Kai-Xin-San, a traditional Chinese medicine formulation, exerts antidepressive and neuroprotective effects by promoting pCREB upstream pathways

Abstract

Kai-Xin-San (KXS) is a traditional Chinese medicine that has been widely used for the treatment of emotion-related disease. However, the underlying mechanism remains largely unknown. The present study aimed to examine whether phospho-cAMP response element-binding protein (pCREB) and upstream components, such as extracellular signal-regulated kinase (ERK), phospho-ERK (pERK), phosphatidylinositol-3-kinase (PI3K), protein kinase B (Akt), glycogen synthase kinase 3β (GSK3β) and pGSK3β are associated with the antidepressive effect of KXS. In total, 24 male Wistar rats were randomly divided into three groups, including control (n=8, no treatment), induced with chronic unpredictable mild stress (CMS) (n=8), and CMS rats treated with KXS at dosage of 370 mg/kg/day orally. Primary hippocampal neuronal cultures were prepared from Wistar rats for cell survival and proliferation assays. In KXS rats, increased protein expression levels of pCREB, BDNF and tyrosine receptor kinase B (TrkB) were observed in the hippocampus and prefrontal cortex, compared with the CMS model group. Furthermore, increased expression levels of ERK, pERK, PI3K, Akt, and GSK3β were also detected in the hippocampus and prefrontal cortex of KXS-treated rats compared with CMS model rats and in primary hippocampal neuronal cells treated with KXS. These results suggest that pCREB and upstream components, including TrkB/ERK/CREB and TrkB/PI3 K/CREB, may contribute to the antidepressive effect induced by KXS. Further studies are required to confirm these findings.

Keywords: Kai-Xin-San; antidepressant effect; brain-derived neurotrophic factor; extracellular signal-regulated kinase signal pathway; traditional Chinese medicinal formula.

Figure 1.

Effects of KXS on the TrkB/BDNF signaling pathway in the control and CMS rats. KXS (370 mg/kg) was given daily for three weeks. Data are presented as the mean ± standard deviation. *P<0.05 vs. the control group, ^##P<0.01 vs. the model group. CMS, chronic unpredictable mild stress; KXS, Kai-Xin-San; BDNF, brain-derived neurotrophic factor; p, phospho; CREB, cAMP response element-binding protein; TrkB, tyrosine receptor kinase B.

Figure 2.

Effects of KXS on the TrkB/BDNF/ERK and TrkB/BDNF/PI3K signaling pathway in CMS rats. KXS (370 mg/kg) was administered daily for three weeks. Data are presented as the mean ± standard deviation, *P<0.05 vs. the control group; ^##P<0.01 and ^#P<0.05 vs. the CMS group. CMS, chronic unpredictable mild stress; KXS, Kai-Xin-San; BDNF, brain-derived neurotrophic factor; Akt, protein kinase B; p, phospho; ERK, extracellular signal-regulated kinase; TrkB, tyrosine receptor kinase B; GSK3β, glycogen synthase kinase 3β; PI3K, phosphatidylinositol-3-kinase.

Figure 3.

Morphological observation and identification of primary hippocampal neurons (magnification, ×200) at (A) 4 h, (B) 24 h, (C) 2 days, (D) 3 days, (E) 5 days, (F) 7 days. Primary hippocampal neurons incubated with (G) DIPA and MAP-2 at (H) day 5 and (I) day 7.

Figure 4.

Effect of KXS on cell survial and BDNF release from primary hippocampal neurons. (A) For cell viability assay, cells were treated with 25 pg/l BDNF or 50, 100 and 200 µg/ml of KXS for 48 h. **P<0.01 vs. the control group. (B) In the BDNF release test, cultured cells were treated with 100 µg/ml KXS for 0, 3, 6, 12, 24 and 48 h. **P<0.01 vs. the 0 h group. Data are presented as the mean ± standard deviation. KXS, Kai-Xin-San; BDNF, brain-derived neurotrophic factor; OD, optical density.

Figure 5.

KXS rapidily activates the TrkB/ERK signaling pathway. Effects of K252a on the KXS (100 µg/ml for 48 h) -induced TrkB and ERK phosphorylation or BDNF expression were evaluated by western blot. Data are presented as the mean ± standard deviation **P<0.01 vs. the basal group,^##P<0.01 vs. the KXS alone. KXS, Kai-Xin-San; BDNF, brain-derived neurotrophic factor; ERK, extracellular signal-regulated kinase; TrkB, tyrosine receptor kinase B.