5-HT1A and histamine H1 receptors in HeLa cells stimulate phosphoinositide hydrolysis and phosphate uptake via distinct G protein pools.
Regulation of phosphate uptake was studied in a HeLa cell line after transfection with DNA encoding the human 5-HT1A receptor. In these cells, 5-HT stimulates sodium-dependent phosphate uptake via protein kinase C activation. Endogenous histamine H1 receptors (739 +/- 20 fmol/mg protein) were identified with [3H]pyrilamine. Histamine (i) stimulated phosphoinositide hydrolysis (EC50 = 8.6 +/- 4.1 microM), (ii) activated protein kinase C (2.4-fold increase in activity), and (iii) increased phosphate uptake (EC50 = 3.2 +/- 1.8 microM) by increasing maximal transport (Vmax(basal) = 6.2 +/- 0.3 versus Vmax(histamine) = 9.1 +/- 0.4) without changing the affinity of the transport process for phosphate. Prolonged treatment with 16 microM phorbol 12-myristate 13-acetate completely blocked protein kinase C activation and markedly attenuated the stimulation of phosphate uptake induced by histamine, establishing that 5-HT and histamine stimulate phosphate uptake through the common pathway of protein kinase C activation. The linkages of the histamine H1 and 5-HT1A receptors to G protein pools were assessed in two ways. (i) The stimulation of phosphoinositide hydrolysis, protein kinase C activity, and phosphate uptake associated with histamine were insensitive to pertussis toxin, whereas those associated with 5-HT were very sensitive to pertussis toxin. (ii) The stimulation of phosphoinositide hydrolysis, protein kinase C activity, and phosphate uptake induced by histamine and 5-HT were additive. These findings suggest that distinct receptor types can stimulate phosphoinositide hydrolysis, protein kinase C, and phosphate uptake in an additive fashion through distinct pools of G proteins in a single cell type.
Raymond
J R
JR
Medical Service (Nephrology Section), Durham Veterans Administration Medical Center, North Carolina.
Albers
F J
FJ
Middleton
J P
JP
Lefkowitz
R J
RJ
Caron
M G
MG
Obeid
L M
LM
Dennis
V W
VW
D001692:Biological Transport; D015152:Blotting, Northern; D015536:Down-Regulation; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006367:HeLa Cells / Q000187:drug effects / Q000378:metabolism; D006632:Histamine / Q000494:pharmacology*; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D037342:Pertussis Toxin; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010710:Phosphates / Q000378:metabolism*; D010716:Phosphatidylinositols / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011738:Pyrilamine / Q000378:metabolism; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics / Q000302:isolation & purification; D011969:Receptors, Histamine H1 / Q000235:genetics / Q000378:metabolism*; D011985:Receptors, Serotonin / Q000235:genetics / Q000378:metabolism*; D012701:Serotonin / Q000494:pharmacology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D010710:Phosphates; D010716:Phosphatidylinositols; D012333:RNA, Messenger; D011969:Receptors, Histamine H1; D011985:Receptors, Serotonin; D010566:Virulence Factors, Bordetella; D012701:Serotonin; D015240:Phorbol 12,13-Dibutyrate; D006632:Histamine; D037342:Pertussis Toxin; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D011738:Pyrilamine; D013755:Tetradecanoylphorbol Acetate