Anton P A, Shanahan F, Sun X P, Diehl D, Kodner A, Mayer E A
MacDonald Research Laboratories, UCLA School of Medicine 90024.
Immunopharmacol Immunotoxicol. 1993 Aug;15(4):429-46. doi: 10.3109/08923979309035238.
Vasoactive intestinal polypeptide (VIP) has been shown to stimulate adenylate cyclase in a human lymphoblast cell line (MOLT 4). In the present study, we monitored fluorescence in cell suspensions and in single fura-2 loaded MOLT 4 lymphoblasts to determine if VIP modulates intracellular calcium concentrations ([Ca2+]i), and if this modulation is mediated by adenylate cyclase. The distribution of [Ca2+]i in resting and stimulated cells was non-homogeneous, with gradients of high [Ca2+]i present in the subplasmalemmal space. In a subset of cells (10-30% of all cells studied), [Ca2+]i showed La(3+)-sensitive, temporal changes in the form of [Ca2+]i oscillations with a baseline [Ca2+]i value of 115 +/- 10 nM, an oscillation amplitude of 150 +/- 18 nM and a mean period of 9.2 +/- 2 s. The remaining non-oscillating cells showed a constant [Ca2+]i level of 75 +/- 5 nM (n = 65 cells from 4 experiments). In the subset of cells with spontaneous [Ca2+]i oscillations, VIP dose-dependently (10(-12) to 10(-8) M) increased the amplitude of oscillations but did not stimulate their frequency. The stimulatory effect of VIP was correlated with baseline [Ca2+]i in these cells, was attenuated in the presence of La3+ (25 microM), but was unaffected by cell depolarization (126 mM KCl). Dibutyryl cyclic AMP (10(-4) to 10(-3) M) and forskolin (10(-4) M) had no effect on [Ca2+]i oscillations, or on [Ca2+]i in cells without oscillations. In cell suspensions, baseline [Ca2+]i was found to be 55.1 +/- 11.2 nM (mean +/- S.E.M., n = 11); VIP, cyclic AMP analogues or forskolin had no significant effect on [Ca2+]i. These findings suggest that: a) VIP modulates the amplitude of [Ca2+]i oscillations generated by a cytosolic [Ca2+] oscillator in a subset of cells at a concentration of 10(-12) M, a thousand-fold below the KD for the VIP receptor; b) baseline [Ca2+] values may be related to both the ability of cells to generate spontaneous [Ca2+] oscillations and of oscillating cells to respond to VIP; c) due to the small number of responding cells, VIP-induced [Ca2+]i changes are not detectable when studied in cell suspensions.
血管活性肠肽（VIP）已被证明可刺激人淋巴母细胞系（MOLT 4）中的腺苷酸环化酶。在本研究中，我们监测了细胞悬液和单个负载fura - 2的MOLT 4淋巴母细胞中的荧光，以确定VIP是否调节细胞内钙浓度（[Ca2 +]i），以及这种调节是否由腺苷酸环化酶介导。静息和受刺激细胞中[Ca2 +]i的分布不均匀，在质膜下空间存在高[Ca2 +]i梯度。在一部分细胞（所有研究细胞的10 - 30%）中，[Ca2 +]i呈现出对镧（3 +）敏感的、以[Ca2 +]i振荡形式出现的时间变化，基线[Ca2 +]i值为115±10 nM，振荡幅度为150±18 nM，平均周期为9.2±2秒。其余非振荡细胞的[Ca2 +]i水平保持恒定，为75±5 nM（来自4个实验的65个细胞）。在具有自发[Ca2 +]i振荡的细胞亚群中，VIP以剂量依赖性方式（10（-12）至10（-8）M）增加振荡幅度，但不刺激其频率。VIP的刺激作用与这些细胞中的基线[Ca2 +]i相关，在存在镧（3 +）（25 microM）时减弱，但不受细胞去极化（126 mM KCl）的影响。二丁酰环磷腺苷（10（-4）至10（-3）M）和福斯可林（10（-4）M）对[Ca2 +]i振荡或无振荡细胞中的[Ca2 +]i没有影响。在细胞悬液中，发现基线[Ca2 +]i为55.1±11.2 nM（平均值±标准误，n = 11）；VIP、环磷腺苷类似物或福斯可林对[Ca2 +]i没有显著影响。这些发现表明：a）VIP在浓度为10（-12）M时调节一部分细胞中由胞质[Ca2 +]振荡器产生的[Ca2 +]i振荡幅度，该浓度比VIP受体的KD低一千倍；b）基线[Ca2 +]值可能与细胞产生自发[Ca2 +]振荡的能力以及振荡细胞对VIP的反应能力都有关；c）由于响应细胞数量较少，在细胞悬液中研究时无法检测到VIP诱导的[Ca2 +]i变化。
