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Mitochondria buffer NCX-mediated Ca2+-entry and limit its diffusion into vascular smooth muscle cells
Damon Poburko1, Kathryn Potter, Edo van Breemen
1Department of Anesthesiology, Pharmacology & Therapeutics, University of British Columbia, Vancouver, Canada V6T 1Z1; Child & Family Research Institute, Vancouver, Canada V5Z 4H4. Damon.Poburko@medecine.unige.ch
Cell Calcium
|June 30, 2006
Summary
Peripheral mitochondria and sarcoplasmic reticulum buffer calcium influx during sodium-calcium exchanger (NCX) reversal in vascular smooth muscle cells, crucial for regulating blood pressure.
Area of Science:
- Cardiovascular Physiology
- Cellular Biology
- Biochemistry
Background:
- The reverse mode of the sodium-calcium exchanger (NCX) facilitates calcium entry in vascular smooth muscle cells (VSMCs) during agonist stimulation.
- NCX reversal is implicated in the pathogenesis of salt-sensitive hypertension.
Purpose of the Study:
- To investigate the role of peripheral mitochondria in buffering calcium entry mediated by NCX reversal in rat aortic smooth muscle cells (RASMCs).
- To elucidate the interplay between mitochondria and the sarcoplasmic reticulum in managing intracellular calcium dynamics during NCX reversal.
Main Methods:
- Measurement of mitochondrial ([Ca2+]MT) and sub-plasmalemmal ([Ca2+]subPM) calcium using targeted aequorins.
- Quantification of bulk cytosolic calcium ([Ca2+]i) with fura-2.
- Pharmacological inhibition of sarcoplasmic reticulum and mitochondrial calcium uptake using cyclopiazonic acid, FCCP, and ruthenium red.
- Assessment of NCX activity using extracellular sodium substitution and ATP stimulation, with selective NCX blocker KB-R7943.
Main Results:
- Sodium substitution induced rapid mitochondrial and sub-plasmalemmal calcium increases, followed by slower cytosolic calcium elevation.
- Inhibition of sarcoplasmic reticulum or mitochondrial calcium uptake enhanced sub-plasmalemmal calcium and abolished the delay in cytosolic calcium response.
- Extracellular ATP triggered a peak and plateau in cytosolic calcium, with the plateau sensitive to the NCX blocker KB-R7943.
- Mitochondrial NCX inhibition revealed an agonist-induced mitochondrial calcium flux blocked by KB-R7943.
Conclusions:
- Mitochondria and sarcoplasmic reticulum collaborate to buffer cytosolic calcium fluctuations resulting from agonist-induced NCX reversal.
- This buffering mechanism plays a significant role in regulating calcium homeostasis in vascular smooth muscle cells.
- Understanding these processes offers insights into potential therapeutic targets for hypertension.