Considering that the brain constantly consumes a substantial amount of energy, the nature of its energy reserve is an important issue. Although the brain is rich in lipid content encompassing membranes, myelin sheath, and astrocytic lipid droplets, it is devoid of adipose tissue which serves as an energy reserve. Notably, glycogen represents the major energy store in the brain. While glycogen has been observed mainly in astrocytes for decades by electron microscopy, glycogen distribution in the brain has only been partially documented. The involvement of glycogen metabolism in memory consolidation, demonstrated by several research groups, has reiterated the functional significance of this macromolecule and the need for description of its comprehensive distribution in the brain. The combination of focused microwave-assisted brain fixation and glycogen immunohistochemistry permits assessment of glycogen distribution in the rodent brain. In this article, we describe glycogen distribution in the mouse brain using glycogen immunohistochemistry. We find heterogeneous glycogen storage patterns at multiple spatial scales. The heterogeneous glycogen distribution patterns may underlie local energy metabolism or synaptic activity, and its mechanistic understanding should extend our knowledge on brain metabolism in health and disease.