Neurogenesis-dependent antidepressant-like activity of Hericium erinaceus in an animal model of depression

Abstract

Background: Depression is a severe neuropsychiatric disorder that affects more than 264 million people worldwide. The efficacy of conventional antidepressants are barely adequate and many have side effects. Hericium erinaceus (HE) is a medicinal mushroom that has been reported to have therapeutic potential for treating depression.

Methods: Animals subjected to chronic restraint stress were given 4 weeks HE treatment. Animals were then screened for anxiety and depressive-like behaviours. Gene and protein assays, as well as histological analysis were performed to probe the role of neurogenesis in mediating the therapeutic effect of HE. Temozolomide was administered to validate the neurogenesis-dependent mechanism of HE.

Results: The results showed that 4 weeks of HE treatment ameliorated depressive-like behaviours in mice subjected to 14 days of restraint stress. Further molecular assays demonstrated the 4-week HE treatment elevated the expression of several neurogenesis-related genes and proteins, including doublecortin, nestin, synaptophysin, brain-derived neurotrophic factor (BDNF), tropomyosin receptor kinase B (TrkB), phosphorylated extracellular signal-regulated kinase, and phosphorylated cAMP response element-binding protein (pCREB). Increased bromodeoxyuridine-positive cells were also observed in the dentate gyrus of the hippocampus, indicating enhanced neurogenesis. Neurogenesis blocker temozolomide completely abolished the antidepressant-like effects of HE, confirming a neurogenesis-dependent mechanism. Moreover, HE induced anti-neuroinflammatory effects through reducing astrocyte activation in the hippocampus, which was also abolished with temozolomide administration.

Conclusion: HE exerts antidepressant effects by promoting neurogenesis and reducing neuroinflammation through enhancing the BDNF-TrkB-CREB signalling pathway.

Keywords: Anti-neuroinflammatory; Depression; Hericium erinaceus; Hippocampus; Neurogenesis.

Fig. 1

Structure of adenosine (1), herierin III (2), and herierin IV (3) (A). The compound-target network. The purple diamonds represent the 8 previously reported active compounds in HE. The green rectangles represent gene targets retrieved from PubChem database (B). Genes related to depression were retrieved from DisGeNET and compared with HE gene targets. Overlapped gene targets are shown (C)

Fig. 2

Bubble maps of GO annotation analysis (A) and KEGG pathway analysis (B). The Y axis shows the GO terms and the names of the enriched pathways. The area of the bubble represents the number of enriched genes. Adjusted p-value represented by a colour scale, with the statistical significance increasing from blue to red (blue represents low significance and red represents high significance)

Fig. 3

Schematic representation of the study of the effects of HE in the CRS model of depression (A). The assessment of anxiety-like behaviour by escape latency in the cage emergence test (B) and by latency to feed in the novelty suppressed feeding test. C The evaluation of depressive-like behaviour by percentage sucrose preference (D) and by immobility time in the tail suspension test (E). The results are presented as mean ± S.E.M. Indicators: *Significant difference between the CRS groups. *p ≤ 0.05. **p ≤ 0.01; ***p ≤ 0.001

Fig. 4

Effects of HE on the relative expression of neuroplasticity- and neurogenesis-related genes including Bdnf (A), Trkb (B), NeuN (C), Dcx (D), Syp (E), Nes (F), Creb (G), and Psd-95 (H) in the dorsal hippocampus of non-CRS + saline, CRS + saline, CRS + 10 mg/kg HE and CRS + 25 mg/kg HE groups. BDNF was significantly downregulated in the CRS + saline group. Notably, 10 and 25 mg/kg HE increased the expressions of Bdnf, Syp, Nes, and Psd-95. Relative expression was calculated by normalising the relative quantifications to the reference gene GAPDH as a ratio of the 2^CT(reference) and 2^CT(test). *p ≤ 0.05. **p ≤ 0.01

Fig. 5

Western blot analysis of proteins associated with BDNF/TrkB signalling in dorsal hippocampal tissue from non-CRS + saline, CRS + saline, CRS + 10 mg/kg HE, and CRS + 25 mg/kg HE groups. Note, dividing lines represent cropped blots and white spaces represent cropped bands. Graphical representation of the effects of HE on the expression of neuroplasticity-related proteins (A) including TrkB (B), pTrkB (C), pro-BDNF (D), mature BDNF (E), CREB (F), and pCREB (G). The expression of target proteins was normalised to the expression of GAPDH. Note, there were significant increases in the expression of pTrkB, mBDNF, and pCREB proteins in the CRS + 25 mg/kg HE group compared with the CRS + saline group. *p ≤ 0.05. **p ≤ 0.01

Fig. 6

Western blot analysis of proteins associated with CREB phosphorylation in dorsal hippocampal tissue from non-CRS + saline, CRS + saline, CRS + 10 mg/kg HE, and CRS + 25 mg/kg HE groups. Note, dividing lines represent cropped blots and white spaces represent cropped bands. Blot images (A) and graphical representation of the effects of HE on the expression of neuroplasticity-related proteins including ERK1/2 (B), pERK1/2 (C), AKT (D), and pAKT (E). The expression of target proteins was normalised to the expression of GAPDH. Note, in CRS + 10 mg/kg HE group, there was a significant increase in pERK1/2 protein expression compared with the CRS + saline group and increase in pAKT compared to the non-CRS control group. *p ≤ 0.05

Fig. 7

Immunofluorescence staining of BrdU and NeuN in the DG reveals a dosage-dependent effect of HE treatment on neurogenesis. Scale bar: 100 µm (A). Note, there was a significant increase in BrdU⁺ cells in the DG of CRS animals receiving 10 mg/kg HE, but not 25 mg/kg HE (B). Double labelling of BrdU and NeuN showed deceased neuronal maturation in the CRS + 25 mg/kg HE group (C). **p ≤ 0.01; ***p ≤ 0.001

Fig. 8

Graphical representation of the behavioural data in TMZ experimental groups. No significant differences were found in all the behavioural testing for the assessment of anxiety-like behaviour including cage emergence test (A) and novelty suppressed feeding test (B), and depressive-like behaviour including sucrose preference test (C) and tail suspension test (D). Scatter plots of the correlations between the depression-related behavioural tests in the non-TMZ (E) and TMZ experimental groups (F). Notably, in the non-TMZ experimental group, there were significant correlations between the sucrose preference and immobility time in CRS + 10 mg/kg HE, CRS + 25 mg/kg HE, and non + CRS control groups. No correlation was found between the sucrose preference and immobility time in CRS + saline group or all TMZ experimental groups. p ≤ 0.05 considered statistically significant

Fig. 9

Immunohistochemical analysis of GFAP (astrocyte marker) in the hippocampus of non-CRS + saline, CRS + saline, CRS + 10 mg/kg HE, and CRS + 25 mg/kg HE group. Scale bar: 200 µm (hippocampus) and 50 µm (subregions) (A). The degree of astrocyte activation was examined in the CA1 (B), CA3 (C), and DG (D), separately. Notably, the CRS + saline group exhibited increased astrocyte activation in the CA1 and DG, which was suppressed by 10 and 25 mg/kg HE. To probe whether the observed suppression was mediated by hippocampal neurogenesis, the hippocampus of TMZ-treated animals was stained with GFAP in the same subregions. TMZ administration abolished the suppressive effects of HE (E–H)