An unhealthy “Western Style” high-fat diet, lack of exercise and smoking all contribute to increase cardiovascular diseases. This type of high-fat diet can cause cardiomyopathy directly by altering cardiac metabolism or indirectly due to other factors (e.g. obesity). In addition to cardiomyopathies, “Western Style” high-fat diet predisposes hearts to the damaging effects of cardiac insults. We have recently shown that high-fat diet alters mitochondria morphology, consistent with increased fission. Whether such a change is due to changes in mitochondrial proteins associated with fission/fusion is not known. The aim of this study was to determine whether a chronic “Western Style” high-fat diet, without obesity and associated co-morbidities, alters mitochondrial protein expression. Male C57/BL 6 mice aged 6 weeks were fed either normal chow diet (13% calories from fat) or a high-fat diet (45% calories from fat) for 20 weeks. In this model high-fat diet hearts and cardiomyocytes are more vulnerable to ischemia-reperfusion injury. The mice do not show signs of obesity, diabetes or hypertrophy. Hearts from high-fat diet and control were used to detect the relative expression of mitofusin-1 (Mfn-1), mitofusin-2 (Mfn-2) and FIS1 using western blotting. Mfn-1 and Mfn-2 are thought to control mitochondrial fusion and FIS1 is a regulator of mitochondrial fission. Data are presented as mean±SEM and were analysed using students t-test. A P-value of <0.05 was assumed to be significant. n = 6 hearts/group. In the high-fat diet group there was no change in the protein expression of Mfn-1 when compared to the normal diet. However, there was an increase in Mfn-2 (0.929±0.046 vs. 0.653±0.025 relative protein expression, P<0.001) in the high-fat group compared to the normal diet group. There was a reduction in FIS1 protein expression in the high-fat group versus the normal diet (0.684±0.039 vs. 0.812±0.033 relative protein expression, P<0.05). These data suggest that in the high-fat diet group there is an up regulation of mitochondrial fusion and a decrease in mitochondrial fission. This information does not correlate with our mitochondrial morphology measurements where we see a decrease in mitochondrial area (P<0.01) and length (fewer mitochondria >2.1µm (P<0.05)) in the high-fat group, suggesting more fission. Interestingly, recent work using an Mfn-2 knockout mouse model (1) also reported longer and larger mitochondria as well as increased resistance of cardiomyocytes to insults when Mfn-2 was lower. It is evident therefore that alterations in mitochondrial proteins expression are not simple indicators of fission and fusion. Our work indicates that the effects of high-fat diet on mitochondrial morphology and the associated reduced resistance of cardiomyocytes to cardiac insults could be largely due to an increase in Mfn-2.