Effects of purine and pyrimidine nucleotides on intracellular Ca2+ in human eosinophils: activation of purinergic P2Y receptors.
BACKGROUND
Extracellular adenosine 5'-triphosphate (ATP) increases human eosinophil intracellular Ca(2+) concentration; the mechanism of action is not fully known. ATP, a physiologic regulator, acts through 2 purinergic receptor types: cation channels (P2X) and G protein-coupled receptors (P2Y).


OBJECTIVE
This study is aimed at identifying the functional purinergic receptors in human eosinophils.


METHODS
The relative potency of ATP, uridine (UTP), cytidine (CTP), and inosine (ITP) 5'-triphosphates (P2Y agonists); 2-methylthio-ATP (P2Y(1) agonist); and 2 P2X agonists, alpha,beta-methylene-ATP and beta,gamma-methylene-ATP on intracellular Ca(2+) concentration was examined in Ca(2+)-sensitive Fura-2-labeled human eosinophils. For comparison, ATP effects were similarly studied in human neutrophils. P2X/P2Y mRNA expression in cells was examined by reverse transcription and PCR.


RESULTS
The nucleotide potency order was UTP > or = ATP > ITP >>> 2-methylthio-ATP > alpha,beta-methylene-ATP = beta,gamma-methylene-ATP = CTP = 0 in eosinophils. Pertussis toxin (500 ng/mL) pretreatment abolished the effect of lower (10(-6) mol/L) but not higher (10(-5) mol/L) concentrations of ATP in eosinophils, whereas it attenuated the effects of 10(-4) mol/L ATP in neutrophils. The phospholipase C inhibitor U73122 (2 micromol/L) partially inhibited the effect of ATP in eosinophils but totally blocked it in neutrophils. Both cells constitutively express mRNA for P2X(1), P2X(4), P2X(5), P2Y(1), and P2Y(2), but not P2X(7), with much weaker expressions of P2X(4) and P2X(5) in neutrophils. Eosinophils cultured with the T(H)1 cytokine, IFN-gamma, expressed mRNA for P2X(7), a receptor linked to apoptosis.


CONCLUSIONS
These results suggest that the P2 purinergic receptor signal transduction pathways in eosinophils and neutrophils are different and are mediated by more than 1 subtype of functional P2Y receptors.
Mohanty
J G
JG
Division of Pulmonary/Critical Care, Department of Medicine, MCP Hahnemann University, Philadelphia 19102, USA.
Raible
D G
DG
McDermott
L J
LJ
Pelleg
A
A
Schulman
E S
ES
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology*; D001249:Asthma / Q000097:blood; D002118:Calcium / Q000097:blood*; D020013:Calcium Signaling / Q000187:drug effects*; D003570:Cytidine Triphosphate / Q000494:pharmacology*; D003907:Dexamethasone / Q000494:pharmacology; D004804:Eosinophils / Q000187:drug effects* / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000037:antagonists & inhibitors / Q000502:physiology; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D006967:Hypersensitivity / Q000097:blood; D007293:Inosine Triphosphate / Q000494:pharmacology*; D007371:Interferon-gamma / Q000494:pharmacology; D017136:Ion Transport / Q000187:drug effects; D009504:Neutrophils / Q000187:drug effects; D037342:Pertussis Toxin; D012333:RNA, Messenger / Q000096:biosynthesis; D018048:Receptors, Purinergic P2 / Q000096:biosynthesis / Q000145:classification / Q000187:drug effects* / Q000235:genetics; D011994:Recombinant Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013873:Thionucleotides / Q000494:pharmacology; D014544:Uridine Triphosphate / Q000494:pharmacology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D012333:RNA, Messenger; D018048:Receptors, Purinergic P2; D011994:Recombinant Proteins; D013873:Thionucleotides; D010566:Virulence Factors, Bordetella; D007293:Inosine Triphosphate; C005147:5'-adenylyl (beta,gamma-methylene)diphosphonate; D003570:Cytidine Triphosphate; D003907:Dexamethasone; D007371:Interferon-gamma; D000255:Adenosine Triphosphate; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins; C002630:alpha,beta-methyleneadenosine 5'-triphosphate; D002118:Calcium; D014544:Uridine Triphosphate; C008501:2-methylthio-ATP