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Inhibitory effects of pertussis toxin on a depolarization-evoked Ca2+ influx in NG108-15 cells
Abstract:
Depolarized stimulation 1.5-fold increased Ca2+ influx which was inhibited by pretreatment with verapamil or LaCl3. Treatment with pertussis toxin, islet-activating protein (IAP), induced a reduction in 50 mM K+-induced Ca2+ influx and stimulated adenylate cyclase (AC) activity in NG108-15 cells. However, addition of dibutyryl cAMP or forskolin treatment elevating cAMP level exerted no effects on a depolarization-induced Ca2+ influx. Dissociated B-oligomer of IAP after treatment with dithiothreitol and ATP increased a depolarization-evoked Ca2+ influx. It is suggested that inhibitory GTP-binding protein (G1) or other IAP substrate proteins could directly be involved in Ca2+ influx via voltage-sensitive Ca2+ channel.
Insights
Pertussis toxin (PTX) inhibits calcium influx in NG108-15 cells, suggesting inhibitory GTP-binding proteins regulate voltage-sensitive calcium channels.
Area of Science:
- Neuroscience
- Cell Biology
- Pharmacology
Background:
- Voltage-sensitive calcium channels (VSCCs) are crucial for neuronal function.
- G-protein coupled receptors modulate ion channel activity.
- Pertussis toxin (PTX) is a tool to study G-protein signaling.
Purpose of the Study:
- To investigate the role of PTX-sensitive proteins in regulating calcium (Ca2+) influx through VSCCs in NG108-15 cells.
- To elucidate the mechanism by which PTX affects Ca2+ influx.
Main Methods:
- Cells were stimulated with depolarizing agents (K+ or veratridine).
- Calcium influx was measured using indicator dyes.
- Adenylate cyclase (AC) activity was assessed.
- Cells were pretreated with PTX, verapamil, LaCl3, or cAMP modulators.
Main Results:
- Depolarization increased Ca2+ influx 1.5-fold, inhibited by verapamil and LaCl3.
- PTX pretreatment reduced K+-induced Ca2+ influx and stimulated AC activity.
- Elevating cAMP levels did not affect depolarization-induced Ca2+ influx.
- The B-oligomer of PTX increased depolarization-evoked Ca2+ influx.
Conclusions:
- PTX-sensitive inhibitory GTP-binding proteins (Gi) or other PTX substrates are involved in regulating Ca2+ influx.
- These proteins likely act directly on voltage-sensitive calcium channels.