Yeast-like and filamentous fungi contain vacuoles which are similar to mammalian lysosomes and plant vacuoles, and have a variety of important functions in solute storage, protein turnover, ion homeostasis and apoptosis. Fungal vacuoles are unusual in their wide variety of architectures and roles in different species and in different cell types – their morphology and dynamics reflecting their ecological specialisation. Filamentous fungi have a network of interconnected spherical and tubular vacuole structures which may form the basis of a solute transport system that acts as nutrient transport pipelines. Some filamentous fungi fill entire cellular compartments with vacuole, which reduces the metabolic demands for cytoplasm biosynthesis and markedly affects cell cycle timing. Vacuoles are also highly dynamic, undergoing a continuous balance of fusion and fission reactions to allow changes in size, shape and number during cell division and in response to osmotic stress. This article summarises recent developments in our understanding of the dynamics, regulation and functions of fungal vacuoles.