Dimethyl Fumarate Alleviates Adult Neurogenesis Disruption in Hippocampus and Olfactory Bulb and Spatial Cognitive Deficits Induced by Intracerebroventricular Streptozotocin Injection in Young and Aged Rats

Abstract

The disorder of adult neurogenesis is considered an important mechanism underlying the learning and memory impairment observed in Alzheimer's disease (AD). The sporadic nonhereditary form of AD (sAD) affects over 95% of AD patients and is related to interactions between genetic and environmental factors. An intracerebroventricular injection of streptozotocin (STZ-ICV) is a representative and well-established method to induce sAD-like pathology. Dimethyl fumarate (DMF) has antioxidant and anti-inflammatory properties and is used for multiple sclerosis treatment. The present study determines whether a 26-day DMF therapy ameliorates the disruption of adult neurogenesis and BDNF-related neuroprotection in the hippocampus and olfactory bulb (OB) in an STZ-ICV rat model of sAD. Considering age as an important risk factor for developing AD, this study was performed using 3-month-old (the young group) and 22-month-old (the aged group) male Wistar rats. Spatial cognitive functions were evaluated with the Morris water maze task. Immunofluorescent labelling was used to assess the parameters of adult neurogenesis and BDNF-related neuroprotection in the hippocampus and OB. Our results showed that the STZ-ICV evoked spatial learning and memory impairment and disturbances in adult neurogenesis and BDNF expression in both examined brain structures. In the aged animals, the deficits were more severe. We found that the DMF treatment significantly alleviated STZ-ICV-induced behavioural and neuronal disorders in both age groups of the rats. Our findings suggest that DMF, due to its beneficial effect on the formation of new neurons and BDNF-related neuroprotection, may be considered as a promising new therapeutic agent in human sAD.

Keywords: adult neurogenesis; age; dimethyl fumarate; hippocampus; neuroprotection; olfactory bulb; spatial memory impairment; sporadic Alzheimer’s disease; streptozotocin.

Figure 1

Effects of STZ-ICV and DMF treatment on spatial learning (3 days of sessions, 4 trials per session, platform position fixed) evaluated by Morris water maze in young (3-month-old) and aged (22-month-old) rats. (A) Latency to reach the platform in 1–3 sessions (D1, D2, D3) in the MWM. (B) Percentage of total distance travelled in the critical quadrant (CQ) in 1–3 sessions in the MWM. Data are expressed as mean ± SEM of trials; n = 10 animals/group. Explanations: $: p < 0.05 vs. both Sham groups within the age group; &: p < 0.05 vs. only Sham+DMF within the age group; @: p < 0.05 vs. STZ within the age group; *: p < 0.05 vs. previous day within the experimental group; Y: p < 0.05 young vs. aged rats within the experimental group by three-way ANOVA followed by Tukey’s post-hoc test.

Figure 2

Effects of STZ-ICV and DMF treatment on reference memory performance in probe test evaluated by Morris water maze in young (3-month-old) and aged (22-month-old) rats. (A) Latency to reach the critical quadrant (CQ) in which the platform was located in the previous sessions of spatial learning in the MWM. (B) Percentage of the total distance travelled in the CQ. (C) Number of goal crossings indicates how often an animal crossed the former platform position. (D) Visualisation of the path shape and the number of goal (black circle) crossings of representative rat of each group. Data (A–C) are expressed as mean ± SEM; n = 10 animals/group. Explanations: $: p < 0.05 vs. both Sham groups within the age group; &: p < 0.05 vs. only Sham+DMF within the age group; @: p < 0.05 vs. STZ within the age group; Y: p < 0.05 young vs. aged rats within the experimental group by three-way ANOVA followed by Tukey’s post-hoc test.

Figure 3

Effects of STZ-ICV and DMF treatment on working memory (3 days of sessions, 4 trials per session, platform position changed daily) evaluated by Morris water maze in young (3-month-old) and aged (22-month-old) rats. (A) Latency to reach the platform in 4 trials (T1–T4) during the sessions 5 7 in the MWM. (B) Percentage of the total distance travelled in the CQ in 4 trials (T1–T4) during the sessions 5–7 in the MWM. Data are expressed as mean ± SEM of days; n = 10 animals/group. Explanations: $: p < 0.05 vs. both Sham groups within the age group; &: p < 0.05 vs. only Sham+DMF within the age group; @: p < 0.05 vs. STZ within the age group; *: p < 0.05 vs. previous trial within the experimental group; Y: p < 0.05 young vs. aged rats within the experimental group by three-way ANOVA followed by Tukey’s post-hoc test.

Figure 4

Effects of STZ-ICV and DMF treatment on neural proliferation in young (3-month-old) and aged (22-month-old) rats. Quantification of BrdU-containing cells in the dentate gyrus of the hippocampus (A) and in the olfactory bulb (B). (C) Representative photomicrographs of BrdU-labelled cells in the DG and OB of young and aged animals of all experimental groups. Scale bar = 100 μm. Data (A,B) are expressed as mean ± SEM; n = 10 animals/group. Explanations: $: p < 0.05 vs. both Sham groups within the age group; &: p < 0.05 vs. only Sham+DMF within the age group; @: p < 0.05 vs. STZ within the age group; Y: p < 0.05 young vs. aged rats within the experimental group by three-way ANOVA followed by Tukey’s post-hoc test.

Figure 5

Effects of STZ-ICV and DMF treatment on neuronal differentiation in young (3-month-old) and aged (22-month-old) rats. Quantification of DCX-containing cells in the dentate gyrus of the hippocampus (A) and in the olfactory bulb (B). (C) Representative photomicrographs of DCX-labelled cells in the DG and OB of young and aged animals of all experimental groups. Scale bar = 100 μm. Data (A,B) are expressed as mean ± SEM; n = 10 animals/group. Explanations: $: p < 0.05 vs. both Sham groups within the age group; &: p < 0.05 vs. only Sham+DMF within the age group; @: p < 0.05 vs. STZ within the age group; Y: p < 0.05 young vs. aged rats within the experimental group by three-way ANOVA followed by Tukey’s post-hoc test.

Figure 6

Effects of STZ-ICV and DMF treatment on adult neurogenesis level in young (3-month-old) and aged (22-month-old) rats. Percentage of double-labelled cells (BrdU+DCX-containing cells—newly formed immature neurons) of the sum of all fluorescent cells in the dentate gyrus of the hippocampus (A) and in the olfactory bulb (B). (C) Representative photomicrographs of BrdU+DCX-labelled cells in DG and OB of young and aged animals of all experimental groups. Double labelled (BrdU+DCX) cells are indicated by white arrows. Scale bar = 100 μm. Data (A,B) are expressed as mean ± SEM; n = 10 animals/group. Explanations: $: p < 0.05 vs. both Sham groups within the age group; &: p < 0.05 vs. only Sham+DMF within the age group; @: p < 0.05 vs. STZ within the age group; Y: p < 0.05 young vs. aged rats within the experimental group by three-way ANOVA followed by Tukey’s post-hoc test.

Figure 7

Effects of STZ-ICV and DMF treatment on neuroprotection in young (3-month-old) and aged (22-month-old) rats. (A) Quantification of BDNF-containing cells in both neurogenic regions: the dentate gyrus of the hippocampus and olfactory bulb, and in CA1–CA3 areas of the hippocampus. Data are expressed as mean ± SEM; young n = 10 animals/group, aged n = 8–10 animals/group. (B) Representative photomicrographs of BDNF-labelled cells in the DG of young animals in all experimental groups and in aged STZ and STZ+DMF rats. Scale bar = 100 μm. Data (A) are expressed as mean ± SEM; n = 10 animals/group. Explanations: $: p < 0.05 vs. both Sham groups within the age group; &: p < 0.05 vs. only Sham+DMF within the age group; @: p < 0.05 vs. STZ within the age group; Y: p < 0.05 young vs. aged rats within the experimental group by three-way ANOVA followed by Tukey’s post-hoc test.

Figure 8

Diagram of experimental procedures and group assignments. Explanations: f.ch.—change in feed; ICV1/ICV2—intracerebroventricular injection (1—the first injection, 2—the second injection) of STZ (a total dose of 3 mg/kg divided into 2 injections to both lateral ventricles: 0.75 mg/kg STZ in 2 µL of citrate buffer per ventricle) or vehicle (citrate buffer 2 µL per ventricle); BrdU ip—intraperitoneal injection of 5-bromo-2’deoxyuridine (BrDU) at a dose of 50 mg/kg for three consecutive days; MWM—Morris water maze test; SL—spatial learning, the phase of acquisition of reference memory; probe—single trial without the platform (reference memory evaluation); WOR—working memory performance.