An optimized method to visualize lipid droplets in brain tissue demonstrates their substantial accumulation in aged brains

Lipid droplets (LDs) are cellular stores for lipids. These organelles have recently gained interest in neuroscience because they accumulate in various cell types in neurodegenerative diseases. However, their role under physiological conditions is still not fully understood. Classical LD staining methods, which use lipophilic dyes like BODIPY 493/503 (BD493) or antibodies against LD coat proteins, show very few LDs in healthy brain tissue. Our recently developed novel endogenous LD reporter mouse challenges this view. We have been able to detect numerous LDs in healthy brain tissue from both adult and developing mice without staining. To understand why classical staining and endogenous labeling yield different results, we thoroughly investigated the effects of tissue preparation and detergent used in LD detection. We found that BD493 works poorly in brain tissue, while other lipophilic dyes visualize many LDs. We also found that antibody-based LD detection depends on tissue pretreatment and detergent concentration but can reveal a similar number of LDs as observed with the endogenous LD reporter mouse. Taken together, we here present an optimized procedure for LD detection in brain tissue using commercially available dyes and antibodies. Using these methods, we demonstrate that LDs are numerous in healthy brain tissue and substantially accumulate in aged brains in various cell types, including neurons.