A Novel Ashwagandha (Withania somnifera) Formulation Mitigates Sleep Deprivation-Induced Cognitive Impairment and Oxidative Stress in a Rat Model

Abstract

Ashwagandha (Withania somnifera) is a well-known adaptogenic herb traditionally used to enhance sleep quality and mitigate stress-induced cognitive decline. This study investigated the effects of different doses of ashwagandha root extract (AE) formulations on cognitive function, oxidative stress, and neuronal plasticity in a rat model of sleep deprivation (SD). Forty-nine rats were randomly assigned to seven groups: control, wide platform (WP), SD, SD + A1 (15 mg/kg AE 1.5%), SD + A2 (30 mg/kg AE 1.5%), SD + A3 (5.5 mg/kg AE 8.0%), and SD + A4 (11 mg/kg AE 8.0%). The extract was administered orally for four weeks. SD induced via a modified wide platform model significantly impaired spatial memory, increased oxidative stress, and suppressed GABA receptor activity. Treatment with all AE doses, except 15 mg/kg AE 1.5%, considerably reduced serum corticosterone (12% for SD + A2, 15% for SD + A3, and 32% for SD + A4), CRH (11% for SD + A2, 14% for SD + A3, and 17% for SD + A4), ACTH (22% for SD + A2, 26% for SD + A3, and 38% for SD + A4), and MDA levels (31% for SD + A2, 34% for SD + A3, and 46% for SD + A4) (p < 0.05). All doses improved antioxidant enzyme activity and memory performance, while AE 8.0% doses notably increased serotonin (19% for SD + A3 and 33% for SD + A4) and dopamine levels (40% for SD + A3 and 50% for SD + A4). Moreover, AE treatment enhanced markers of neuronal plasticity and partially improved GABAergic function. These findings suggest that AE formulations, particularly at higher concentrations, exert neuroprotective effects against SD-induced cognitive impairment by modulating oxidative stress, neurotransmitter balance, and neuroplasticity, indicating their potential application in managing stress-related neurological disorders.

Keywords: GABAergic pathway; ashwagandha; memory; sleep deprivation; stress; withanolides.

Figure 1

Experimental procedures.

Figure 2

Sleep deprivation (SD) platform images for rats.

Figure 3

Effects of different ashwagandha doses on corticosterone (A), CRH (B), ACTH (C), serotonin (D), and dopamine (E) in sleep-deprived rats. Data were analyzed using one-way ANOVA followed by Tukey’s post hoc test for multiple comparisons. Different superscript letters (a–e) indicate statistically significant differences between groups (p < 0.05). The groups not sharing the same letter are significantly different. The error lines indicate SEM, and symbols show individual values. CRH: corticotropin-releasing hormone, ACTH: adrenocorticotropic hormone, C: control, WP: wide platform, SD: sleep deprivation, SD + A1: SD + ashwagandha 1.5% (15 mg/kg), SD + A2: SD + ashwagandha 1.5% (30 mg/kg), SD + A3: SD + ashwagandha 8% (5.5 mg/kg), SD + A4: SD + ashwagandha 8% (11 mg/kg).

Figure 4

Effects of different ashwagandha doses on oxidative stress markers in sleep-deprived rats. (A) serum MDA, (B) liver MDA, (C) brain MDA, (D) brain superoxide dismutase (SOD), (E) brain catalase (CAT), (F) brain glutathione peroxidase (GSH-Px), and (G) total antioxidant capacity (TAC). Data were analyzed using one-way ANOVA followed by Tukey’s post hoc test for multiple comparisons. Different superscript letters (a–e) indicate statistically significant differences between groups (p < 0.05). The groups not sharing the same letter are significantly different. The error lines indicate SEM, and symbols show individual values. MDA: malondialdehyde, SOD: superoxide dismutase, CAT: catalase, GSH-Px: glutathione peroxidase, TAC: total antioxidant capacity. C: control, WP: wide platform, SD: sleep deprivation, SD + A1: SD + ashwagandha 1.5% (15 mg/kg), SD + A2: SD + ashwagandha 1.5% (30 mg/kg), SD + A3: SD + ashwagandha 8% (5.5 mg/kg), SD + A4: SD + ashwagandha 8% (11 mg/kg).

Figure 5

Effects of different ashwagandha doses on entries to target quadrant (A), probe trial (B), and spatial learning memory acquisition phase (SMAP) (C) in sleep-deprived rats. One-way ANOVA was used for group differences, and Tukey post hoc analysis for multiple comparisons. Kruskal–Wallis and Mann–Whitney U tests were used to compare the probe trials. Different superscripts (a–d) indicate the mean differences between the groups (p < 0.05). The groups not sharing the same letter are significantly different. The error lines indicate SEM for entries to the target quadrant and 95% confidence interval for the probe trial. Symbols show individual values. C: control, WP: wide platform, SD: sleep deprivation, SD + A1: SD + ashwagandha 1.5% (15 mg/kg), SD + A2: SD + ashwagandha 1.5% (30 mg/kg), SD + A3: SD + ashwagandha 8% (5.5 mg/kg), SD + A4: SD + ashwagandha 8% (11 mg/kg).

Figure 6

Effects of different ashwagandha doses on brain tissue NCAM (A), ICAM-1 (B), BDNF (C), NGF (D), and GAP-43 (E) protein levels and representative Western blot bands ((F) and Figure S1) in sleep-deprived rats. Western blot analysis was performed with incorporated β-actin to ensure equal protein loading. Representative bands are shown in panel (F). Data were analyzed using one-way ANOVA followed by Tukey’s post hoc test for multiple comparisons. Different superscript letters (a–g) indicate statistically significant differences between groups (p < 0.05). The groups not sharing the same letter are significantly different. The error lines indicate SEM. NCAM: neural cell adhesion molecule, ICAM: intercellular adhesion molecule-1, BDNF: brain-derived neurotrophic factor, NGF: nerve growth factor. C: control, WP: wide platform, SD: sleep deprivation, SD + A1: SD + ashwagandha 1.5% (15 mg/kg), SD + A2: SD + ashwagandha 1.5% (30 mg/kg), SD + A3: SD + ashwagandha 8% (5.5 mg/kg), SD + A4: SD + ashwagandha 8% (11 mg/kg). The original Western blot image can be found in the Supplementary Materials.

Figure 7

Effects of different ashwagandha doses on brain tissue GABA_AR2 (A), GABA_BR1 (B), GABA_BR2 (C), and 5-HT1A (D) protein levels and representative Western blot bands ((E) and Figure S2) in sleep-deprived rats. Western blot analysis was performed with incorporated β-actin to ensure equal protein loading. Representative bands are shown in panel E. Data were analyzed using one-way ANOVA followed by Tukey’s post hoc test for multiple comparisons. Different superscript letters (a–f) indicate statistically significant differences between groups (p < 0.05). The groups not sharing the same letter are significantly different. The error lines indicate SEM. GABA_A: gamma-aminobutyric acid type A receptor subunit alpha, GAP-43: growth-associated protein-43. C: control, WP: wide platform, SD: sleep deprivation, SD + A1: SD + ashwagandha 1.5% (15 mg/kg), SD + A2: SD + ashwagandha 1.5% (30 mg/kg), SD + A3: SD + ashwagandha 8% (5.5 mg/kg), SD + A4: SD + ashwagandha 8% (11 mg/kg). The original Western blot image can be found in the Supplementary Materials.