A pertussis toxin-sensitive G-protein mediates the alpha 2-adrenergic receptor inhibition of melatonin release in photoreceptive chick pineal cell cultures.
The avian pineal gland is a photoreceptive organ that has been shown to contain postjunctional alpha 2-adrenoceptors that inhibit melatonin synthesis and/or release upon receptor activation. Physiological response and [32P]ADP ribosylation experiments were performed to investigate whether pertussis toxin-sensitive guanine nucleotide-binding proteins (G-proteins) were involved in the transduction of the alpha 2-adrenergic signal. For physiological response studies, the effects of pertussis toxin on melatonin release in dissociated cell cultures exposed to norepinephrine were assessed. Pertussis toxin blocked alpha 2-adrenergic receptor-mediated inhibition in a dose-dependent manner. Pertussis toxin-induced blockade appeared to be noncompetitive. One and 10 ng/ml doses of pertussis toxin partially blocked and a 100 ng/ml dose completely blocked norepinephrine-induced inhibition. Pertussis toxin-catalyzed [32P]ADP ribosylation of G-proteins in chick pineal cell membranes was assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. Membranes were prepared from cells that had been pretreated with 0, 1, 10, or 100 ng/ml pertussis toxin. In the absence of pertussis toxin pretreatment, two major proteins of 40K and 41K mol wt (Mr) were labeled by [32P]NAD. Pertussis toxin pretreatment of pineal cells abolished [32P] radiolabeling of the 40K Mr G-protein in a dose-dependent manner. The norepinephrine-induced inhibition of both cAMP efflux and melatonin release, as assessed by RIA of medium samples collected before membrane preparation, was also blocked in a dose-dependent manner by pertussis toxin. Collectively, these results suggest that a pertussis toxin-sensitive 40K Mr G-protein labeled by [32P]NAD may be functionally associated with alpha 2-adrenergic signal transduction in chick pineal cells.
Pratt
B L
BL
Department of Neurobiology and Physiology, Northwestern University, Evanston, Illinois 60208.
Takahashi
J S
JS
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D002645:Chickens; D000242:Cyclic AMP / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000502:physiology*; D007700:Kinetics; D008027:Light; D008297:Male; D008550:Melatonin / Q000378:metabolism*; D008970:Molecular Weight; D009638:Norepinephrine / Q000494:pharmacology*; D037342:Pertussis Toxin; D010870:Pineal Gland / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects; D011942:Receptors, Adrenergic, alpha / Q000187:drug effects / Q000502:physiology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*
D011942:Receptors, Adrenergic, alpha; D010566:Virulence Factors, Bordetella; D000246:Adenosine Diphosphate Ribose; D000242:Cyclic AMP; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins; D008550:Melatonin; D009638:Norepinephrine