Ca(2+) entry into primary cultured pig coronary smooth muscle cells after previous store depletion by repetitive P2Y purinoceptor stimulation.
Store-operated Ca(2+) entry, stimulated by depletion of intracellular Ca(2+) pools, has not been fully elucidated in vascular smooth muscle cells of pig coronary arteries. Therefore, [Ca(2+)](i) was measured in cultured cells derived from extramural pig coronary arteries using the Fura-2/AM fluorometry. Divalent cation entry was visualized with the Fura-2 Mn(2+)-quenching technique. Ca(2+) stores were depleted either by repetitive stimulation of P2Y purinoceptors with ATP (10 micromol/L), or by the sarcoendoplasmic Ca(2+)-ATPase inhibitor 2,5-Di-(tert-butyl)-1,4-benzohydroquinone (BHQ; 1 micromol/L) in Ca(2+)-free medium (EGTA 1 mmol/L). Addition of Ca(2+)(1 mmol/L) induced refilling of ATP-sensitive Ca(2+) stores and an increase in [Ca(2+)](i) in the presence of BHQ. Both could be significantly diminished by Ni(2+)(5 and 1mmol/L), La(3+)(10 micromol/L), Gd(3+)(10 micromol/L), and Mg(2+)(5.1 mmol/L). In contrast to the BHQ-mediated rise in [Ca(2+)](i), refilling of ATP-depleted stores was affected by neither flufenamate (0.1 mmol/L), nor by nitrendipine, nifedipine, and nisoldipine (each 1 micromol/L). The data suggest that after store depletion in pig coronary smooth muscle cells ATP and BHQ may converge on a common, Ni(2+)-, La(3+)-, Gd(3+)-, and Mg(2+)- sensitive Ca(2+) entry pathway, i.e. on a store-operated Ca(2+) entry. An additional contribution of the Na(+)/Ca(2+) exchanger cannot be excluded. Flufenamate-sensitive non-selective cation channels and dihydropyridine-sensitive L-type Ca(2+) channels are not involved in refilling of Ca(2+) stores after previous depletion by repetitive P2Y purinoceptor stimulation. The store-operated Ca(2+) entry in-between repetitive purinoceptor stimulation, i.e. in the absence of the agonist, may be responsible for the maintenance of agonist-induced rhythmic Ca(2+) responses.