Cyanotoxins produced by cyanobacteria are a significant threat to human health. However, their responses to nitrogen (N) supplies could differ between N-fixing and non-N-fixing species, which has been poorly understood. This study aimed to compare the responses of the non-N-fixing Microcystis aeruginosa and N-fixing Nostoc sp. to varying concentrations of nitrate and ammonium. This comparison had been conducted by analyzing chlorophyll-a contents, maximum quantum efficiencies of photosystem II, microcystin production, and related gene expressions. Our findings revealed that nitrate substantially stimulated the growth of both M. aeruginosa and Nostoc sp. with biomass increase by 366.2 ± 56.5 and 93.0 ± 14.0%, respectively, at 16 mg-N/L. In contrast, high ammonium concentrations suppressed their growth. Furthermore, the intracellular concentration of microcystins produced by M. aeruginosa was higher under high nitrate. Extracellular microcystins showed an opposite trend to increases in nitrate and ammonium. Ammonium increases the production and releases microcystin from Nostoc sp. N metabolism genes showed a similar trend with toxin formation genes, which were up-regulated under the high N treatments. This study provides valuable insights into the impacts of N supplies on growths of N- and non-N-fixing cyanobacteria, as well as microcystin production, which helps to develop effective strategies for managing cyanobacterial blooms.