Carraway Robert E, Gui Xianyong, Cochrane David E
Department of Physiology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA.
J Pharmacol Exp Ther. 2003 Nov;307(2):640-50. doi: 10.1124/jpet.102.052688.
Neurotensin (NT) stimulates Ca2+ release and Ca2+ influx in many cells. Its contractile effects in smooth muscle are inhibited by removal of Ca2+ and by Ca2+ channel blockers (CCBs). To better understand NT signaling in prostate cancer PC3 cells, blockers of voltage-gated and store-operated Ca2+ channels (VGCC and SOCC) were tested for effects on NT-binding and signaling. Eight chemical types of agents, including VGCC-blocker nifedipine and SOCC-blocker SKF-96365 (1-[beta-[3-(4-methoxyphenyl)-propoxy]-4-methoxyphenyl]-1H-imidazole), enhanced cellular NT binding up to 3-fold, while inhibiting (by congruent with 70%) NT-induced inositol phosphate (IP) formation. The ability to enhance NT binding correlated with the ability to inhibit NT-induced IP formation, and both effects were relatively specific for NT. Although cellular binding for beta2-adrenergic, V1a-vasopressin, and epidermal growth factor receptors was not enhanced by these drugs, bombesin receptor binding was increased approximately equal to 19% and bombesin-induced IP formation was inhibited approximately equal to 15%. One difference was that the effect on NT binding was Ca2+-independent, whereas the effect on IP formation was Ca2+-dependent (in part). The Ca2+-dependent part of the IP response seemed to involve SOCC-mediated Ca2+ influx to activate phospholipase C (PLC)delta, while the Ca2+-independent part probably involved PLCbeta. Photoaffinity labeling of the NT receptor NTR1 was enhanced in CCB-treated cells. NTR1 affinity was increased but NTR1 number and internalization were unchanged. Since CCBs did not alter NT binding to isolated cell membranes, the effects in live cells were indirect. These results suggest that CCBs exert two effects: 1) they inhibit NT-induced IP formation, perhaps by preventing Ca2+ influx-dependent activation of PLCdelta; and 2) they enhance NTR1 affinity by an unexplained Ca2+-independent mechanism.
神经降压素（NT）可刺激许多细胞中的Ca2+释放和Ca2+内流。其在平滑肌中的收缩作用可通过去除Ca2+和使用Ca2+通道阻滞剂（CCB）来抑制。为了更好地理解NT在前列腺癌PC3细胞中的信号传导，测试了电压门控性和储存性Ca2+通道（VGCC和SOCC）的阻滞剂对NT结合和信号传导的影响。八种化学类型的药物，包括VGCC阻滞剂硝苯地平和SOCC阻滞剂SKF-96365（1-[β-[3-(4-甲氧基苯基)-丙氧基]-4-甲氧基苯基]-1H-咪唑），可使细胞NT结合增强达3倍，同时抑制（约70%）NT诱导的肌醇磷酸（IP）形成。增强NT结合的能力与抑制NT诱导的IP形成的能力相关，且这两种作用对NT相对具有特异性。尽管这些药物未增强细胞对β2-肾上腺素能、V1a-血管加压素和表皮生长因子受体的结合，但蛙皮素受体结合增加了约19%，且蛙皮素诱导的IP形成受到约15%的抑制。一个不同之处在于，对NT结合的作用不依赖Ca2+，而对IP形成的作用部分依赖Ca2+。IP反应中依赖Ca2+的部分似乎涉及SOCC介导的Ca2+内流以激活磷脂酶C（PLC）δ，而不依赖Ca2+的部分可能涉及PLCβ。在CCB处理的细胞中，NT受体NTR1的光亲和标记增强。NTR1亲和力增加，但NTR1数量和内化未改变。由于CCB不改变NT与分离细胞膜的结合，其在活细胞中的作用是间接的。这些结果表明，CCB发挥两种作用：1）它们抑制NT诱导的IP形成，可能是通过阻止Ca2+内流依赖性激活PLCδ；2）它们通过一种未明的不依赖Ca2+的机制增强NTR1亲和力。
