Nomura Y, Asano M, Ito K, Uyama Y, Imaizumi Y, Watanabe M
Department of Pharmacology, Nagoya City University Medical School, Japan.
J Pharmacol Exp Ther. 1996 Nov;279(2):830-7.
To determine the Ca(+2)-buffering function of sarcoplasmic reticulum (SR) in the resting state of arterial smooth muscle, the effects of cyclopiazonic acid (CPA) and thapsigargin, which inhibit the Ca(+20-ATPase of SR, on tension and cytosolic Ca+2 level ¿[Ca+2]¿ were examined in endothelium-denuded strips of rat femoral artery. In resting strips preloaded with fura-PE3, the addition of CPA (10 microM) or thapsigargin (100 nM) caused an elevation of [Ca+2]i and a contraction. These responses were inhibited by blockers of L-type voltage-dependent Ca+2 channels (verapamil, nifedipine and diltiazem). The inhibition by verapamil was much greater for contraction than for [Ca+2]i Verapamil itself decreased the resting [Ca+2]i without affecting the resting tone. In a Ca(+2)-free solution, CPA caused a small elevation of [Ca+2]i that was not accompanied by a contraction. In the presence of CPA or thapsigargin, the effects of caffeine (20 mM) on [Ca+2]i and tension were greatly inhibited. In strips where the SR Ca+2 was depleted by the addition of caffeine and a Ca(+2)-free solution, the Ca+2 influx from the extracellular space was completely taken up into the SR. This process was inhibited by CPA or thapsigargin, which resulted in a contraction. This contraction was also inhibited by verapamil. However, the basal Ca+2 influx measured by a 5-min incubation of the artery with 45Ca+2 was not increased by CPA or thapsigargin. The net Ca+2 entry measured in a 30-min incubation of the artery with 45Ca+2 was decreased by CPA or thapsigargin. The basal 45Ca+2 efflux from the artery was increased during the addition of CPA. These results suggested that the basal Ca+2 influx was completely buffered by Ca+2 uptake into the SR in the resting state of rat femoral artery and therefore the inhibition of SR Ca+2 uptake by CPA or thapsigargin caused an elevation of [Ca+2]i and a contraction. During this process, the basal Ca+2 influx via L-type voltage-dependent Ca+2 channels mainly contributed to the contraction. The present study also showed the existence of a relatively large compartment of cytosolic Ca+2 that does not contribute to contraction during the addition of CPA or thapsigargin.
为了确定动脉平滑肌静息状态下肌浆网（SR）的Ca(+2)缓冲功能，我们在大鼠股动脉去内皮条上研究了环匹阿尼酸（CPA）和毒胡萝卜素对张力和胞质Ca+2水平¿[Ca+2]¿的影响，这两种物质可抑制SR的Ca(+20-ATP酶。在预先加载fura-PE3的静息条中，加入CPA（10 microM）或毒胡萝卜素（100 nM）会导致[Ca+2]i升高和收缩。这些反应被L型电压依赖性Ca+2通道阻滞剂（维拉帕米、硝苯地平和地尔硫卓）抑制。维拉帕米对收缩的抑制作用比对[Ca+2]i的抑制作用大得多。维拉帕米本身可降低静息[Ca+2]i，而不影响静息张力。在无Ca(+2)溶液中，CPA可导致[Ca+2]i小幅升高，但不伴有收缩。在存在CPA或毒胡萝卜素的情况下，咖啡因（20 mM）对[Ca+2]i和张力的影响被大大抑制。在通过加入咖啡因和无Ca(+2)溶液使SR Ca+2耗尽的条中，细胞外空间的Ca+2内流完全被SR摄取。这一过程被CPA或毒胡萝卜素抑制，导致收缩。这种收缩也被维拉帕米抑制。然而，用45Ca+2孵育动脉5分钟测得的基础Ca+2内流并未因CPA或毒胡萝卜素而增加。用45Ca+2孵育动脉30分钟测得的净Ca+2内流因CPA或毒胡萝卜素而减少。在加入CPA期间，动脉的基础45Ca+2外流增加。这些结果表明，在大鼠股动脉静息状态下，基础Ca+2内流被SR摄取Ca+2完全缓冲，因此CPA或毒胡萝卜素抑制SR Ca+2摄取会导致[Ca+2]i升高和收缩。在此过程中，通过L型电压依赖性Ca+2通道的基础Ca+2内流主要导致收缩。本研究还表明，在加入CPA或毒胡萝卜素期间，存在一个相对较大的胞质Ca+2区室，其对收缩无贡献。
