Khalifa S, Mills S, Hruska K A
J Biol Chem. 1983 Dec 10;258(23):14400-6.
The effect of parathyroid hormone (PTH) on Ca2+ uptake was studied in brush-border membrane vesicles (BBMV) prepared from the kidneys of dogs administered 4-5 micrograms/kg of bovine PTH 1-84 in vivo. PTH stimulated Ca2+ uptake at 20 s of incubation from control values of 231 +/- 21 to 306 +/- 30 pmol/mg of protein, p less than 0.001. The stimulation of Ca2+ uptake by PTH was not reversed by incubation of the BBMV with the Ca2+ ionophore, despite the fact that Ca2+ uptake was several times greater than the expected uptake at equilibrium, indicating that most of the uptake represented Ca2+ binding to the BBMV. In BBMV from kidneys exposed to PTH, hypotonic lysis or increasing the osmolality of the solution external to the BBMV did not affect Ca2+ uptake. These data also indicated that the largest fraction of Ca2+ uptake in the presence of a chemical potential represented binding of Ca2+ to BBMV. Ca2+ binding was initially to the exterior of the BBMV, then translocated within the membrane and to the interior vesicular face as assessed by chelation of Ca2+ bound to the BBMV by ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid. Incubation of BBMV from kidneys exposed to PTH with gentamicin, which competes with Ca2+ for anionic phospholipid-binding sites, reversed the stimulatory effects of PTH on Ca2+ uptake. Phosphorylation of BBMV and PTH treatment in vivo had similar effects on BBMV phospholipid composition increasing the levels of anionic phospholipids. Phosphorylation of the BBMV also produced gentamicin-inhibitable increases in membrane Ca2+ binding. Phosphorylation of BBMV from kidneys exposed to PTH was inhibited suggesting a higher state of phosphorylation in vivo. The data demonstrate that PTH administered in vivo stimulated Ca2+ binding in BBMV that was gentamicin inhibitable and associated with an increase in the membrane content of anionic phospholipids.
研究了甲状旁腺激素（PTH）对从体内注射4 - 5微克/千克牛PTH 1 - 84的犬肾制备的刷状缘膜囊泡（BBMV）中Ca2+摄取的影响。在孵育20秒时，PTH刺激Ca2+摄取，从对照值231±21增加到306±30皮摩尔/毫克蛋白质，p＜0.001。尽管Ca2+摄取比平衡时预期的摄取量大几倍，但用Ca2+离子载体孵育BBMV并不能逆转PTH对Ca2+摄取的刺激作用，这表明大部分摄取代表Ca2+与BBMV结合。在暴露于PTH的肾脏的BBMV中，低渗裂解或增加BBMV外部溶液的渗透压均不影响Ca2+摄取。这些数据还表明，在存在化学势的情况下，Ca2+摄取的最大部分代表Ca2+与BBMV结合。Ca2+最初结合到BBMV的外部，然后通过用乙二醇双（β-氨基乙基醚）-N，N，N'，N'-四乙酸螯合结合到BBMV的Ca2+来评估，其在膜内转运并到达囊泡内部表面。用庆大霉素孵育暴露于PTH的肾脏的BBMV，庆大霉素与Ca2+竞争阴离子磷脂结合位点，可逆转PTH对Ca2+摄取的刺激作用。BBMV磷酸化和体内PTH处理对BBMV磷脂组成有类似影响，增加了阴离子磷脂水平。BBMV磷酸化也使膜Ca2+结合产生庆大霉素可抑制的增加。暴露于PTH的肾脏的BBMV的磷酸化受到抑制，提示体内存在较高的磷酸化状态。数据表明，体内给予PTH可刺激BBMV中的Ca2+结合，该结合可被庆大霉素抑制，并与阴离子磷脂的膜含量增加有关。
