Fish oil promotes survival and protects against cognitive decline in severely undernourished mice by normalizing satiety signals

Abstract

Severe malnutrition resulting from anorexia nervosa or involuntary starvation leads to low weight, cognitive deficits and increased mortality rates. In the present study, we examined whether fish oil supplementation, compared with that of canola oil, would ameliorate the morbidity and mortality associated with these conditions by normalizing endocannabinoid and monoaminergic systems as well as other systems involved in satiety and cognitive function within the hypothalamus and hippocampus. Female Sabra mice restricted to 40% of their daily food intake exhibited decreased body weight, were sickly in appearance, displayed cognitive deficits and had increased mortality rates. Strikingly, fish oil supplementation that contains high omega-3 fatty acids levels decreased mortality and morbidity, and normalized the expression of genes and neurotransmitters in the hippocampus and hypothalamus. Fish oil supplementation, but not canola oil, increased survival rates, improved general appearance and prevented cognitive decline, despite the facts that both diets contained an equivalent number of calories and that there were no differences in weight between mice maintained on the two diets in 100% but decrease in the 40%. In the hypothalamus, the beneficial effects of fish oil supplementation were related to normalization of the endocannabinoid 2-arachidonylglycerol, serotonin (5-HT) (P<.056), dopamine, neuropeptide Y (NPY) and Ca(2+)/calmodulin (CaM)-dependent protein kinase (Camkk2). In the hippocampus, fish oil supplementation normalized 5-HT, Camkk2, silent mating type information regulation 1 and brain-derived neurotrophic factor. In conclusion, dietary supplements of fish oil, as source of omega-3 fatty acids, may alleviate cognitive impairments associated with severe diet restriction and prolong survival independently of weight gain by normalizing neurochemical systems.

Fig. 1

Female Sabra mice were assigned to one of the following four groups: Group 1: 100% Diet with canola oil (weight maintenance) Group 2 :100% diet with fish oil .Group 3: 40% diet with canola oil .Group 4: 40% diet with fish oil .The study was terminated at 12 days. The subjects were weighed on days 1, 5, 8, and 12. Mice weight decreased following DR to 40% and was further decreased by fish oil supplementation on the 8^th d. Values are means ±SEM. N=10 mice per group. Means with a common letter differ,*: p<0.05 corrected for multiple comparisons.

Fig. 2

Mice appearance following control, DR and fish oil supplementation (Representative mice from each group are presented). Mice maintained on the 100% diet presented a healthy appearance and appeared well groomed, regardless of oil supplementation (Fig2a, b). However, the 40% diet supplemented with canola oil displayed matted, ungroomed, and piloerect fur, and were sickly in appearance (Fig 2c). Strikingly, fish oil supplementation to mice on the 40% diet appeared was protective in terms of general appearance(Fig 2d), as these mice appeared more similar to the mice maintained on the 100% diet than those supplemented with canola oil on the 40% diet, despite their profound weight loss.

Fig. 3

Cognitive function in the eight arm maze test. Cognitive function was impaired following DR to 40% and was improved by fish oil supplementation. Values are means±SEM. N=34 for control, 20 for 4% fish oil, 32 for DR to 40% and 25 for DR to 40% + 4% fish oil. Means with a common letter differ,. p<0.05 corrected for multiple comparisons.

Fig. 4

Survival rates after 2wk of experiment. 100% of the mice not subjected to diet restriction survived while only 55% of the DR to 40% mice and 82.5% of the DR mice fed with fish oil survived. N=40 mice per group at the beginning of the experiment. N=40 for control and 4% fish oil, 22 for DR to 40% and 33 for DR to 40% + 4% fish oil at the end of the experiment. Means with a common letter differ, p<0.05,p<0.01.

Fig. 5

Alterations in the endocannabinoid system in the hypothalamus. (a) 40% diet with canola oil supplement resulted in a significant decrease in hypothalamic 2-AG compared to the 100% diet groups p<0.05a. Fish oil supplement normalized 2-AG levels in this brain region of mice receiving 40% diet p<0.05b. (b) Diet restriction decreased significantly p<0.05a while oil supplementation did not affect anandamide levels. (C) More CB1 receptor binding and(D) GTPγS binding activity were found in the 100% diet group given canola oil compared to each of the other three groups, which did not differ from one another.

Fig. 6. Monoamine concentrations and gene expression in the hypothalamus

(A)The 40% diet group supplemented with canola oil possessed significantly decreased levels of 5-HT in the hypothalamus compared to the 100% diet group supplemented with canola oil (p<0.05a). Fish oil supplementation increased 5-HT levels in the hypothalamus in 40% (p<0.056)diet groups . (B)Dopamine concentration decreased in the hypothalamus in the 40% diet group given canola oil extract p<0.05a. Fish oil supplement restored dopamine concentration in the 40% diet group (p<0.05b). (C)NPY mRNA expression increased following DR to 40% (p<0.05b) and was restored by fish oil supplementation (p<0.05c). Camkk2 mRNA expression increased following DR to 40% (p<0.05a) and was restored by fish oil supplementation (p<0.05b). Values are means±SEM.

Fig. 7

Biochemical alterations in the endocannabinoid system in the hippocampus.(A) No differences were detected in hippocampal 2-AG levels following diet restriction while there was significant increase following oil supplementation. (b) Niether DR nor Fish oil Fish oil change anandamide levels. More CB1 receptor binding and GTPγS binding activity was found in the 100% diet group given canola oil compared to each of the other three groups, which did not differ from one another.

Fig 8. Monoamine concentrations and gene expression in the hippocampus

(A)The 40% diet group supplemented with canola oil possessed significantly decreased levels of 5-HT in the hippocampus compared to the 100% diet group supplemented with canola oil (p<0.05a). Fish oil supplementation increased 5-HT levels in the hippocampus in the 40% diet group compared to the canola oil supplemented 40% diet group (p<0.05b). (B) The 40% diet group supplemented with fish oil possessed significantly higher concentrations of norepinephrine than 40% groups (p<0.05a) which is consistent with the enhanced performance of these mice in acquisition of the radial arm task. Gene expression (C)The expression of mRNA for Camkk2 increased in the hippocampi of the 40% diet group supplemented with canola oil compared to the corresponding 100% diet group (p<0.05a). The 4% fish oil supplementation in the 40% diet group fully normalized Camkk2 mRNA (p<0.05b). (D)BDNF expression increased following fish oil supplementation to both the control group p<0.05a and the DR group p<0.05b. (E) SIRT-1 expression increased following fish oil supplementation, both in the control group (p<0.05a) and the DR group(p<0.05b)