Tacrine(10)-Hupyridone Prevents Post-operative Cognitive Dysfunction via the Activation of BDNF Pathway and the Inhibition of AChE in Aged Mice

Abstract

Post-operative cognitive dysfunction (POCD) could cause short-term or long-term cognitive disruption lasting weeks or months after anesthesia and surgery in elderly. However, no effective treatment of POCD is currently available. Previous studies indicated that the enhancement of brain-derived neurotrophic factor (BDNF) expression, and the elevation the cholinergic system, might be effective to prevent POCD. In this study, we have discovered that tacrine(10)-hupyridone (A10E), a novel acetylcholinesterase (AChE) inhibitor derived from tacrine and huperzine A, could prevent surgery-induced short-term and long-term impairments of recognition and spatial cognition, as evidenced by the novel object recognition test and Morris water maze (MWM) tests, in aged mice. Moreover, A10E significantly increased the expression of BDNF and activated the downstream Akt and extracellular regulated kinase (ERK) signaling in the surgery-treated mice. Furthermore, A10E substantially enhanced choline acetyltransferase (ChAT)-positive area and decreased AChE activity, in the hippocampus regions of surgery-treated mice, indicating that A10E could prevent surgery-induced dysfunction of cholinergic system, possibly via increasing the synthesis of acetylcholine and the inhibition of AChE. In conclusion, our results suggested that A10E might prevent POCD via the activation of BDNF pathway and the inhibition of AChE, concurrently, in aged mice. These findings also provided a support that A10E might be developed as a potential drug lead for POCD.

Keywords: acetylcholinesterase; brain-derived neurotrophic factor; choline acetyltransferase; post-operative cognitive dysfunction; tacrine(10)-hupyridone.

Figure 1

Design of animal experiments. Day 1, mice were administrated with a mixture of 0.02 mg/kg fentanyl and 5.0 mg/kg droperidol to cause anesthesia. Then, a laparotomy was conducted to induce post-operative cognitive dysfunction (POCD). Each day, tacrine(10)-hupyridone (A10E; 0.2–0.6 mg/kg) was injected intraperitoneally (i.p). for 32 days. After surgery, mice were allowed to recover for 4 days. At day 4 post-operation, the motor functions of mice were evaluated by the open field test. The effect of A10E on recognition and spatial cognition was tested by novel objective recognition (NOR) or morris water maze (MWM) testing at each of two phases on days 5–12 and days 24–31 post-operation, respectively. On the final day, animals were sacrificed.

Figure 2

Treatments did not significantly change motor functions of mice after surgery. The open field test was performed on day 4 post-operation. The number of line crossing and rearing events were calculated, and the results are shown in (A,B), respectively (n = 8).

Figure 3

A10E significantly prevented surgery-induced impairment of recognition in the early stage. The 1st NOR test was performed on days 5–7 post-operation. (A) In the training session of the 1st NOR test, treatments did not significantly change the recognition index in aged mice. (B) In the retention session of the 1st NOR test, A10E treatment at 0.4–0.6 mg/kg significantly prevented the reduction of recognition index in surgery-treated mice (n = 8). ^#p < 0.05 vs. the control group, **p < 0.01 vs. the surgery group (Dunn’s multiple comparison test).

Figure 4

A10E significantly prevented surgery-induced impairment of spatial learning and memory in the early stage. The 1st MWM test was performed on days 8–12 post operation. (A) In the training trials of the 1st MWM test, treatment with A10E at 0.6 mg/kg significantly reduced the escape latency at the last day of test in surgery-treated mice. (B) In the probe trial session of the 1st MWM test, A10E treatment at 0.6 mg/kg significantly increased the time duration spent by animals in the target quadrant in surgery-treated mice (n = 8). ^#p < 0.05 vs. the control group, *p < 0.05 vs. the surgery group (Dunn’s multiple comparison test).

Figure 5

A10E significantly prevented surgery-induced impairment of recognition in the late stage. The 2nd NOR test was performed on days 24–26 post-operation. (A) In the training session of the 2nd NOR test, treatments did not significantly change the recognition index in aged mice. (B) In the retention session of the 2nd NOR test, treatment with A10E at 0.4–0.6 mg/kg significantly prevented the reduction of recognition index in surgery-treated mice (n = 8). ^#p < 0.05 vs. the control group, *p < 0.05 vs. the surgery group (Dunn’s multiple comparison test).

Figure 6

A10E significantly prevented surgery-induced impairment of spatial learning and memory in the late stage. The 2nd MWM test was performed on days 27–31 post operation. (A) In the training trials of the 2nd MWM test, treatment with A10E at 0.6 mg/kg significantly reduced the escape latency at the last day of testing in surgery-treated mice. (B) In the probe trial session of the 2nd MWM test, A10E treatment at 0.6 mg/kg significantly increased the time duration spent of animals in the target quadrant in surgery-treated mice (n = 8). ^#p < 0.05 vs. the control group, *p < 0.05 vs. the surgery group (Dunn’s multiple comparison test).

Figure 7

A10E significantly increased the brain-derived neurotrophic factor (BDNF)-positive area in the hippocampal regions of surgery-treated mice. (A) Representative images of BDNF staining in the hippocampal region of mice in various groups, as indicated (scale bar = 150 μm). (B) Quantitative results demonstrated that treatment with A10E at 0.6 mg/kg significantly increased BDNF-positive area in surgery-treated mice (n = 4). ^##p < 0.01 vs. control group, *p < 0.05 vs. the surgery group (Dunn’s multiple comparison test).

Figure 8

A10E significantly increased the expression of pERK and pAkt in the hippocampal regions of surgery-treated mice. (A) The expression of pAkt, Akt, pERK, extracellular regulated kinase (ERK) and β-actin in the hippocampal regions of mice was detected by Western blotting. Quantitative results demonstrated that treatment with A10E at 0.6 mg/kg significantly increased the expressions of (B) pAkt and (C) pERK in surgery-treated mice (n = 4). ^#p < 0.05 vs. control group and *p < 0.05 vs. the surgery group (Dunn’s multiple comparison test).

Figure 9

A10E significantly increased the choline acetyltransferase (ChAT)-positive area in the hippocampal regions of surgery-treated mice. (A) Representative images of ChAT staining in the hippocampal region of mice in various groups, as indicated (scale bar = 150 μm). (B) Quantitative results demonstrated that treatment with A10E at 0.6 mg/kg significantly increased ChAT-positive area in surgery-treated mice (n = 4). ^#p < 0.05 vs. control group, *p < 0.05 vs. the surgery group (Dunn’s multiple comparison test).

Figure 10

A10E significantly prevented surgery-induced inhibition of acetylcholinesterase (AChE) activity ex vivo. A10E was injected 30 min before the sacrifice. AChE activity were measured ex vivo (n = 4). ^#p < 0.05 vs. the control group, *p < 0.05 vs. the surgery group (Dunn’s multiple comparison test).