Subplasmalemmal ryanodine-sensitive Ca2+ release contributes to Ca2+-dependent K+ channel activation in a human umbilical vein endothelial cell line

1. The whole-cell configuration of the patch clamp technique was used to as sess the involvement of ryanodine-sensitive Ca2+ release (RsCR) in histamin e-activated Ca2+-dependent K+ (K-Ca) channels in the human umbilical vein e ndothelial cell line EA.hy926. 2. Histamine (10 mu M) induced a transient outward current that reached 18. 9 +/- 5.5 pA pF(-1) at +20 mV. This current was diminished by 1 mM tetraeth ylammonium or 50 nM iberiotoxin, by 90 % and 80 %, respectively, suggesting that this current results from the stimulation of large-conductance K-Ca ( BKCa) channels. 3. In about 50% of the cells tested, stimulation of RsCR with 200 nM ryanod ine initiated a small outward current that was also sensitive to iberiotoxi n. 4. Following the ryanodine-mediated RsCR, the potency of 10 mu M histamine to activate K-Ca channels was reduced by about 60%. In agreement, an inhibi tion of RsCR with 25 mu M ryanodine diminished K-Ca current in response to histamine by about 70%. 5. The effect of 100 mu M histamine on K-Ca channel activity was not reduce d by previous RsCR with 200 nM ryanodine, or by an inhibition of RsCR by 25 mu M ryanodine. 6. Histamine (10 mu M)-induced Ca2+ elevation was reduced by 30% following ryanodine-mediated RsCR, whereas no inhibition occurred in the case of 100 mu M histamine stimulation. 7. In cells treated with 10 mu M nocodazole for 16 h to collapse the superf icial endoplasmic reticulum, 200 nM ryanodine failed to initiate any K-Ca c urrent. Furthermore, the inhibitory effect of previous RsCR on 10 mu M hist amine-induced K-Ca current was not obtained in nocodazole-treated cells. 8. Our data suggest that during moderate cell stimulation (10 mu M histamin e), subplasmalemmal RsCR greatly contributes to the activation of K-Ca chan nels in endothelial cells. Thus, the function of the subplasmalemmal Ca2+ c ontrol unit (SCCU) described previously must be extended as a regulator for K-Ca channels.