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Protein-tyrosine kinases activate while protein-tyrosine phosphatases inhibit L-type calcium channel activity in
M Cataldi1, M Taglialatela, S Guerriero
1Section of Pharmacology, Department of Neurosciences, University of Naples Federico II, Naples, Italy.
Abstract:
The aim of this study was to evaluate the effect of protein-tyrosine kinase (PTK) and protein tyrosine phosphatase (PTP) inhibitors on Ca2+ channels in GH3 cells. The activity of Ca2+ channels was monitored either by single-cell microfluorometry or by the whole-cell configuration of the patch-clamp technique. Genistein (20-200 micron) and herbimycin A (1-15 micron) inhibited [Ca2+]i rise induced either by 55 mM K+ or 10 micron Bay K 8644. In addition, genistein and lavendustin A inhibited whole-cell Ba2+ currents. By contrast, daidzein, a genistein analogue devoid of PTK inhibitory properties, did not modify Ca2+ channel activity. The inhibitory action of genistein on the [Ca2+]i increase was completely counteracted by the PTP inhibitor vanadate (100 micron). Furthermore, vanadate alone potentiated -Ca2+-i response to both 55 mM K+ and 10 micron Bay K 8644. The possibility that genistein could decrease the [Ca2+]i elevation by enhancing Ca2+ removal from the cytosol seems unlikely since genistein also reduced the increase in fura-2 fluorescence ratio induced by Ba2+, a cation that enters into the cells through Ca2+ channels but cannot be pumped out by Ca2+ extrusion mechanisms. Finally, in unstimulated GH3 cells, genistein caused a decline of [Ca2+]i and the disappearance of [Ca2+]i oscillations, whereas vanadate induced an increase of [Ca2+]i and the appearance of [Ca2+]i oscillations in otherwise non-oscillating cells. The present results suggest that in GH3 cells PTK activation causes an increase of L-type Ca2+ channel function, whereas PTPs exert an inhibitory role.
Insights
Protein-tyrosine kinase (PTK) activation increases L-type Ca2+ channel function in GH3 cells, while protein tyrosine phosphatases (PTPs) inhibit it. PTK inhibitors like genistein reduced Ca2+ influx, and PTP inhibitors like vanadate enhanced it.
Area of Science:
- Cellular signaling
- Ion channel physiology
- Biochemistry
Background:
- Protein-tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs) are key regulators of cellular processes.
- Calcium (Ca2+) channels play critical roles in various cellular functions, including hormone secretion.
- GH3 cells are a commonly used model for studying pituitary cell function and calcium signaling.
Purpose of the Study:
- To investigate the impact of PTK and PTP inhibitors on Ca2+ channels in GH3 cells.
- To elucidate the roles of PTKs and PTPs in regulating Ca2+ channel activity and intracellular calcium levels.
Main Methods:
- Single-cell microfluorometry was used to monitor intracellular Ca2+ ([Ca2+]i) changes.
- The whole-cell configuration of the patch-clamp technique assessed ion currents.
- Specific inhibitors of PTKs (genistein, herbimycin A) and PTPs (vanadate) were employed.
Main Results:
- PTK inhibitors genistein and herbimycin A inhibited Ca2+ influx induced by K+ depolarization and Bay K 8644.
- Genistein and lavendustin A reduced whole-cell Ba2+ currents, indicating direct effects on Ca2+ channels.
- The PTP inhibitor vanadate counteracted genistein's inhibitory effects and potentiated Ca2+ responses, suggesting opposing roles for PTKs and PTPs.
Conclusions:
- PTK activation enhances L-type Ca2+ channel function in GH3 cells.
- PTPs exert an inhibitory influence on L-type Ca2+ channel activity.
- These findings highlight the intricate regulation of Ca2+ channels by kinase and phosphatase signaling pathways in GH3 cells.