BACKGROUND : Dietary interventions can improve gastrointestinal (GI) symptoms. We determined the effects of fatty acids (FAs) supplementation with medium- and long-chain saturated FAs on mouse GI motility and correlated them with the expression of genes for free FA receptors (FFAR)1-4, FA binding protein 4 (FABP4) and inflammation. METHODS : Forty-eight BalbC were assigned to: standard diet (STD), diet rich in medium-chain saturated FAs (COCO) and long-chain saturated FAs (HF) (7% by weight). Body weight (BW) and food intake (FI) were monitored for 8-weeks. GI motility was determined by fecal pellet output (FPO) and colon bead expulsion tests. FABP4 inhibitor, BMS309403 (1mg/kg, ip) was injected to half of each group 2 days/week. mRNA expression of FABP4, (FFAR)1-4, and pro-inflammatory cytokines were measured in colonic and splenic tissues using real-time PCR. RESULTS : COCO and HF decreased FI. COCO accelerated overall GI transit (p<0.05). COCO increased the mRNA expression of FFAR2 (p<0.001) and TNFα (p<0.01); HF increased the expression of FABP4 and FFAR4 (p<0.05), and FFAR2 (p<0.001) in the colon, and decreased FFAR1 and FFAR4 (p<0.001), TNFα (p<0.01) and IL-1β (p<0.05) in splenic tissues. BMS309403 decreased the FI and delayed colonic transit in STD+BMS and COCO+BMS vs. STD (p<0.05). HF+BMS increased colonic expression of FFAR3 (p<0.01), TNFα (p<0.01), IL-6 (p<0.01), and reduced FFAR4 (p<0.05); COCO+BMS decreased TNFα (p<0.01). CONCLUSIONS : Diversification in the dietary lipid content affected GI motility in mice and the expression of FFARs and pro-inflammatory cytokines in vivo.