Characterization and possible mechanisms of alpha 2-adrenergic receptor-mediated sensitization of forskolin-stimulated cyclic AMP production in HT29 cells.
Preincubation with an alpha 2-adrenergic agonist sensitized subsequent forskolin- and vasoactive intestinal peptide-stimulated cyclic AMP production in HT29 cells, a human colonic adenocarcinoma cell line. Preincubation with somatostatin, another agonist negatively coupled to adenylate cyclase, sensitized forskolin-stimulated cyclic AMP production to a lesser extent. alpha 2-Adrenergic agonist preincubation also resulted in desensitization as indicated by a shift to the right in the dose-response curve of a subsequent challenge by an alpha 2-adrenergic agonist. In an effort to elucidate the mechanism for sensitization, we examined protein kinase C and the Na+/H+ antiporter. Whereas these components had marked effects on forskolin stimulation, there was no effect on sensitization. Changes in the concentration of extra-cellular Ca2+ or Mg2+ had no effect on either forskolin stimulation or sensitization. Pertussis toxin pretreatment caused a time-dependent decrease in sensitization, an attenuation of inhibition of cyclic AMP production, and a decrease in subsequent [32P]ADP-ribosylation by pertussis toxin. The time course for these three events was similar, implicating the inhibitory guanine nucleotide regulatory protein in the mechanism for alpha 2-adrenergic receptor-mediated sensitization of forskolin-stimulated cyclic AMP production. In addition, pertussis toxin dramatically decreased forskolin-stimulated cyclic AMP production, although with a different time course. These results suggest that the mechanism of sensitization is via an as yet undefined sequence of biochemical events that includes the inhibitory guanine nucleotide regulatory protein, but does not include inhibition of adenylate cyclase nor activation of the Na+/H+ antiporter.
Jones
S B
SB
Department of Pharmacology, School of Medicine, University of Missouri-Columbia 65212.
Bylund
D B
DB
D000230:Adenocarcinoma; D037361:Adenylate Cyclase Toxin; D001792:Blood Platelets / Q000378:metabolism; D002352:Carrier Proteins / Q000037:antagonists & inhibitors; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology*; D003110:Colonic Neoplasms; D000242:Cyclic AMP / Q000096:biosynthesis*; D003513:Cycloheximide / Q000494:pharmacology; D004789:Enzyme Activation; D006801:Humans; D007700:Kinetics; D009638:Norepinephrine / Q000494:pharmacology; D037342:Pertussis Toxin; D011493:Protein Kinase C / Q000378:metabolism; D011942:Receptors, Adrenergic, alpha / Q000187:drug effects / Q000502:physiology*; D012964:Sodium / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014660:Vasoactive Intestinal Peptide / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D037361:Adenylate Cyclase Toxin; D002352:Carrier Proteins; D011942:Receptors, Adrenergic, alpha; D017923:Sodium-Hydrogen Exchangers; D010566:Virulence Factors, Bordetella; D005576:Colforsin; D014660:Vasoactive Intestinal Peptide; D003513:Cycloheximide; D012964:Sodium; D000242:Cyclic AMP; D037342:Pertussis Toxin; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D009638:Norepinephrine