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Regulation of L-type calcium channels in GH4 cells via A1 adenosine receptors
R Zapata1, A Navarro, E I Canela
1Departament de Bioquímica i Biologia Molecular, Facultat de Química, Universitat de Barcelona, Catalonia, Spain.
Abstract:
Identification of A1 adenosine receptors (A1Rs) in a tumor cell line derived from rat pituitary (GH4 cells) was performed by ligand binding and immunological experiments. Subsequently, the involvement of A1Rs in the regulation of calcium conductance was studied in these cells. The agonist N6-(R)-(2-phenylisopropyl)adenosine (R-PIA) did not modify the intracellular calcium basal levels, whereas it inhibited the increase produced by 15 mM KCl depolarization. The antagonist 1,3-dipropyl-8-cyclopentylxanthine led to the opening of voltage-dependent cell surface calcium channels in the absence of exogenous KCl. The channels were of the L type because the effect was abolished by calciseptine and by verapamil. These results suggest that endogenous adenosine exerts a tonic inhibitory effect on calcium transport. This was confirmed by the high adenosine concentration found in cell supernatants (up to 1 microM) and by the calcium mobilization produced by exogenously added adenosine deaminase. In depolarizing conditions, the calcium peak in the presence of adenosine deaminase was reduced when cells were preincubated with R-PIA, thus suggesting that A1R activation regulates the intensity of depolarization. These results demonstrate that adenosine is an important regulator of the physiological state of pituitary tumor cells by modulating, in an autocrine manner, the activity of L-type voltage-dependent calcium channels.
Insights
Adenosine A1 receptors (A1Rs) regulate calcium channels in pituitary tumor cells. Endogenous adenosine tonically inhibits calcium transport, modulating L-type calcium channel activity in an autocrine manner.
Area of Science:
- Endocrinology
- Neuropharmacology
- Cell Physiology
Background:
- A1 adenosine receptors (A1Rs) are present in GH4 rat pituitary tumor cells.
- The role of A1Rs in regulating calcium conductance in these cells was investigated.
Purpose of the Study:
- To identify A1Rs in GH4 cells.
- To elucidate the involvement of A1Rs in calcium channel regulation.
- To determine the autocrine role of adenosine in pituitary tumor cell physiology.
Main Methods:
- Ligand binding and immunological experiments for receptor identification.
- Pharmacological manipulation using A1R agonist (R-PIA) and antagonist (1,3-dipropyl-8-cyclopentylxanthine).
- Assessment of intracellular calcium levels following KCl depolarization and treatment with adenosine deaminase.
- Inhibition studies using calciseptine and verapamil to characterize calcium channels.
Main Results:
- A1R agonist (R-PIA) inhibited KCl-induced calcium increase, but not basal levels.
- A1R antagonist induced opening of voltage-dependent L-type calcium channels.
- Endogenous adenosine demonstrated a tonic inhibitory effect on calcium transport.
- Adenosine deaminase increased calcium mobilization, which was reduced by R-PIA pre-incubation.
Conclusions:
- Adenosine exerts tonic inhibitory control over calcium transport in pituitary tumor cells.
- A1R activation modulates the intensity of depolarization by regulating L-type voltage-dependent calcium channels.
- Adenosine acts in an autocrine fashion to regulate the physiological state of pituitary tumor cells.