Vitisin A, a Resveratrol Tetramer, Improves Scopolamine-Induced Impaired Learning and Memory Functions in Amnesiac ICR Mice

Abstract

Resveratrol has been reported to exhibit neuroprotective activities in vitro and in vivo. However, little is known about resveratrol tetramers of hopeaphenol, vitisin A, and vitisin B with the same molecular mass in the improvement of degenerative disorders. In this study, two 95% ethanol extracts (95EE) from stem parts of Vitis thunbergii Sieb. & Zucc. (VT-95EE) and from the root (R) parts of Vitis thunbergii var. taiwaniana (VTT-R-95EE) showed comparable acetylcholinesterase (AChE) inhibitory activities. It was found that VT-95EE and VTT-R-95EE showed different distribution patterns of identified resveratrol and resveratrol tetramers of hopeaphenol, vitisin A, and vitisin B based on the analyses of HPLC chromatographic profiles. The hopeaphenol, vitisin A, and vitisin B, showed AChE and monoamine oxidase-B inhibitions in a dose-dependent manner, among which vitisin B and vitisin A exhibited much better activities than those of resveratrol, and had neuroprotective activities against methylglyoxal-induced SH-SY5Y cell deaths. The scopolamine-induced amnesiac ICR mice treated with VT-95EE and its ethyl acetate-partitioned fraction (VT-95EE-EA) at doses of 200 and 400 mg/kg, or vitisin A at a dose of 40 mg/kg, but not vitisin B (40 mg/kg), were shown significantly to improve the impaired learning behaviors by passive avoidance tests compared to those in the control without drug treatments (p < 0.05). Compared to mice in the control group, the brain extracts in the vitisin A-treated mice or donepezil-treated mice showed significant reductions in AChE activities and malondialdehyde levels (p < 0.05), and elevated the reduced protein expressions of brain-derived neurotrophic factor (BDNF) and BDNF receptor, tropomyosin receptor kinase B (TrkB). These results revealed that vitisin A was the active constituent in the VT-95EE and VTT-95EE, and the VT medicinal plant and that the endemic variety of VTT has potential in developing functional foods for an unmet medical need for neurodegenerative disorders.

Keywords: acetylcholinesterase; brain-derived neurotrophic factor (BDNF); scopolamine; vitisin A.

Figure 1

Effects of (A) VT-HWE, VT-95EE, VT-95EE-EA, VT-95EE-BuOH, and VT-95EE-H₂O; and (B) HWE and 95EE of VTT-S, VTT-L, and VTT-R, and VTT-R-95EE-EA, VTT-R-95EE-BuOH, and VTT-R-95EE-H₂O on acetylcholinesterase (AChE) inhibitory activities. The stem part of Vitis thunbergii Sieb. & Zucc. (VT) or the Vitis thunbergii var. taiwaniana (VTT) of different parts, root (VTT-R), stem (VTT-S), and leaf (VTT-L), each was extracted either by hot-water (HW) or 95% ethanol (95E) to get HW extracts (HWE) and 95EE. Data were expressed as mean ± SD of three independent quantitative experiments.

Figure 2

(A) The structures of the resveratrol and resveratrol oligomers, including resveratrol dimer of (+)-ε-viniferin, resveratrol trimer of ampelopsin C, and resveratrol tetramers of (+)-vitisin A, (+)-hopeaphenol, and (−)-vitisin B. The HPLC chromatograms of (B) VT-95EE; (C) VT-95EE-EA; (D) VTT-R-95EE; and (E) VTT-R-95EE-EA. The identified compounds included resveratrol (peak 1), (+)-hopeaphenol (resveratrol tetramer, peak 2), (+)-ampelopsin C (resveratrol trimer, peak 3), (+)-ε-viniferin (resveratrol dimer, peak 4), (+)-vitisin A (resveratrol tetramer, peak 5), and (−)-vitisin B (resveratrol tetramer, peak 6) based on the elution sequence.

Figure 2

(A) The structures of the resveratrol and resveratrol oligomers, including resveratrol dimer of (+)-ε-viniferin, resveratrol trimer of ampelopsin C, and resveratrol tetramers of (+)-vitisin A, (+)-hopeaphenol, and (−)-vitisin B. The HPLC chromatograms of (B) VT-95EE; (C) VT-95EE-EA; (D) VTT-R-95EE; and (E) VTT-R-95EE-EA. The identified compounds included resveratrol (peak 1), (+)-hopeaphenol (resveratrol tetramer, peak 2), (+)-ampelopsin C (resveratrol trimer, peak 3), (+)-ε-viniferin (resveratrol dimer, peak 4), (+)-vitisin A (resveratrol tetramer, peak 5), and (−)-vitisin B (resveratrol tetramer, peak 6) based on the elution sequence.

Figure 3

(A) Effects of the same molecular mass of resveratrol tetramers, vitisin A, hopeaphenol, and vitisin B, on acetylcholinesterase (AChE) inhibitory activities in comparisons with resveratrol, and the donepezil was used as the positive control; (B) Effects of the same molecular mass of resveratrol tetramers, vitisin A, hopeaphenol, and vitisin B, on monoamine oxidase (MAO)-B inhibitory activities in comparisons with resveratrol, and the deprenyl was used as the positive control; (C) Effects of resveratrol, vitisin A, hopeaphenol, and vitisin B, on neuroprotective activities (2.5, 5, 10, and 20 μM) in methylglyoxal-induced SH-SY5Y cell deaths. Data were expressed as mean ± SD of three independent quantitative experiments. The Student’s t-test was used to compare the neuroprotective activity of resveratrol or resveratrol tetramers in 500 μM methylglyoxal-treated SH-SY5Y cells (C) with the control. The different cell viability was considered as the significant differences when p < 0.001 (***).

Figure 4

Effects of VT-95EE or VT-95EE-EA pretreatments (200 and 400 mg/kg) on the improvements of impaired learning and memory functions in scopolamine-treated amnesiac ICR mice. (A) The experimental protocol. There were seven groups (six heads of ICR mice/group), including the blank, the control, the VT-95EE (200, 400 mg/kg) groups, the VT-95EE-EA (200, 400 mg/kg) groups, and donepezil group (the positive control); (B) the learning and memory behaviors (the step-through latency, s) in the passive avoidance test. The scopolamine-injected mice without any sample treatments were used as the control. The one-way analysis of variance (ANOVA) and the post hoc Tukey’s test were used to compare the differences among multiple groups in the step-through latency (B). It was considered as the significant difference (p < 0.05) among groups which marked the different uppercase letters in the acquisition trial or marked the different lowercase letters in the retention trial.

Figure 4

Effects of VT-95EE or VT-95EE-EA pretreatments (200 and 400 mg/kg) on the improvements of impaired learning and memory functions in scopolamine-treated amnesiac ICR mice. (A) The experimental protocol. There were seven groups (six heads of ICR mice/group), including the blank, the control, the VT-95EE (200, 400 mg/kg) groups, the VT-95EE-EA (200, 400 mg/kg) groups, and donepezil group (the positive control); (B) the learning and memory behaviors (the step-through latency, s) in the passive avoidance test. The scopolamine-injected mice without any sample treatments were used as the control. The one-way analysis of variance (ANOVA) and the post hoc Tukey’s test were used to compare the differences among multiple groups in the step-through latency (B). It was considered as the significant difference (p < 0.05) among groups which marked the different uppercase letters in the acquisition trial or marked the different lowercase letters in the retention trial.

Figure 5

Effects of vitisin A or vitisin B pretreatments (40 mg/kg) on the improvements impaired learning and memory functions in scopolamine-treated amnesiac ICR mice. (A) The experimental protocol. There were five groups and each group contained 6 heads of ICR mice, including the blank, the control, the vitisin A (40 mg/kg) group, the vitisin B (40 mg/kg) group, and the donepezil group (the positive control); (B) the learning and memory behaviors (the step-through latency, sec) in the passive avoidance test. The scopolamine-injected mice without any sample treatments were used as the control. The one-way analysis of variance (ANOVA) and the post hoc Tukey’s test were used to compare the differences among multiple groups in the step-through latency (B). It was considered as the significant difference (p < 0.05) among groups which the marked different uppercase letters in the acquisition trial or the marked different lowercase letters in the retention trial.

Figure 6

The changes of (A) AChE activity, (B) the MDA levels, and (C) BDNF and TrkB protein expressions, and expressed as ratios of BDNF/β-actin and TrkB/β-actin in the brain tissue extracts of amnesiac mice after vitisin A or vitisin B treatments (40 mg/kg). The scopolamine-injected mice without any sample treatments were used as the control. Data were expressed as mean ± SD of three independent quantitative experiments. The one-way analysis of variance (ANOVA) and the post hoc Tukey’s test were used to compare the differences among multiple groups. It was considered as the significant difference (p < 0.05) among groups which marked the different lowercase letters.