Docosahexaenoic acid (DHA), is the major polyunsaturated fatty acid in the brain and is important for both the structure and the function of the nervous system. Mice were fed either an n-3 fatty acid deficient (n-3 Def) or adequate (n-3 Adq) diet for two generations. The mice were housed under two conditions, as a group or in isolation and the major point of the study was to determine whether n-3 fatty acid deficiency would enhance isolation-induced anxiety. Isolation stress was assessed using the novelty suppressed feeding paradigm (NSF) after a 3-week period and the test lasted a maximal duration of 10 min. The number of successful mice consuming food pellets within 5 min in the n-3 Def diet group was low in both housing conditions (group housing, 33% and isolated, 30%), but was 92% in the group housed and 50% in the isolated group when fed the n-3 Adq diet. In the subsequent 5 min period, the isolated housing group consuming the n-3 Adq diet increased up to 79% and the group housed animals fed the n-3 Def diet increased to 67%. However, those that consumed the n-3 deficient diet combined with isolation stress exhibited no increase. These results suggested that the n-3 deficient mice had increased anxiety that was enhanced by the chronic mild stress of social isolation.