Sobrerol Improves Memory Impairment in the Scopolamine-Induced Amnesia Mouse Model

Abstract

Memory impairment is a defining characteristic of Alzheimer's disease (AD), with amnesia often appearing as its earliest symptom. Given the multifactorial nature of AD pathogenesis, this study investigates the multi-target therapeutic potential of sobrerol (coded as NRM-331) in a scopolamine-induced amnesia mouse model, focusing specifically on its effects in ameliorating memory deficits and enhancing neuronal plasticity. Sixty male C57BL/6NCrljOri mice were divided into six groups (10 mice/group): vehicle control (CTL, saline), scopolamine (SPA, 10 mg/kg/day), Aricept (APT, 2 mg/kg/day), and three treatment groups receiving NRM-331 at doses of 40, 80, and 100 mg/kg/day. Several behavioral tests were conducted, including the Y-maze test, passive avoidance test, and Morris water maze test. Additionally, biochemical assays were performed in serum (to measure Aß 1-40 and Aß 1-42) and in the brain (to assess ACh and AChE levels), along with histopathological examination of the brain using Nissl staining and p-tau IHC. No significant change was observed in the Y-maze test or the acquisition trial of the passive avoidance test. However, improvements were noted in the retention trial of the passive avoidance test and the Morris water maze test (including escape latency, swim distance, and number of platform crossed) for the NRM-331 groups compared to the SPA group. Serum levels of Aß 1-40 and Aß 1-42 decreased in the NRM-331 groups compared to the SPA group. In the brain, levels of ACh significantly increased, while AChE levels significantly decreased compared to the SPA group. The number of neuronal cells improved in the CA1, CA3, and DG regions of the hippocampus, as indicated by Nissl staining. A significant reduction in p-tau accumulation was also observed in the NRM-331 groups. In conclusion, NRM-331 demonstrated an anti-amnesic effect by enhancing hippocampal cholinergic signaling, alongside exhibiting anti-tau and anti-Aβ synthesis properties. These therapeutic effects suggest that NRM-331 significantly mitigates memory impairment induced by SPA through a neuroprotective mechanism.

Keywords: acetylcholine; amnesia; amyloid beta; memory; phospho-tau; scopolamine.

Figure 1

Body weights and gained body weight during the study. (A) Weekly body weight. (B) Gained body weight. Data were represented by mean ± S.E. The results were statistically analyzed by Welch’s t-test and ONE-WAY ANOVA. The SPA group was compared to the CTL group, with significance indicated by *; the remaining groups were compared to the SPA group, with significance indicated by #. *** p < 0.001, # p < 0.05, ns: not significant. CTL (saline vehicle control); SPA (2 mg/kg/day scopolamine, negative control); ACT (2 mg/kg/day Aricept, positive control); N40 (40 mg/kg/day NRM-331); N80 (80 mg/kg/day NRM-331); N100 (100 mg/kg/day NRM-331).

Figure 2

Behavioral tests of the Y maze, passive avoidance, and Morris water maze test. (A) Spontaneous alterations in the Y-maze test, (B) escape latencies in the passive avoidance test, (C) escape latencies in the Morris water maze test, (D) swim time in the Morris water maze test, (E) swim distances in the Morris water maze test, (F) number of platform crossing in the Morris water maze test. Data were represented by mean ± S.E. The results were statistically analyzed by ONE-WAY and TWO-WAY ANOVA. SPA group was compared to the CTL group, with significance indicated by *; the remaining groups were compared to SPA, with significance indicated by #. *** or ### p < 0.001, # p < 0.05, ns: not significant. CTL (saline vehicle control); SPA (2 mg/kg/day scopolamine, negative control); ACT (2 mg/kg/day Aricept, positive control); N40 (40 mg/kg/day NRM-331); N80 (80 mg/kg/day NRM-331); N100 (100 mg/kg/day NRM-331).

Figure 3

Biochemical analysis in serum for Aβ 1-40 and Aβ 1-42 levels and in brain tissue for ACh and AChE levels by ELISA. (A) Serum amyloid-β 1-40 level, (B) Serum amyloid- β 1-42 level, (C) Brain tissue ACh level, and (D) Brain tissue AChE level. The results were statistically analyzed by ONE-WAY ANOVA. The SPA group was compared to the CTL group, with significance indicated by *; the remaining groups were compared to SPA, with significance indicated by #. *** or ### p < 0.001, ns: not significant. CTL (saline vehicle control); SPA (2 mg/kg/day scopolamine, negative control); ACT (2 mg/kg/day Aricept, positive control); N40 (40 mg/kg/day NRM-331); N80 (80 mg/kg/day NRM-331); N100 (100 mg/kg/day NRM-331); Aβ: Amyloid beta; ACh: Acetylcholine; AChE: Acetylcholine esterase.

Figure 4

Histopathology of the brain for the Nissl stain and tau-accumulation by ELISA. (A) Representative images of Nissl staining neuronal cells in the hippocampus, (B) Quantification of Nissl stain, (C) Representative images of immunohistochemical staining for p-tau, and (D) Quantification of p-tau. The results were statistically analyzed by ONE-WAY and TWO-WAY ANOVA. SPA group was compared with the CTL group, and the remaining groups were compared with the SPA group. *** or ^###: p < 0.001, ** or ^##: p < 0.01, * or ^#: p < 0.05, ns: not significant, Arrow: shrunken neuron or degenerated neuronal cell, or p-tau signal intensity. Scale bars: 500 μm or 50 μm. CTL (saline vehicle control); SPA (2 mg/kg/day scopolamine, negative control); ACT (2 mg/kg/day Aricept, positive control); N40 (40 mg/kg/day NRM-331); N80 (80 mg/kg/day NRM-331); N100 (100 mg/kg/day NRM-331).