Study of the mechanisms regulating species richness in aquatic systems is essential for understanding the functioning and conservation of these environments, especially in the Neotropics, given the lack of relevant information. In 15 lakes in three geographical regions in South America, we studied the effects of lake size, the concentration of chlorophyll a, the variety of photosynthetic pigments and the temporal stability of the habitat on crustacean zooplankton richness. Lake size and concentration of chlorophyll a did not show a significant relationship with crustacean species richness. However, the degree of temporal fluctuation in habitat conditions and the variety of photosynthetic pigments explained 83% of the observed differences in species richness between habitats. These results show that the richness of Neotropical crustacean zooplankton depends mainly on the negative relationship with temporal variability of the habitat and, to a lesser extent, on the positive relationship with the variation in resource diversity. Therefore, our results show the importance that possible impacts generating environmental instability could have on the species richness of Neotropical aquatic crustaceans.