Curcumin treatment leads to better cognitive and mood function in a model of Gulf War Illness with enhanced neurogenesis, and alleviation of inflammation and mitochondrial dysfunction in the hippocampus

Abstract

Diminished cognitive and mood function are among the most conspicuous symptoms of Gulf War Illness (GWI). Our previous studies in a rat model of GWI have demonstrated that persistent cognitive and mood impairments are associated with substantially declined neurogenesis, chronic low-grade inflammation, increased oxidative stress and mitochondrial dysfunction in the hippocampus. We tested the efficacy of curcumin (CUR) to maintain better cognitive and mood function in a rat model of GWI because of its neurogenic, antiinflammatory, antioxidant, and memory and mood enhancing properties. Male rats were exposed daily to low doses of GWI-related chemicals, pyridostigmine bromide, N,N-diethyl-m-toluamide (DEET) and permethrin, and 5-minutes of restraint stress for 28 days. Animals were next randomly assigned to two groups, which received daily CUR or vehicle treatment for 30 days. Animals also received 5'-bromodeoxyuridine during the last seven days of treatment for analysis of neurogenesis. Behavioral studies through object location, novel object recognition and novelty suppressed feeding tests performed sixty days after treatment revealed better cognitive and mood function in CUR treated GWI rats. These rats also displayed enhanced neurogenesis and diminished inflammation typified by reduced astrocyte hypertrophy and activated microglia in the hippocampus. Additional studies showed that CUR treatment to GWI rats enhanced the expression of antioxidant genes and normalized the expression of multiple genes related to mitochondrial respiration. Thus, CUR therapy is efficacious for maintaining better memory and mood function in a model of GWI. Enhanced neurogenesis, restrained inflammation and oxidative stress with normalized mitochondrial respiration may underlie better memory and mood function mediated by CUR treatment.

Keywords: Curcumin; Gulf War Illness; Hippocampal neurogenesis; Memory impairment; Mitochondria; Mood dysfunction; Neuroinflammation; Oxidative stress; Reactive oxygen species.

Fig. 1.

Timeline of various studies. Nine-week old Sprague-Dawley rats (n = 32) were exposed daily to Gulf War Illness related chemicals (GWIR-C’s) and 5-minutes of restraint stress for 28 days. A day later, rats exposed to GWIR-Cs and stress were randomly assigned to two groups. One group received daily curcumin (CUR, 30 mg/Kg i.p., n = 16) and the other group received vehicle (VEH, n = 16) for 30 days. Sixty days later, animals were subjected to a series of behavioral tests to assess cognitive and mood function. A group of age-matched naïve control rats (n = 11) were also included in behavioral studies. Animals in all groups were perfused with 4% paraformaldehyde following the conclusion of behavioral tests, and hippocampal tissues were processed for analyses of neurogenesis and inflammation.

Fig. 2.

Curcumin (CUR) treated Gulf War Illness (GWI) rats displayed better ability to discern minor changes in the environment. Fig. A1 shows the various phases involved in an object location test (OLT). The bar charts in A2–A4 illustrate the performance of animals belonging to naïve control (naïve CTRL, A2), GWI animals receiving vehicle (GWI + VEH, A3), and GWI animals receiving CUR (GWI + CUR, A4) groups. Animals in GWI + VEH group showed impairment. This was evidenced by the lack of preference for the novel place object (NPO) in OLT over the familiar place object (FPO; A3). In contrast, animals in naïve control and GWI + CUR groups showed better cognitive function. This was apparent from their preference for NPO in OLT (A4). The bar charts in A5–A7 demonstrate that cognitive test results were not influenced by variability in object exploration time (A5), the total distance moved (A6) or the velocity of movement (A7) between groups. *p < 0.05, **p < 0.01, and ***p < 0.001.

Fig. 3.

Curcumin (CUR) treated Gulf War Illness (GWI) rats exhibited better novel object recognition memory and mood function. The Fig. A1 shows the various phases involved in a novel object recognition test (NORT). The bar charts in A2–A4 illustrate the performance of animals belonging to naïve control (naïve CTRL, A2), GWI animals receiving vehicle (GWI + VEH, A3), and GWI animals receiving CUR (GWI + CUR, A4) groups. Animals in GWI + VEH group showed novel object recognition memory impairment. This was evidenced by the lack of preference for the novel object (NO) in NORT (A3). In contrast, animals in naïve control and GWI + CUR groups showed normal novel object recognition memory. This was apparent from their preference for NO in NORT (A4). The bar charts in A5–A7 demonstrate that cognitive test results were not influenced by variability in object exploration time (A5), the total distance moved (A6) or the velocity of movement (A7) between groups. The Fig. B1 shows the test apparatus used for a novelty suppressed feeding test (NSFT). The bar chart in B2 compares latency values to the first bite of food between different groups. Note that, animals in GWI + VEH group showed longer latencies than naïve control and GWI + CUR groups, implying a decreased motivation and an increased anxiety-like behavior in this group. In contrast, animals in GWI + CUR group displayed similar latencies as naïve control animals. *p < 0.05, **p < 0.01, and ***p < 0.001.

Fig. 4.

Curcumin (CUR) treatment enhanced neurogenesis in animals with Gulf War Illness (GWI). A1–B3 show distribution of BrdU+ cells in naïve control animals (A1, B1), GWI animals receiving vehicle (GWI-VEH; A2, B2), and GWI animals receiving CUR (GWI-CUR; A3, B3). The bar chart in I compares the number of BrdU+ cells in the subgranular zone-granule cell layer (SGZ-GCL) across the three groups. C1–E3 illustrate examples of BrdU+ cells that differentiated into neurons in naïve control (C1–C3), GWI-VEH (D1–D3) and GWI-CUR (E1–E3) groups. The bar charts in J and K compare percentages of BrdU+ cells expressing NeuN (J) and net neurogenesis (K) across the three groups. F1–H2 show the distribution of doublecortin+ (DCX+) neurons from animals belonging to naïve control (F1, F2), GWI-VEH (G1, G2), and GWI-CUR (H1, H2) groups. The bar chart in L compares the number of DCX+ neurons across the three groups. *p < 0.05, **p < 0.01, and ***p < 0.001. Scale bar, A1, A2, A3, F1, G1, and H1 = 200 μm; B1, B2, B3, F2, G2 and H2 = 50 μm; C1–E3 = 25 μm.

Fig. 5.

Curcumin (CUR) treatment reduced astrocyte hypertrophy in animals with Gulf War Illness (GWI). A1–C3 show the distribution and morphology of GFAP+ astrocytes in the dentate gyrus (DG; A1–A3), the CA1 subfield (B1–B3) and the CA3 subfield (C1–C3) from a naïve control animal (A1, B1, C1), a GWI animal receiving vehicle (GWI + VEH, A2, B2, C2) and a GWI animal receiving CUR (GWI-CUR; A3, B3, C3). The bar charts in D-G compare the area fraction (AF) of GFAP+ structures in the DG (D), the CA1 subfield (E), the CA3 subfield (F), and the entire hippocampus (G) between different groups. *p < 0.05, **p < 0.01, and ***p < 0.001. Scale bar, A1–C3 = 50 μm.

Fig. 6.

Curcumin (CUR) treatment reduced the occurrence of activated microglia. A1–B3 show the distribution of ED-1+ structures in the dentate gyrus (DG; A1,B1), the CA1 subfield (A2,B2) and the CA3 subfield (A3,B3) from a GWI animal receiving vehicle (GWI + VEH; A1–A3) and GWI animal receiving CUR (GWI-CUR; B1–B3). The bar charts in CF compare the numbers of ED-1+ structures in the DG (C), the CA1 subfield (D), the CA3 subfield (E) and the entire Hippocampus (F). G1–H3 show the distribution of IBA-1 positive microglia expressing ED-1 in animals belonging to GWI-VEH (G1–G3) and GWI-CUR (H1–H3) groups. The bar chart in I compares percentages of IBA1+ microglia expressing ED-1 across the three groups. *p < 0.05, and **p < 0.01. Scale bar, A1–B3 = 100 μm; G1–H3 = 25 μm.

Fig. 7.

Curcumin (CUR) treatment increased the expression of genes encoding multiple antioxidant proteins in Gulf War Illness (GWI) animals. Figure A is a cluster diagram comparing the relative expression of antioxidant genes between naive control animals receiving vehicle (VEH group), animals receiving exposure to GWI chemicals and stress (GWI group), and animals receiving GWI-related chemicals and stress followed by CUR treatment (GWI + CUR group). The bar charts (B1–B17) illustrate the increased expression of 18 genes encoding different antioxidant proteins in GWI + CUR group, in comparison to VEH and GWI groups. p < 0.05, **p < 0.01, and ***p < 0.001.

Fig. 8.

Curcumin (CUR) treatment normalized the expression of genes encoding multiple proteins relevant to the mitochondrial electron transport chain (ETC) in Gulf War Illness (GWI) rats. The figure A is a cluster diagram comparing the relative expression of genes related to the mitochondrial ETC between naive control animals receiving vehicle (VEH group), animals receiving exposure to GWI chemicals and stress (GWI group), and animals receiving GWI-related chemicals and stress followed by CUR treatment (GWI + CUR group). Arrows in A point to genes that showed increased expression in the GWI group but displayed normalized expression in the GWI + CUR group. The bar charts (B1–H5) illustrate the normalized expression of 24 genes related to the mitochondrial ETC in GWI + CUR group, in comparison to their increased expression in the GWI group. These include genes related to mitochondrial complexes I (B1–B6), II (C1–C2), III (D1–D2), IV (E1–E5) and V (F1–F2) and some other genes related to mitochondrial respiration (G1–G6). p < 0.05, **p < 0.01, and ***p < 0.001.