To explore the possible mechanism of high rates of ammonium nitrate (NH₄NO₃) in the regulation of root hair formation, we grew wild type (WT), auxininsensitive and ethyleneinsensitive mutant seedlings on vertically oriented agar plates. Our study found that: (1) high rates of ammonium nitrate (NH₄NO₃) supply stimulated the branched root hair formation and inhibited root hair elongation. (2) The effect of NH₄NO₃ on root hair formation were blocked by reactive oxygen species (ROS) and diphenylene iodonium chloride (DPI), suggesting that ROS was involved in NH₄NO₃induced branched root hair formation. (3) The treatment of auxin or ethylene under high NH₄NO₃ condition completely inhibited high NH₄NO₃ induced branched root hair formation. (4) Either eir13 seedlings treated with auxin or aux17 and axr13 treated with ACC, the NH₄NO₃ induced branched root hair formation was inhibited. The results above showed that ROS, auxin and ethylene involved in high NH₄NO₃induced branched root hair formation. Auxin played a compensating role in the absence of an ethylene response and ethylene played a supplemental role in the absence of an auxin response. A higher level of auxin was required for preventing branched root hair formation in the absence of ethylene signaling.