (ube3a m+/p+) (ube3a m-/p+) (ube3a m+/p+) (ube3a m-/p+) Jelissa Reynoso-Garcia

  11. Macromolecular permeability was increased in AS mice in both ileum and colon 8000 4000 FITC flux FITC flux FITC flux (ng/ml/cm2/h) FITC flux (ng/ml/cm2/h) 6000 3000 * 4000 2000 ** 2000 1000 0 0 WT HET (ube3a m+/p-) AS WT HET (ube3a m+/p-) AS (ube3a m+/p+) (ube3a m-/p+) (ube3a m+/p+) (ube3a m-/p+) Jelissa Reynoso-Garcia

  12. Altered serotonergic and GABA signaling in the distal colon of AS mice GABA Serotonin 5HTR2c GABAb1b GABAa2a 2.0 3 2.0 2.0 5HTR3a Sert Tph2 Tph1 3 * * 3 5 (normalized to b-actin) (normalized to b-actin) (normalized to b-actin) * * * (normalized to b-actin) * 1.5 1.5 1.5 Fold expression Fold expression Fold expression (normalized to b-actin) (normalized to b-actin) 4 Fold expression 2 2 Fold expression Fold expression 2 3 1.0 1.0 1.0 2 1 1 1 0.5 0.5 0.5 1 0.0 0.0 0.0 0 0 0 0 WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS (ube3a m-/p+) (ube3a m-/p+) (ube3a m-/p+) (ube3a m-/p+) (ube3a m-/p+) (ube3a m-/p+) (ube3a m-/p+)

  13. Altered GABA signaling in the ileum of AS mice Serotonin GABA TPH1 ileum TPH2 ileum GABAA1a GABAB1a GABAB2B SERT ileum 3 3 * 3 5 GABAb1b 3 3 * 3 * (normalized to b-actin) (normalized to b-actin) * (normalized to b-actin) (normalized to b-actin) (normalized to b-actin) 4 (normalized to b-actin) (normalized to b-actin) Fold expression * Fold expression Fold expression Fold expression Fold expression Fold expression Fold expression 2 2 2 2 2 2 3 2 1 1 1 1 1 1 1 0 0 0 0 0 0 0 WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS (ube3a m-/p+) (ube3a m-/p+) (ube3a m-/p+) (ube3a m-/p+) (ube3a m-/p+) (ube3a m-/p+) (ube3a m-/p+)

  14. Altered serotonergic and GABA signaling in the hippocampus of AS mice Serotonin GABA tph2 sert GABAb1b GABAa2a 3 3 3 3 * (normalized to b-actin) (normalized to b-actin) (normalized to b-actin) (normalized to b-actin) * * Fold expression Fold expression Fold expression Fold expression 2 2 2 2 p=0.06 1 1 1 1 0 0 0 0 WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS (ube3a m-/p+) (ube3a m-/p+) (ube3a m-/p+) (ube3a m-/p+)

  15. Evidence of neuroinflammation and increased neurogenesis in the hippocampus of AS mice Iba1 Bdnf 4 5 * * (normalized to b-actin) (normalized to b-actin) 4 3 Fold expression Fold expression 3 2 2 1 1 0 0 WT (ube3a m+/p+) AS WT (ube3a m+/p+) AS (ube3a m-/p+) (ube3a m-/p+)

  16. Summary - Decreased secretory state in colon coupled with increased permeability in colon & ileum in AS mice is suggestive of altered motility and permeability in the gut Altered serotonin and GABA gene expression in the gut and brain in AS mice is suggestive of altered neuronal signaling -

  17. Conclusion AS mice show evidence of constipation mediated by altered enteric nerves, which may provide insight to the mechanisms of constipation in AS patients.

  18. Future directions and speculations - Administration of Tributyrin (butyrate) as a therapeutic strategy to improve GI symptoms and MGB axis signaling by beneficially modulating the gut microbiome in AS - Short-chain fatty acid (SCFA) that beneficially modulates the gut microbiome - Determine whether these findings are common across other neurodevelopmental/degenerative diseases - Provide a better understanding of the signaling pathways involved in constipation across species

  19. Acknowledgements Research Funding: Special Thanks to- Melanie Gareau, PhD FT2022-007 Jelissa Reynoso-Garcia, PhD Student Funding: Olivia Orahood Students Training in Advanced Research (STAR) Program T35 OD010956

  20. Questions?

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