Modulation of cyclic AMP in purified rat mast cells. I. Responses to pharmacologic, metabolic, and physical stimuli.
Modulation of cyclic AMP in purified rat mast cells. I. Responses to pharmacologic, metabolic, and physical stimuli.
复制标题
纯化大鼠肥大细胞中环磷酸腺苷的调节。
作者：
T. Sullivan;K. Parker;W. Stenson;C. Parker
研究了分离的未受刺激肥大细胞环腺苷3′，5′-单磷酸腺苷（cAMP）含量及其在各种药理、代谢和物理刺激下的变化。改良的牛血清白蛋白密度梯度纯化方法始终提供95%或更高纯度的肥大细胞制剂，没有明显的损伤，并且应用于梯度的肥大细胞回收率为73%。在未受刺激的肥大细胞中测量到的cAMP很高，平均每百万细胞16皮摩尔。适度搅拌或与玻璃接触可使cAMP含量增加约2倍。当培养基中不含钙时，肥大细胞cAMP显著升高；添加钙离子浓度从1 muM增加到1 mM， cAMP含量呈线性下降。茶碱（3 ~ 20 mM）引起肥大细胞cAMP含量的剂量相关增加，约为20 mM茶碱时的2倍。肾上腺素（0.01 ~ 1 mM）引起cAMP轻度剂量相关升高。在茶碱存在的情况下，肾上腺素增加的cAMP水平等于或大于单独使用的药物的效果总和。100 muM普萘洛尔完全抑制肾上腺素引起的cAMP升高，10 muM普萘洛尔部分抑制cAMP升高，提示与β肾上腺素能受体有关。前列腺素E1 （PGE1）和组胺（在茶碱存在下）也升高cAMP。氨甲酰胆碱（1 nM）降低cAMP 38%；0.1 mM的阿托品对1 nM氨甲酰胆碱诱导的cAMP下降有83%的抑制作用，表明其与毒蕈碱受体有关。因此，在这些均质单细胞悬浮液中，胆碱能剂和肾上腺素能剂对cAMP水平有相反的影响。二氮氧化物（10 muM）和腺嘌呤（1 muM）分别使cAMP降低37%和32%。高纯度肥大细胞的可用性和降低或增加cAMP含量的试剂的鉴定使得直接检查cAMP在组胺释放中的作用成为可能。
The cyclic adenosine 3', 5'-monophosphate (cAMP) content of isolated unstimulated mast cells and the changes induced by a variety of pharmacologic, metabolic, and physical stimuli were studied. A modified bovine serum albumin density gradient purification method consistently provided mast cell preparations which were 95% or more pure, without apparent damage, and a 73% recovery of the mast cells applied to the gradients. The measured cAMP in unstimulated mast cells was high, a mean of 16 picomoles per million cells. Moderate agitation or contact with glass increased cAMP content about 2-fold. When calcium was omitted from the medium mast cell cAMP was markedly elevated; incremental increases in added calcium ion concentration from 1 muM to 1 mM caused a linear decrease in cAMP content. Theophylline (3 to 20 mM) caused a dose-related increase in mast cell cAMP content, approximately 2-fold at 20 mM theophylline. Epinephrine (0.01 to 1 mM) caused a modest, dose-related increase in cAMP. In the presence of theophylline, epinephrine increased cAMP levels equal to or greater than the sum of the effects of the agents used individually. The increase in cAMP induced by epinephrine was completely inhibited by 100 muM propranolol and partially inhibited by 10 muM propranolol, thus suggesting that a beta adrenergic receptor is involved. Prostaglandin E1 (PGE1) and histamine (in the presence of theophylline) also raised cAMP. Carbamylcholine (1 nM) lowered cAMP 38%; Atropine (0.1 mM) inhibited the decrease in cAMP induced by 1 nM carbamylcholine by 83% indicating that a muscarinic receptor is involved. In these homogeneous single cell suspensions, therefore, cholinergic and beta adrenergic agents have opposing effects on cAMP levels. Diazoxide (10 muM) and adenine (1 muM) caused 37 and 32% decreases in cAMP, respectively. The availability of highly purified mast cells and the identification of agents which either decrease or increase cAMP content allows a direct examination of the role of cAMP in histamine release.