Auxins play an essential role in regulating plant growth and adaptation to abiotic stresses, such as nutrient stress. Our current understanding of auxins is based almost entirely on the results of research on the eudicot Arabidopsis thaliana, however, the role of the rice PIN-FORMED (PIN) auxin efflux carriers in the regulation of the ammonium-dependent response remains elusive. Here, we analyzed the expression patterns in various organs/tissues and the ammonium-dependent response of rice PIN-family genes (OsPIN genes) via qRT–PCR, and attempted to elucidate the relationship between nitrogen (N) utilization and auxin transporters. To investigate auxin distribution under ammonium-dependent response after N deficiency in rice roots, we used DR5::VENUS reporter lines that retained a highly active synthetic auxin response. Subsequently, we confirmed that ammonium supplementation reduced the DR5::VENUS signal compared with that observed in the N-deficient condition. These results are consistent with the decreased expression patterns of almost all OsPIN genes in the presence of the ammonium-dependent response to N deficiency. Furthermore, the ospin1b mutant showed an insensitive phenotype in the ammonium-dependent response to N deficiency and disturbances in the regulation of several N-assimilation genes. These molecular and physiological findings suggest that auxin is involved in the ammonium assimilation process of rice, which is a model crop plant.