Black seed oil reverses chronic antibiotic-mediated depression and social behaviour deficits via modulation of hypothalamic mitochondrial-dependent markers and insulin expression

Abstract

Chronic antibiotic use has been reported to impair mitochondrial indices, hypothalamus-mediated metabolic function, and amygdala-regulated emotional processes. Natural substances such as black seed (Nigella sativa) oil could be beneficial in mitigating these impairments. This study aimed to assess the impact of black seed oil (NSO) on depression and sociability indices, redox imbalance, mitochondrial-dependent markers, and insulin expression in mice subjected to chronic ampicillin exposure. Forty adult male BALB/c mice (30 ± 2 g) were divided into five groups: the CTRL group received normal saline, the ABT group received ampicillin, the NSO group received black seed oil, the ABT/NSO group concurrently received ampicillin and black seed oil, and the ABT+NSO group experienced pre-exposure to ampicillin followed by subsequent treatment with black seed oil. The ampicillin-exposed group exhibited depressive-like behaviours, impaired social interactive behaviours, and disruptions in mitochondrial-dependent markers in plasma and hypothalamic tissues, accompanied by an imbalance in antioxidant levels. Moreover, chronic antibiotic exposure downregulated insulin expression in the hypothalamus. However, these impairments were significantly ameliorated in the ABT/NSO, and ABT+NSO groups compared to the untreated antibiotic-exposed group. Overall, findings from this study suggest the beneficial role of NSO as an adjuvant therapy in preventing and abrogating mood behavioural and neural-metabolic impairments of chronic antibiotic exposure.

Keywords: Antibiotic misuse; Black seed oil; Depression; Insulin expression; Mitochondrial function; Social interaction.

Graphical abstract

Fig. 1

Experimental design.

Fig. 2

Effects of Nigella sativa oil on depressive-like behaviours following antibiotic exposure in mice from the TST (A) and FST (B) tests. Data are expressed as ± SD; n = 5 and analysed by one-way ANOVA followed by Tukey posthoc test. Legend: CTRL = Control; ABT = Antibiotics toxicity; NSO = Nigella sativa oil, ABT/NSO = Antibiotic toxicity and Nigella sativa oil simultaneously; ABT+NSO = Antibiotics toxicity with Nigella sativa oil post-treatment.

Fig. 3

Evaluation of social interaction behaviour (sociability index from the three-chamber sociability test) following Nigella sativa administration and/or chronic antibiotic exposure. Data are expressed as ± SD; n = 5 and analysed by one-way ANOVA followed by Tukey posthoc test. Legend: CTRL = Control; ABT = Antibiotics toxicity; NSO = Nigella sativa oil, ABT/NSO = Antibiotic toxicity and Nigella sativa oil simultaneously; ABT+NSO = Antibiotics toxicity with Nigella sativa oil post-treatment.

Fig. 4

Effects of Nigella sativa oil on oxidative following antibiotic exposure in mice from the MDA (A) and SOD (B) estimation. Data are expressed as ± SD; n = 4 and analysed by one-way ANOVA followed by Tukey posthoc test. Legend: CTRL = Control; ABT = Antibiotics toxicity; NSO = Nigella sativa oil, ABT/NSO = Antibiotic toxicity and Nigella sativa oil simultaneously; ABT+NSO = Antibiotics toxicity with Nigella sativa oil post-treatment.

Fig. 5

Effects of Nigella sativa oil on mitochondrial activity following antibiotic exposure in mice from the plasma lactate (A), hypothalamic lactate (B), and hypothalamic CK (C) evaluation. Data are expressed as ± SD; n = 4 and analysed by one-way ANOVA followed by Tukey posthoc test. Legend: CTRL = Control; ABT = Antibiotics toxicity; NSO = Nigella sativa oil, ABT/NSO = Antibiotic toxicity and Nigella sativa oil simultaneously; ABT+NSO = Antibiotics toxicity with Nigella sativa oil post-treatment.

Fig. 6

Effects of Nigella sativa oil on hypothalamic acetylcholinesterase activity following antibiotic exposure in mice. Data are expressed as ± SD; n = 4 and analysed by one-way ANOVA followed by Tukey posthoc test. Legend: CTRL = Control; ABT = Antibiotics toxicity; NSO = Nigella sativa oil, ABT/NSO = Antibiotic toxicity and Nigella sativa oil simultaneously; ABT+NSO = Antibiotics toxicity with Nigella sativa oil post-treatment.

Fig. 7

Photomicrograph of amygdala in mice following Nigella sativa administration and/or chronic antibiotic exposure. Group A = control (CTRL); Group B = Antibiotic toxicity (ABT); Group C = Nigella sativa oil (NSO); Group D = Antibiotic toxicity and Nigella sativa oil simultaneously (ABT/NSO); Group E = Antibiotics toxicity with Nigella sativa oil post treatment (ABT+NSO). Yellow arrow (vacuolation of the neuropil), blue arrow (viable cell), white arrow (degenerated cell) red arrow (pyknotic cell), F. Amygdala cell counts X800 (H&E).

Fig. 8

Photomicrograph of hypothalamus in mice following Nigella sativa administration and/or chronic antibiotic exposure. Group A = control (CTRL) – with numerous viable cells; Group B = Antibiotic toxicity (ABT) – with reduction in number of cells in the arcuate nucleus region of the hypothalamus; Group C = Nigella sativa oil (NSO) – numerous viable cells; Group D = Antibiotic toxicity and Nigella sativa oil simultaneously (ABT/NSO) – presenting viable cells more than ABT group; Group E = Antibiotics toxicity with Nigella sativa oil post treatment (ABT+NSO) – with numerous viable cells compared to ABT group, Yellow arrow (vacuolation of the neuropil), blue arrow (viable cell), white arrow (degenerated cell) red arrow (pyknotic cell), F. Hypothalamic cell counts X800 (H&E). X800 (H&E).

Fig. 9

Photomicrograph of amygdala in mice following Nigella sativa administration and/or chronic antibiotic exposure. Group A = control (CTRL); Group B = Antibiotic toxicity (ABT); Group C = Nigella sativa oil (NSO); Group D = Antibiotic toxicity and Nigella sativa oil simultaneously (ABT/NSO); Group E = Antibiotics toxicity with Nigella sativa oil post treatment (ABT+NSO). Yellow arrow (vacuolated neuropil), white arrow (degenerated cell) red arrow (pyknotic cell), F. Amygdala Nissl substance area. X800 (CFV).

Fig. 10

Photomicrograph of hypothalamus in mice following Nigella sativa administration and/or chronic antibiotic exposure. Group A = control (CTRL) – with numerous viable cells; Group B = Antibiotic toxicity (ABT) – with reduction in number of cells in the arcuate nucleus region of the hypothalamus; Group C = Nigella sativa oil (NSO) – numerous viable cells; Group D = Antibiotic toxicity and Nigella sativa oil simultaneously (ABT/NSO) – presenting viable cells more than ABT group; Group E = Antibiotics toxicity with Nigella sativa oil post treatment (ABT+NSO) – with numerous viable cells compared to ABT group. Yellow arrow (vacuolated neuropil), white arrow (degenerated cell) red arrow (pyknotic cell), F. Hypothalamic Nissl substance area. X800 (CFV).

Fig. 11

Photomicrograph of amygdala in mice following Nigella sativa administration and/or chronic antibiotic exposure. Group A = control (CTRL); Group B = Antibiotic toxicity (ABT); Group C = Nigella sativa oil (NSO); Group D = Antibiotic toxicity and Nigella sativa oil simultaneously (ABT/NSO); Group E = Antibiotics toxicity with Nigella sativa oil post treatment (ABT+NSO). Yellow arrow (insulin expression), white arrow (degenerated neuron) red arrow (pyknotic nucleus) F. Amygdala INS percent area occupied G. Amygdala INS staining intensity. X400 (Immunohistochemical staining techniques).

Fig. 12

Photomicrograph of hypothalamus insulin expression in mice following Nigella sativa administration and/or chronic antibiotic exposure. Group A = control (CTRL); Group B = Antibiotic toxicity (ABT); Group C = Nigella sativa oil (NSO); Group D = Antibiotic toxicity and Nigella sativa oil simultaneously (ABT/NSO); Group E = Antibiotics toxicity with Nigella sativa oil post treatment (ABT+NSO); 3 V = third ventricle; ARC = arcuate nucleus. Yellow arrow = (insulin expression); red arrow = (degenerated neuron). F. Hypothalamic INS percent area occupied G. Hypothalamic INS staining intensity. X800 Immunohistochemical staining techniques.