| Abstrakt: |
Stimulation of human neutrophils with f-met-leu-phe, platelet-activating factor, or leukotriene B4 resulted in an increase in [Ca2+]i. The [Ca2+]irise was greater in the presence than absence of external Ca2+; the component that was dependent on external Ca2+was blocked by Ni2+, or could be reconstituted by addition of external Ca2+following discharge of the internal Ca2+store. These measurements of [Ca2+]iresponses provide only indirect evidence for agonist-stimulated Ca2+entry, and here we have used an alternative approach to demonstrate directly agonist-stimulated divalent cation entry. In the presence of extracellular Mn2+, f-met-leu-phe, leukotriene B4, and platelet-activating factor stimulate a quench in fluorescence of fura-2-loaded human neutrophils. This quench was due to stimulated Mn2+influx and was blocked by Ni2+. When Mn2+was added in the continued presence of agonist, after discharge of the internal store of Ca2+, a stimulated quench was seen; this result shows that an elevated [Ca2+]iis not needed for the stimulation of Mn2+entry. Depolarization by high [K+] or addition of the L-type Ca2+channel agonist, BAY-R-5417, had little or no effect on either [Ca2+]ior Mn2+entry. These results show that agonists stimulate divalent cation entry (Ca2+or Mn2+) by a mechanism independent of changes in [Ca2+]iand unrelated to voltage-dependent Ca2+channels. |
