A pertussis toxin-sensitive g-protein mediates the α2-adrenergic receptor inhibition of melatonin release in photoreceptive chick pineal cell cultures

B. L. Pratt, J. S. Takahashi

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Abstract

The avian pineal gland is a photoreceptive organ that has been shown to contain postjunctional α2-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 α2-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 α2- 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 dosedependent 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 α2-adrenergic signal transduction in chick pineal cells.

Original languageEnglish (US)
Pages (from-to)277-283
Number of pages7
JournalEndocrinology
Volume123
Issue number1
DOIs
StatePublished - Jul 1 1988

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ASJC Scopus subject areas

  • Endocrinology

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