18β-Glycyrrhetinic Acid Ameliorates Neuroinflammation Linked Depressive Behavior Instigated by Chronic Unpredictable Mild Stress via Triggering BDNF/TrkB Signaling Pathway in Rats

Abstract

Evidence shows that inflammatory responses may encompass the onset of severe depressive illness. Traditionally used licorice contains 18β-glycyrrhetinic acid (18βGA), which has been demonstrated to reduce inflammation and oxidative stress. This study investigates the antidepressant effects of 18βGA and the underlying mechanism in rats exposed to chronic unpredictable mild stress (CUMS). Wistar rats were exposed to CUMS for 36 consecutive days to establish depression. 18βGA (10, 20, and 50 mg/kg) or fluoxetine was given once daily (from day 30 to day 36). Thereafter, behavior parameters (sucrose preference test, forced-swimming test, open-field test, body weight), pro-inflammatory cytokines, neurotransmitters, adrenocorticotropic hormone (ACTH), corticosterone (CORT), and liver biomarkers were studied. Immunohistochemistry and western blot analyses were conducted to investigate the protein's expression. 18βGA (20 and 50 mg/kg) treatment increased sucrose intake, locomotion in the open-field test, decreased immobility time in the forced swim test, and improved body weight in CUMS-exposed rats. The therapy of 18βGA dramatically declined cytokines, ACTH and CORT and improved 5HT and norepinephrine in CUMS rats. Furthermore, BDNF and TrkB proteins were down-regulated in CUMS group, which was increased to varying degrees by 18βGA at doses of 20 and 50 mg/kg. Therefore, 18βGA ameliorates depressive-like behavior persuaded by chronic unpredictable mild stress, decreases neuroinflammation, liver biomarkers, stress hormones, and improves body weight, brain neurotransmitter concentration via activating on BDNF/TrkB signaling pathway in both PFC and hippocampus in rats.

Keywords: 18β-Glycyrrhetinic acid; Brain-derived neurotrophic factor; Chronic unpredictable mild stress; Depression; Tropomyosin receptor kinase B.

Fig. 1

In the OFT, 18βGA therapy influences square crossing numbers (A), rearing number (B), grooming counts (C), total distance traveled (D), frequency of entering the center (E) in the CUMS and non-CUMS rats. Percent weekly weight changes (F) after 18βGA therapy are also presented in the CUMS and non-CUMS rats. The results are displayed as mean ± S.E.M. of 6 animals in each group with ^###p < 0.001, ^####p < 0.0001 vs non-CUMS rats and *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 vs CUMS rats, “one-way (A–E) and two-way (F) analysis of variance TMC test”. CUMS stands for chronic unpredictable mild stress; 18βGA stands for 18β-glycyrrhetinic acid; OFT is for the open field test; i.g. stands for intragastrically

Fig. 2

The effects of CUMS and 18βGA on the FST immobility time (A) and sucrose preference % (B). Rats were exposed to the FST and SPT after receiving 18βGA (10, 20, and 50 mg/kg, intragastrically), FXT (10 mg/kg, intragastrically), or vehicle. The results are displayed as mean ± S.E.M. of 6 animals in each group with ^###p < 0.001, ^####p < 0.0001 vs non-CUMS rats and *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 vs CUMS rats, “one-way analysis of variance then TMC test”. CUMS stands for chronic unpredictable mild stress; 18βGA stands for 18β-glycyrrhetinic acid; FST stands for forced swimming test; SPT is for sucrose preference test; i.g. stands for intragastrically

Fig. 3

Effects of CUMS and 18βGA on serum markers of the liver (A), ACTH (B), and CORT (C). The results are displayed as mean ± S.E.M. of 6 animals in each group with ^##p < 0.01, ^####p < 0.0001 vs non-CUMS rats and *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 vs CUMS rats, “one-way (ACTH and CORT) and two-way (liver markers) analysis of variance then TMC test”. ALP stands for alkaline phosphatase; CUMS stands for chronic unpredictable mild stress; ALT stands for alanine aminotransferase; 18βGA stands for 18β-glycyrrhetinic acid; i.g. stands for intragastrically; AST stands for aspartate aminotransferase; ACTH stands adrenocorticotropic hormone and CORT for corticosterone

Fig. 4

The effects of CUMS and 18βGA on IL-1beta, IL-6, and TNF-alpha in plasma are shown in Fig. 4. The results are presented as mean ± S.E.M. of 6 animals in each group with ^####p < 0.0001 vs non-CUMS rats and *p < 0.05, **p < 0.01, ****p < 0.0001 vs CUMS rats, “two-way analysis of variance then TMC test”. TNF-alpha, tumor necrosis factor; CUMS stands for chronic unpredictable mild stress; 18βGA stands for 18β-glycyrrhetinic acid; IL-1beta stands for interleukin-1 beta; IL-6, interleukin-6; i.g. stands for intragastrically

Fig. 5

The effects of 18βGA on norepinephrine (A) and serotonin (B) levels in the PFC and the hippocampus in CUMS and non-CUMS rats. The results are displayed as mean ± S.E.M. of 3 animals in each group with ^####p < 0.0001 vs non-CUMS rats and *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 vs CUMS rats, “two-way analysis of variance then TMC test”. CUMS stands for chronic unpredictable mild stress; 18βGA stands for 18β-glycyrrhetinic acid; i.g. stands for intragastrically

Fig. 6

Effects of CUMS and 18βGA at doses 10, 20, and 50 mg/kg intragastrically and FXT at dose 10 mg/kg intragastrically on relative BDNF protein expression by IHC staining in the CA1 area. The scale bar represents 25 μm. The results are displayed as mean ± S.E.M. of 3 animals in each group with ^####p < 0.0001 vs. non-CUMS rats and *p < 0.05, ****p < 0.0001 vs. CUMS rats, “one-way analysis of variance then TMC test.” CUMS stands for chronic unpredictable mild stress; 18βGA stands for 18β-glycyrrhetinic acid; BDNF is a brain-derived neurotrophic factor; i.g. stands for intragastrically

Fig. 7

Effects of CUMS and 18βGA on the PFC’s relative BDNF protein expression by immunohistochemistry staining. The scale bar represents 25 μm. The results are displayed as mean ± S.E.M. of 3 animals in each group with ^####p < 0.0001 vs. non-CUMS rats and *p < 0.05, **p < 0.01 vs. CUMS rats, “one-way analysis of variance post-hoc TMC test.” CUMS stands for chronic unpredictable mild stress; 18βGA stands for 18β-glycyrrhetinic acid; BDNF is a brain-derived neurotrophic factor; i.g. stands for intragastrically

Fig. 8

Effects of CUMS and 18βGA at doses 10, 20, and 50 mg/kg, intragastrically and FXT at dose 10 mg/kg, intragastrically on BDNF (A) and TrkB (B) protein expression by Western blotting analysis in the PFC and the hippocampus. Representative immunoblot images of BDNF, TrkB, and β-actin. The results are displayed as mean ± S.E.M. of 3 animals in each group with ^####p < 0.0001 vs. non-CUMS rats and *p < 0.05, **p < 0.01 vs. CUMS rats, “two-way analysis of variance post-hoc TMC test.” CUMS stands for chronic unpredictable mild stress; 18βGA stands for 18β-glycyrrhetinic acid; BDNF is a brain-derived neurotrophic factor; TrkB stands for tropomyosin receptor kinase B; i.g. stands for intragastrically