Frieden M, Graier W F
Department of Medical Biochemistry and Medical Molecular Biology, Karl-Franzens University of Graz, Harrachgasse 21/III, A-8010 Graz, Austria.
J Physiol. 2000 May 1;524 Pt 3(Pt 3):715-24. doi: 10.1111/j.1469-7793.2000.00715.x.
The whole-cell configuration of the patch clamp technique was used to assess the involvement of ryanodine-sensitive Ca2+ release (RsCR) in histamine-activated Ca2+-dependent K+ (KCa) channels in the human umbilical vein endothelial cell line EA.hy926. Histamine (10 microM) induced a transient outward current that reached 18.9 +/- 5.5 pA pF-1 at +20 mV. This current was diminished by 1 mM tetraethylammonium or 50 nM iberiotoxin, by 90 % and 80 %, respectively, suggesting that this current results from the stimulation of large-conductance KCa (BKCa) channels. In about 50 % of the cells tested, stimulation of RsCR with 200 nM ryanodine initiated a small outward current that was also sensitive to iberiotoxin. Following the ryanodine-mediated RsCR, the potency of 10 microM histamine to activate KCa channels was reduced by about 60 %. In agreement, an inhibition of RsCR with 25 microM ryanodine diminished KCacurrent in response to histamine by about 70 %. The effect of 100 microM histamine on KCa channel activity was not reduced by previous RsCR with 200 nM ryanodine, or by an inhibition of RsCR by 25 microM ryanodine. Histamine (10 microM)-induced Ca2+ elevation was reduced by 30 % following ryanodine-mediated RsCR, whereas no inhibition occurred in the case of 100 microM histamine stimulation. In cells treated with 10 microM nocodazole for 16 h to collapse the superficial endoplasmic reticulum, 200 nM ryanodine failed to initiate any KCa current. Furthermore, the inhibitory effect of previous RsCR on 10 microM histamine-induced KCa current was not obtained in nocodazole-treated cells. Our data suggest that during moderate cell stimulation (10 microM histamine), subplasmalemmal RsCR greatly contributes to the activation of KCa channels in endothelial cells. Thus, the function of the subplasmalemmal Ca2+ control unit (SCCU) described previously must be extended as a regulator for KCa channels.
采用膜片钳技术的全细胞模式，评估了人脐静脉内皮细胞系EA.hy926中，组胺激活的钙依赖性钾（KCa）通道中，对兰尼碱敏感的Ca2+释放（RsCR）的作用。组胺（10 microM）诱导出一个瞬时外向电流，在+20 mV时达到18.9±5.5 pA pF-1。该电流分别被1 mM四乙铵或50 nM埃博霉素减少了90%和80%，这表明该电流是由大电导KCa（BKCa）通道的刺激所导致。在约50%的受试细胞中，用200 nM兰尼碱刺激RsCR引发了一个小的外向电流，该电流也对埃博霉素敏感。在兰尼碱介导的RsCR之后，10 microM组胺激活KCa通道的效力降低了约60%。同样，用25 microM兰尼碱抑制RsCR，可使组胺诱导的KCa电流减少约70%。100 microM组胺对KCa通道活性的作用，并未因先前用200 nM兰尼碱进行的RsCR或25 microM兰尼碱对RsCR的抑制而降低。兰尼碱介导的RsCR后，组胺（10 microM）诱导的Ca2+升高降低了30%，而在100 microM组胺刺激的情况下未发生抑制。在用10 microM诺考达唑处理16小时以使表面内质网解体的细胞中，200 nM兰尼碱未能引发任何KCa电流。此外，在诺考达唑处理的细胞中，先前RsCR对10 microM组胺诱导的KCa电流的抑制作用未出现。我们的数据表明，在适度的细胞刺激（10 microM组胺）期间，质膜下RsCR对内皮细胞中KCa通道的激活有很大贡献。因此，先前描述的质膜下Ca2+控制单元（SCCU）的功能，必须扩展为KCa通道的调节剂。
