Abstract
Cellular energy homeostasis is a crucial function of oxidative tissues but becomes altered with obesity, a major health problem that is rising unabated and demands attention. Maintaining cardiac lipid homeostasis relies on complex processes and pathways that require concerted actions between lipid droplets (LDs) and mitochondria to prevent intracellular accumulation of bioactive or toxic lipids while providing an efficient supply of lipid for conversion into ATP. While cardiac mitochondria have been extensively studied, cardiac LDs and their role in heart function have not been fully characterized. The cardiac LD compartment is highly dynamic and individual LD is small, making their study challenging. Here, we describe a simple procedure to isolate cardiac LDs that provide sufficient amounts of highly enriched material to allow subsequent protein and lipid biochemical characterization. We also present a detailed protocol to image cardiac LDs by conventional transmission electronic microscopy to provide two-dimensional (2D) analyses of cardiac LDs and mitochondria. Finally, we discuss the potential advantages of dual ion beam and electron beam platform (FIB-SEM) technology to study the cardiac LDs and mitochondria by allowing 3D imaging analysis.