P2Y receptor regulation of sodium transport in human mammary epithelial cells.
Primary human mammary epithelial (HME) cells were immortalized by stable, constitutive expression of the catalytic subunit of human telomerase. Purinergic receptors were identified by RT-PCR and quantitative RT-PCR from mRNA isolated from primary and immortalized cells grown to confluence on membrane filters. Several subtypes of P2Y receptor mRNA were identified including P2Y(1), P2Y(2), P2Y(4), and P2Y(6) receptors. RT-PCR experiments also revealed expression of A(2b) adenosine receptor mRNA in primary and immortalized cells. Confluent monolayers of HME cells exhibited a basal short-circuit current (I(sc)) that was abolished by amiloride and benzamil. When monolayers were cultured in the presence of hydrocortisone, mRNA expression of Na(+) channel (ENaC) alpha-, beta-, and gamma-subunits increased approximately threefold compared with that in cells grown without hydrocortisone. In addition, basal benzamil-sensitive Na(+) transport was nearly twofold greater in hydrocortisone-treated monolayers. Stimulation with UTP, UDP, or adenosine 5'-O-(3-thiotriphosphate) (ATPgammaS) produced increases in intracellular calcium concentration that were significantly reduced following pretreatment with the calcium-chelating agent BAPTA-AM. Concentration-response relationships indicated that the rank order of potency for these agonists was UTP > UDP > ATPgammaS. Basolateral stimulation with UTP produced a rapid but transient increase in I(sc) that was significantly reduced if cells were pretreated with BAPTA-AM or benzamil. Moreover, basolateral treatment with either charybdotoxin or clotrimazole significantly inhibited the initial UTP-dependent increase in I(sc) and eliminated the sustained current response. These results indicate that human mammary epithelial cells express multiple P2 receptor subtypes and that Ca(2+) mobilization evoked by P2Y receptor agonists stimulates Na(+) absorption by increasing the activity of Ca(2+)-activated K(+) channels located in the basolateral membrane.
Lee
So Yeong
SY
Department of Veterinary Pharmacology, College of Veterinary Medicine, Seoul National University, Seoul, South Korea.
Palmer
Melissa L
ML
Maniak
Peter J
PJ
Jang
Soo Hwa
SH
Ryu
Pan Dong
PD
O'Grady
Scott M
SM
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D018999:Charybdotoxin / Q000494:pharmacology; D002614:Chelating Agents / Q000494:pharmacology; D003022:Clotrimazole / Q000494:pharmacology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000378:metabolism; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D053503:Epithelial Sodium Channels / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D006854:Hydrocortisone / Q000494:pharmacology; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels / Q000037:antagonists & inhibitors / Q000378:metabolism; D042361:Mammary Glands, Human / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008564:Membrane Potentials; D026902:Potassium Channel Blockers; D012333:RNA, Messenger / Q000378:metabolism; D043704:Receptor, Adenosine A2B / Q000378:metabolism; D018048:Receptors, Purinergic P2 / Q000235:genetics / Q000378:metabolism*; D058472:Receptors, Purinergic P2Y1; D058473:Receptors, Purinergic P2Y2; D012964:Sodium / Q000378:metabolism*; D026941:Sodium Channel Blockers / Q000494:pharmacology; D013997:Time Factors; D014530:Uridine Diphosphate / Q000378:metabolism; D014544:Uridine Triphosphate / Q000378:metabolism
C505032:CFTR protein, human; D002614:Chelating Agents; D053503:Epithelial Sodium Channels; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels; C496923:KCNN4 protein, human; C546382:P2RY1 protein, human; C546385:P2RY2 protein, human; D026902:Potassium Channel Blockers; D012333:RNA, Messenger; D043704:Receptor, Adenosine A2B; D018048:Receptors, Purinergic P2; D058472:Receptors, Purinergic P2Y1; D058473:Receptors, Purinergic P2Y2; C507045:SCNN1A protein, human; C507048:SCNN1B protein, human; C507052:SCNN1G protein, human; D026941:Sodium Channel Blockers; C104749:purinoceptor P2Y4; C099845:purinoceptor P2Y6; C013407:benzamil; D018999:Charybdotoxin; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; C022571:adenosine 5'-O-(3-thiotriphosphate); D004533:Egtazic Acid; D014530:Uridine Diphosphate; D000584:Amiloride; D000255:Adenosine Triphosphate; D012964:Sodium; D003022:Clotrimazole; D002118:Calcium; D014544:Uridine Triphosphate; D006854:Hydrocortisone