Related Experiment Videos
Dual effect of phosphatase inhibitors on calcium channels in intestinal smooth muscle cells
Abstract:
The effects of okadaic acid (OA) and calyculin A (CL-A), potent inhibitors of protein phosphatases type 1 (PP1) and type 2A (PP2A), on inward current carried by Ba2+ through voltage-dependent Ca2+ channel in guinea pig teniae coli smooth muscle cells were investigated using whole-cell patch-clamp technique. High concentrations of OA (5 x 10(-8)-5 x 10(-6) M) and CL-A (10(-9)-10(-7) M) dose dependently increased the inward current. The concentration producing apparent half-maximum enhancing effect values for OA and CL-A were 1.12 x 10(-7) and 1.78 x 10(-9) M, respectively. CL-A appeared to be approximately 100-fold more potent in increasing the inward current than OA. Lower concentrations of OA (10(-10)-2 x 10(-8) M) and CL-A (10(-11)-10(-9) M) decreased the inward current. The maximum inhibitory effects of OA and CL-A were observed at 10(-8) M OA and 5 x 10(-10) M CL-A, respectively. CL-A is approximately 100 times more effective inhibitor of PP1 than OA, and lower concentrations of OA and CL-A used in the present study inhibit PP2A activity, but they have no or little effect on PP1 activity (Ishihara, H., B. L. Martin, D. L. Brautigan, H. Karaki, H. Ozaki, Y. Kato, N. Fusetani, S. Watabe, K. Hashimoto, D. Uemura and D. J. Hartshorne. Biochem. Biophys. Res. Commun. 159: 871-877, 1989). In the absence of ATP in pipette solution, OA and CL-A did not affect the inward current.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
Okadaic acid (OA) and calyculin A (CL-A) affect barium currents in smooth muscle cells by inhibiting protein phosphatases. High concentrations enhance currents, while low concentrations inhibit them, depending on phosphatase activity.
Area of Science:
- Pharmacology and Physiology
- Cellular Electrophysiology
- Smooth Muscle Biology
Background:
- Protein phosphatases type 1 (PP1) and type 2A (PP2A) regulate cellular functions.
- Okadaic acid (OA) and calyculin A (CL-A) are potent inhibitors of PP1 and PP2A.
- Voltage-dependent calcium channels (Ca2+) are crucial for smooth muscle contraction.
Purpose of the Study:
- To investigate the effects of OA and CL-A on Ba2+ currents through voltage-dependent Ca2+ channels.
- To determine the dose-dependent effects of OA and CL-A on guinea pig teniae coli smooth muscle cells.
- To elucidate the role of PP1 and PP2A inhibition in modulating Ca2+ channel activity.
Main Methods:
- Whole-cell patch-clamp technique was employed to measure inward currents.
- Guinea pig teniae coli smooth muscle cells were utilized.
- Varying concentrations of OA and CL-A were applied to assess their effects.
Main Results:
- High concentrations of OA and CL-A dose-dependently increased Ba2+ inward currents.
- CL-A was approximately 100-fold more potent than OA in enhancing currents.
- Low concentrations of OA and CL-A decreased Ba2+ inward currents, with CL-A being more potent.
Conclusions:
- Inhibition of PP2A by OA and CL-A modulates Ca2+ channel activity in smooth muscle.
- The effects are concentration-dependent, with high doses enhancing and low doses inhibiting currents.
- ATP is required for the modulatory effects of OA and CL-A on these currents.