This work demonstrates manganese-enhanced magnetization transfer (MT) MRI to improve the contrast of myelinated structures in mouse brain in vivo. Systemic administration of manganese chloride led to a reduction of the MT ratio by 23% in white matter and 35% in gray matter. The effect increased their contrast-to-noise ratio by 48% and facilitated a mapping of myelin-rich white matter tissues. Relaxation time measurements revealed the manganese-induced shortening of T1 to be smaller in the corpus callosum (-42%) than in the cortex (-52%) or hippocampus (-60%). These findings are in line with the assumption that a high myelin and correspondingly low water content hinder the free diffusion and uptake of manganese ions. The resulting preferential accumulation of manganese in gray matter structures causes a stronger reduction of the MT saturation in gray matter than in white matter. Extending MRI assessments with conventional MT contrast, manganese-enhanced MT MRI at 76 x 80 x 160 microm(3) resolution and 2.35 T field strength allowed for a delineation of small myelinated structures such as the fornix, mammillothalamic tract, and fasciculus retroflexus in the living mouse brain.