Binding, coupling, and mRNA subtype heterogeneity of alpha 1-adrenergic receptors in cultured human RPE.
In retinal pigment epithelium, apically applied epinephrine changes the conductance of specific ions which subsequently affects the membrane voltage and stimulates transepithelial fluid transport. In this investigation, myo-[3H]inositol radiotracer studies, radioligand binding with [125I]HEAT, and ribonuclease protection assays were performed to examine the coupling of this receptor to phosphoinositide hydrolysis and the specific mRNA subtypes expressed in cultured human retinal pigment epithelial cells. After labeling second to sixth passage cells with 3-muCi myo-[3H]inositol for 24 hr, epinephrine caused a dose- and time-dependent increase in [3H]inositol phosphate products (EC50 of 0.7 microM). This stimulation was antagonized by prazosin but not by propranolol. The effect of epinephrine was potentiated by the presence of the monoamine oxidase inhibitor, pargyline (10 microM). Pertussis toxin (1 microgram per well) attenuated the stimulatory effect of epinephrine. In the radioligand binding assays, [125I]HEAT binding sites varied among different cell lines, with a range of 44 to 200 fmol (mg protein)-1. Using a ribonuclease protection assay, alpha 1D and alpha 1B, but not alpha 1C, adrenergic mRNA subtypes were detected in cultured human cells. Collectively, these results show that the catecholamines act on a potentially heterogeneous population of alpha 1-adrenergic receptors coupled to phospholipase C by a pertussis toxin-sensitive G protein.
Moroi-Fetters
S E
SE
Department of Ophthalmology, Duke University Medical Center, Durham, NC, USA.
Earley
O
O
Hirakata
A
A
Caron
M G
MG
Jaffe
G J
GJ
D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004837:Epinephrine / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D010293:Pargyline / Q000494:pharmacology; D037342:Pertussis Toxin; D010716:Phosphatidylinositols / Q000378:metabolism; D010857:Pigment Epithelium of Eye / Q000187:drug effects / Q000378:metabolism*; D011224:Prazosin / Q000494:pharmacology; D011433:Propranolol / Q000494:pharmacology; D011485:Protein Binding; D012333:RNA, Messenger / Q000378:metabolism; D018340:Receptors, Adrenergic, alpha-1 / Q000235:genetics / Q000378:metabolism*; D013268:Stimulation, Chemical; D010738:Type C Phospholipases / Q000378:metabolism; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D010716:Phosphatidylinositols; D012333:RNA, Messenger; D018340:Receptors, Adrenergic, alpha-1; D010566:Virulence Factors, Bordetella; D010293:Pargyline; D011433:Propranolol; D037342:Pertussis Toxin; D010738:Type C Phospholipases; D019204:GTP-Binding Proteins; D011224:Prazosin; D004837:Epinephrine