Temperate insects spend over half their lives overwintering, during which most enter diapause; a pre-programmed state of developmental arrest. One of the most consistent physiological changes associated with diapause is metabolic suppression. All diapausing insects suppress their metabolic rates, but we have a limited understanding of what drives this metabolic suppression. Some insects degrade their muscles during the winter, however we do not know the extent to which this degradation contributes to metabolic suppression, especially at the mitochondrial level. In this study, we investigated the physiological mechanisms that drive metabolic suppression during diapause in the Colorado potato beetle (CPB), which suppress their metabolic rate by ~88% during diapause. We found that there is a gradual suppression of mitochondrial respiration rate as CPB enter diapause, and those in diapause have virtually undetectable respiration rates. This is, in part, driven by the breakdown of mitochondria in flight muscle, which we confirmed using MitoTracker staining, transmission electron microscopy, and citrate synthase enzyme assays. Furthermore, diapausing CPB show increased expression of parkin (a kinase involved in tagging mitochondria for removal), decreased expression of atg5 (an autophagy-related protein involved in later stages of mitochondrial removal), and no change in mitochondrial biogenesis gene expression. These results suggest that mitochondria are tagged, but not fully disposed of during diapause in CPB, resulting in a decrease in functional mitochondria and suppressed metabolism. This study will help contribute to our understanding of how insects regulate mitochondrial abundance and function, and provide new insights into the mechanisms underlying diapause and metabolic suppression.