Cryoconite holes (water-filled reservoirs) are considered ecologically simple ecosystems but represent biological hotspots of biodiversity on glaciers. In order to check for fine-scale spatial distribution of metazoans on the bottom of the holes, in this study, we analysed three groups of grazing invertebrates as a model: tardigrades, rotifers, and mites. We addressed differences within cryoconite holes comparing the distribution of invertebrates within and between separate holes and between glaciers at a worldwide scale. We divided each cryoconite hole into three sampling zones (established in relation to water flow on a glacier) and collected nine subsamples within cryoconite holes on glaciers in the Arctic (Longyearbreen), Norway (Blåisen), the Alps (Forni) and maritime Antarctic (Ecology Glacier). Generally, we found no consistent difference in sampling zones within cryoconite holes, which suggests homogeneity on the hole floors. However, we did find strong differences and high heterogeneity between subsamples, even within the same zone. Invertebrate densities ranged between 52 and 426 individuals per ml in subsamples collected from the same hole. We found from zero to four trdigrade species in the cryoconite hole on Longyearbreen. Our results show that benthic animals in cryoconite holes in various climatic zones have heterogeneous spatial distribution, even if no preference could be highlighted for upstream versus downstream areas with respect to water flow. The distribution of invertebrates may result from ecosystem disturbance by flushing water and animals’ active movement. Cryoconite holes, usually considered to be simple ecosystems, seem to be complex habitats where hidden spatial heterogeneity may affect abundance and diversity of organisms.