SUMMARY
Neutrophils from newborn calves have been shown to be deficient in ability to generate superoxide anion (O2-) after stimulation of the respiratory burst enzyme with the phorbol ester, phorbol 12-myristate 13-acetate (pma). This compound activates the O2--generating enzyme of bovine neutrophils through a pathway involving protein kinase C (pkc). To investigate the biochemical basis underlying this functional difference between neutrophils from newborn and adult cattle, we measured and compared the activity of the enzyme pkc in nonstimulated and pma-stimulated bovine neutrophils. Neutrophils from newborn calves (n = 5) and adult cows (n = 5) were stimulated with various concentrations of pma (0, 10, 100, and 500 ng/ml) for 3 minutes, and pkc activity was assayed in the cytosolic and the membrane fractions. In nonstimulated cells, most pkc activity was detected in the cytosolic fraction of neutrophils from newborn and adult cattle. Activity ofpkc in the cytosol was dependent on the presence of added calcium and phospholipids, whereas membrane-associated pkc in nonstimulated cells did not have such dependence. Significant differences in pkc activity were not observed between newborn and adult cattle in either the cytosolic or the membrane fractions from nonstimulated cells. Stimulation with pma caused redistribution of pkc activity in the cell (translocation) in newborns and adults, consisting of decrease in cytosolic pkc activity and increase in membrane-associated pkc activity. Similar to that in nonstimulated cells, pkc activity in cytosolic fractions from pma-stimulated neutrophils was dependent on the presence of cofactors (calcium and phospholipids), whereas pkc activity in the membrane did not have such requirement. Translocation of pkc from the cytosol to the membrane was expressed as percentage of decrease of activity in the cytosol and increase of activity in the membrane, compared with that in nonstimulated cells. Overall, significant differences were not evident in the ability of neutrophils from newborn and adult cattle to translocate pkc after stimulation with various concentrations of pma. Results indicate that differences in redistribution of pkc activity are not responsible for the difference observed in O2- production between neutrophils from newborn calves and adult cattle after cells are stimulated with pkc agonists.