To understand the process of neutrophil migration into mammary glands during mastitis, secretion of MMP-9 by bovine neutrophils was investigated. The methods of zymograms and RT-PCR were used to measure the secretory MMP-9 activity and its gene transcription, respectively. Both MMP-9 protein and mRNA were found to be expressed constitutively and greatly enhanced by PMA. However, the significant increase in the enzyme activity was found after 1 ng/ml PMA treatment for 1 h, while increasing MMP-9 mRNA transcription was only obvious after 1 ng/ml PMA treatment for 3 h. Moreover, secretion of MMP-9 protein was not inhibited by actinomycin D and cycloheximide. These results suggest that quick MMP-9 protein release from bovine neutrophils by the stimulation of PMA was due to the degranulation. As MMP-9 is an important proteinase in breakdown of the extracellular matrix and can be rapidly secreted by neutrophils through degranulation, it should play a critical role in the recruitment of neutrophils into mammary glands in bovine mastitis.