title: P2 purinoceptors regulate calcium-activated chloride and fluid transport in 31EG4 mammary epithelia.
abstract: It has been reported that secretory mammary epithelial cells (MEC) release ATP, UTP, and UDP upon mechanical stimulation. Here we examined the physiological changes caused by ATP/UTP in nontransformed, clonal mouse mammary epithelia (31EG4 cells). In control conditions, transepithelial potential (apical side negative) and resistance were -4.4 +/- 1.3 mV (mean +/- SD, n = 12) and 517.7 +/- 39.4 Omega. cm(2), respectively. The apical membrane potential was -43.9 +/- 1.7 mV, and the ratio of apical to basolateral membrane resistance (R(A)/R(B)) was 3.5 +/- 0.2. Addition of ATP or UTP to the apical or basolateral membranes caused large voltage and resistance changes with an EC(50) of approximately 24 microM (apical) and approximately 30 microM (basal). Apical ATP/UTP (100 microM) depolarized apical membrane potential by 17.6 +/- 0.8 mV (n = 7) and decreased R(A)/R(B) by a factor of approximately 3. The addition of adenosine to either side (100 microM) had no effect on any of these parameters. The ATP/UTP responses were partially inhibited by DIDS and suramin and mediated by a transient increase in free intracellular Ca(2+) concentration (427 +/- 206 nM; 15-25 microM ATP, apical; n = 6). This Ca(2+) increase was blocked by cyclopiazonic acid, by BAPTA, or by xestospongin C. 31EG4 MEC monolayers also secreted or absorbed fluid in the resting state, and ATP or UTP increased fluid secretion by 5.6 +/- 3 microl x cm(-2) x h(-1) (n = 10). Pharmacology experiments indicate that 31EG4 epithelia contain P2Y(2) purinoceptors on the apical and basolateral membranes, which upon activation stimulate apical Ca(2+)-dependent Cl channels and cause fluid secretion across the monolayer. This suggests that extracellular nucleotides could play a fundamental role in mammary gland paracrine signaling and the regulation of milk composition in vivo.
authors.affiliation: Department of Molecular and Cell Biology, University of California at Berkeley, Berkeley, California 94720-3200, USA.
authors.forename: Sasha
authors.initials: S
authors.lastname: Blaug
authors.forename: Jodi
authors.initials: J
authors.lastname: Rymer
authors.forename: Stephen
authors.initials: S
authors.lastname: Jalickee
authors.forename: Sheldon S
authors.initials: SS
authors.lastname: Miller
mesh_terms: D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D001826:Body Fluids / Q000378:metabolism*; D002118:Calcium / Q000502:physiology*; D002460:Cell Line; D018118:Chloride Channels / Q000378:metabolism; D002712:Chlorides / Q000378:metabolism*; D017097:Electric Impedance; D004594:Electrophysiology; D004847:Epithelial Cells / Q000378:metabolism / Q000502:physiology; D005260:Female; D007425:Intracellular Membranes / Q000378:metabolism; D008321:Mammary Glands, Animal / Q000166:cytology / Q000378:metabolism* / Q000502:physiology; D051379:Mice; D009994:Osmolar Concentration; D018048:Receptors, Purinergic P2 / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013498:Suramin / Q000494:pharmacology; D014544:Uridine Triphosphate / Q000494:pharmacology
chemical_list: D018118:Chloride Channels; D002712:Chlorides; D018048:Receptors, Purinergic P2; D013498:Suramin; D000255:Adenosine Triphosphate; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D002118:Calcium; D014544:Uridine Triphosphate