Roles of Ca2+ in human neutrophil responses to receptor agonists.
Previous studies have concluded that cytosolic Ca2+ [( Ca2+]i) transients are essential for neutrophils (PMN) to degranulate and make superoxide anion when challenged with the receptor agonists N-formyl-methionyl-leucyl-phenylalanine, platelet-activating factor and leukotriene B4. This view is based on the profound unresponsiveness of PMN that have their [Ca2+]i fixed at resting levels by removing storage Ca2+ and loading the cells with greater than or equal to 20 microM of a Ca2+ chelator, quin2 AM. We too observed this unresponsive state in PMN loaded with 10-32 microM-quin2 AM, fura-2 AM or 1,2-bis-(2-aminophenoxy) ethane-NNN'N'-tetra-acetic acid (BAPTA). When loaded with less than or equal to 1 microM fura-2 AM, however, Ca(2+)-depleted PMN failed to alter [Ca2+]i appreciably, yet still had substantial degranulation and superoxide-anion-generating responses to the receptor agonists. Function thus did not require [Ca2+]i transients. Moreover, Ca(2+)-depleted PMN had 20-35% decreases in receptor numbers for each of the three agonists, and chelator loading of these cells decreased receptor availability by 30-50%. All receptor losses were reversed by incubating PMN with Ca2+ at 37 degrees C, but not at 4 degrees C, and agonist binding at 4 degrees C was not influenced by the presence or absence of extracellular Ca2+. Ca2+ thus caused PMN to up-regulate their agonist receptors at 37 degrees C, and the effect persisted at 4 degrees C regardless of ambient Ca2+. We conclude that Ca2+ acts in at least three ways to regulate responses to receptor agonists. First, some pool of (probably cellular) Ca2+ maintains receptor expression. Second, [Ca2+]i transients potentiate, but are not required for, function. The [Ca2+]i pool may or may not be the same as that influencing receptors. Finally, another pool(s) of Ca2+ signals or permits responses. This last pool, rather than [Ca2+]i transients, appears essential for the bioactions of standard Ca(2+)-mobilizing stimuli.
O'Flaherty
J T
JT
Wake Forest University Medical Center, Department of Medicine, Winston-Salem, NC 27103.
Rossi
A G
AG
Jacobson
D P
DP
Redman
J F
JF
D002118:Calcium / Q000097:blood*; D015550:Cell Degranulation; D005966:Glucuronidase / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007975:Leukotriene B4 / Q000494:pharmacology; D009113:Muramidase / Q000378:metabolism; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000502:physiology*; D010972:Platelet Activating Factor / Q000494:pharmacology; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface / Q000378:metabolism; D044042:Receptors, Formyl Peptide; D043562:Receptors, G-Protein-Coupled; D011971:Receptors, Immunologic / Q000378:metabolism; D018102:Receptors, Leukotriene B4; D015398:Signal Transduction; D013481:Superoxides / Q000378:metabolism; D013696:Temperature; D013997:Time Factors
D010972:Platelet Activating Factor; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface; D044042:Receptors, Formyl Peptide; D043562:Receptors, G-Protein-Coupled; D011971:Receptors, Immunologic; D018102:Receptors, Leukotriene B4; C039351:platelet activating factor receptor; D013481:Superoxides; D007975:Leukotriene B4; D009240:N-Formylmethionine Leucyl-Phenylalanine; D009113:Muramidase; D005966:Glucuronidase; D002118:Calcium