SARCOPLASMIC-RETICULUM CA-2+ UPTAKE AND THAPSIGARGIN SENSITIVITY IN PERMEABILIZED RABBIT AND RAT VENTRICULAR MYOCYTES
SARCOPLASMIC-RETICULUM CA-2+ UPTAKE AND THAPSIGARGIN SENSITIVITY IN PERMEABILIZED RABBIT AND RAT VENTRICULAR MYOCYTES
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DOI：
10.1161/01.res.73.5.820
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发表时间：
1993-11-01
影响因子：
20.1
通讯作者：
BERS, DM
中科院分区：
文献类型：
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作者：
HOVEMADSEN, L;BERS, DM
在透化的兔或大鼠心室肌细胞悬液中（几乎等于 10 mg/mL 蛋白质），使用 indo 1 和 Ca2+ 选择性微电极同时测量肌浆网 (SR) 摄取的 Ca2+ 和游离 [Ca2+]。在25 μmol/L钌红和10 mmol/L草酸盐存在下，兔和大鼠心室肌细胞SR吸收Ca2+的K(m)几乎等于250 nmol/L。大鼠最大Ca2+摄取率是兔的2.4倍。每毫克细胞蛋白添加 5 nmol 毒胡萝卜素 (TG) 可完全消除两个物种的 Ca2+ 吸收。摄取率 (K1/2) 半最大降低所需的 [TG] 对于兔为 55 pmol/mg 细胞蛋白，对于大鼠为 390 pmol/mg 细胞蛋白。假设泵位点的数量是抑制一半Ca2+泵活性所需的TG浓度的两倍（即TG亲和力非常高），则泵位点的密度对于兔为7.7μmol/kg湿重，对于大鼠为54.6μmol/kg湿重。尽管 Ca2+ 摄取率降低了五倍，低于最大 [TG]，但透化的肌细胞仍然能够将游离 [Ca2+] 从峰值 > 10 μmol/L 降低至 < 150 nmol/L。 TG 对 Ca2+ 吸收的相对抑制不依赖于游离 [Ca2+]。每毫克细胞蛋白添加超过 5 nmol TG 可完全消除 SR 对 Ca2+ 的吸收
Ca2+ uptake by the sarcoplasmic reticulum (SR) and free [Ca2+] were measured simultaneously with indo 1 and a Ca2+-selective minielectrode in suspensions of permeabilized rabbit or rat ventricular myocytes (almost-equal-to 10 mg/mL protein). In the presence of 25 mumol/L ruthenium red and 10 mmol/L oxalate, the K(m) for Ca2+ uptake by the SR was almost-equal-to 250 nmol/L in rabbit and rat ventricular myocytes. The maximal Ca2+ uptake rate was 2.4 times higher in rat than in rabbit. Addition of 5 nmol thapsigargin (TG) per milligram cell protein abolished Ca2+ uptake completely in both species. The [TG] necessary for a half-maximal reduction of the uptake rate (K1/2) was 55 pmol/mg cell protein for rabbit and 390 pmol/mg cell protein for rat. Assuming that the number of pump sites is two times the concentration of TG necessary to inhibit half of the Ca2+ pump activity (ie, the TG affinity is very high), the density of pump sites is 7.7 mumol/kg wet wt for rabbit and 54.6 mumol/kg wet wt for rat. Despite a fivefold decrease of the Ca2+ uptake rate by a submaximal [TG], the permeabilized myocytes were still able to lower the free [Ca2+] to < 150 nmol/L from a peak value > 10 mumol/L. The relative inhibition of Ca2+ uptake by TG did not depend on the free [Ca2+]. Addition of more than 5 nmol TG per milligram cell protein abolished Ca2+ uptake by the SR completely in