Improved spatial learning performance of fat-1 mice is associated with enhanced neurogenesis and neuritogenesis by docosahexaenoic acid

Abstract

Docosahexaenoic acid (DHA), an n-3 long chain polyunsaturated fatty acid (LC-PUFA), highly enriched in the central nervous system, is critical for brain development and function. It has been shown that DHA deficiency impairs cognitive performance whereas DHA supplementation improves the condition. However, the mechanisms underlying the role of DHA in brain development and function remain to be elucidated. By using transgenic fat-1 mice rich in endogenous n-3 PUFA, we show that increased brain DHA significantly enhances hippocampal neurogenesis shown by an increased number of proliferating neurons and neuritogenesis, evidenced by increased density of dendritic spines of CA1 pyramidal neurons in the hippocampus. Concurrently, fat-1 mice exhibit a better spatial learning performance in the Morris water maze compared with control WT littermates. In vitro experiments further demonstrate that DHA promotes differentiation and neurite outgrowth of neuronal cells derived from mouse ES cells and increases the proliferation of cells undergoing differentiation into neuronal lineages from the ES cells. These results together provide direct evidence for a promoting effect of DHA on neurogenesis and neuritogenesis and suggest that this effect may be a mechanism underlying its beneficial effect on behavioral performance.

Fig. 1.

Fatty acid composition in the hippocampus of WT and fat-1 transgenic mice. Fatty acids were extracted, methylated, and analyzed by gas chromatography. Linoleic acid (LA) and arachidonic acid (AA) remain the same level between WT and fat-1 mice. DHA and docosapentaenoic acid (DPA, n-3) increase whereas docosatrienoic acid (DTA, n-6) and DPA(n-6) decrease significantly in fat-1 mice. Results are means ± SEM. **, P < 0.01; n = 3, Student's t test.

Fig. 2.

Hippocampal neurogenesis is promoted in adult fat-1 mice. More BrdU+ (green) cells were found in dentate gyrus (DG) of fat-1 mice (B) than that of WT mice (A) as shown by confocal microscope. (Scale bar, 50 μm.)

Fig. 3.

Density of dendritic spines is increased in adult fat-1 hippocampus. Golgi-Cox stained dendritic spines from WT (A) and fat-1 (B) mice. Note the density of dendritic spines along dendrite. (Scale bar, 5 μm.)

Fig. 4.

DHA enhances the neurite outgrowth of neuronal cells differentiated from G-olig2 embryonic stem (ES) cells. The ES cells were induced to differentiate into neuronal cells in defined serum-free medium for 13 days. (A–D) Phase contrast microscope. Embryoid bodies (EBs) (B) and singles cells (D) treated with 5 μM DHA have more and longer neurites compared with the control EBs (A) and single cells (C). (E) Neurite analysis of length and number of neurites per cell, branch number per cell, length per neurite, and neuritogenic index (neurite number × neurite length) from C and D. Results are means ± SD (3–5 experiments). **, P < 0.01 compared with the control (Student's t test). (Scale bar for A and B, 100 μm; Scale bar for C and D, 20 μm.)

Fig. 5.

DHA (5 μM) supplementation increases the number of Tuj1+ cells after differentiation in defined serum-free medium for 13 days from G-Olig2 ES cells. Confocal images of Tuj1 immunostaining in control (A) and DHA (B) groups, Tuj1 (green), PI (red). (Scale bar, 100 μm.)

Fig. 6.

Performance in the Morris water maze is improved in fat-1 transgenic mice. The latency per testing session represents the average of 4 trials of all animals in each group. Results are means ± SEM. In hidden platform trial, the escape latency (s) of fat-1 mice (n = 6) is significantly lower than that of WT mice (n = 7); *, P = 0.016 (F_1,11 = 7.992; two-way ANOVA followed by post-hoc Tukey test).