Basal calcium entry in vascular smooth muscle

Area of Science:

• Physiology
• Cell Biology
• Pharmacology

Background:

• Basal calcium leak in smooth muscle remains poorly understood despite being identified 30 years ago.
• Characterizing this leak is crucial for understanding smooth muscle function and potential therapeutic targets.

Purpose of the Study:

• To quantify basal calcium (Ca2+) entry into rat aortic smooth muscle cells.
• To identify the ion channels responsible for this basal Ca2+ leak.
• To investigate the effects of various blockers on basal Ca2+ entry.

Main Methods:

• Utilized 45Ca2+ uptake assays in cultured rat aortic smooth muscle cells.
• Employed lanthanum and EGTA for tracer removal and retention.
• Applied blockers including gadolinium, nifedipine, SKF 96365, and 2-aminoethoxy-diphenylborate (2-APB).
• Performed 3-D image reconstruction to determine cell volumes.
• Used reverse transcriptase polymerase chain reaction (RT-PCR) to analyze TRPC mRNA expression.

Main Results:

• Quantified basal Ca2+ entry at 1.45 x 10(9) Ca2+ x cell(-1) x min(-1).
• Gadolinium blocked 80% of the basal leak, showing biphasic inhibition.
• Organic ion channel blockers inhibited approximately 80% of the leak, with specific contributions from nifedipine, SKF 96365, and 2-APB.
• RT-PCR detected TrpC1, TrpC4, and TrpC6 mRNA expression.
• 2-APB selectively inhibited TRPC4-containing channels.

Conclusions:

• Basal Ca2+ entry in smooth muscle is primarily mediated by the basal open probability of excitable Ca2+ channels.
• TRPC channels, particularly TRPC4, are likely involved in basal Ca2+ leak.
• Gadolinium and specific organic blockers are effective inhibitors of this basal Ca2+ entry pathway.