Hartmann T, Kondo M, Mochizuki H, Verkman A S, Widdicombe J H
Cystic Fibrosis Research Center, University of California, San Francisco 94143.
Am J Physiol. 1992 Feb;262(2 Pt 1):L163-8. doi: 10.1152/ajplung.1992.262.2.L163.
In primary cultures of dog tracheal epithelium, isoproterenol produced a transient increase in short-circuit current (Isc) (duration 30 s; maximal increase, 32 +/- 5 microA/cm2). This was followed by a more slowly developing sustained increase (9 +/- 3 microA/cm2), which mimicked the response to N6, 2'-O-dibutyryladenosine 3',5'-cyclic monophosphate (DBcAMP). The transient and sustained responses had dissociation constants for isoproterenol of 2 x 10(-8) and 2 x 10(-9) M, respectively. Bradykinin (in the presence of indomethacin), substance P, histamine, and thrombin produced only transient increases in Isc. The time courses of these transients closely paralleled changes in concentration of intracellular Ca ([Ca2+]i) as measured with fura 2. For different mediators, there was a significant correlation between the maximal transient increase in Isc and the maximal increase in [Ca2+]i. The transients in Isc were not associated with elevation of adenosine 3',5'-cyclic monophosphate (cAMP) and were unaffected by pretreatment with DBcAMP, which abolishes the steady-state increase in response to isoproterenol. Both the transient increases in Isc and [Ca2+]i were inhibited by pretreatment with the Ca chelator 1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid. The phorbol ester 12-O-tetradecanoylphorbol 13-acetate abolished the transient increases in [Ca2+]i and Isc in response to isoproterenol but not to bradykinin. These results provide evidence that 1) isoproterenol and bradykinin elevate [Ca2+]i by different mechanisms, and 2) Ca elevation is associated with a transient increase in Isc, whereas increased cAMP is associated with a smaller sustained increase.
在犬气管上皮细胞原代培养物中，异丙肾上腺素可使短路电流（Isc）出现短暂增加（持续时间30秒；最大增加幅度为32±5微安/平方厘米）。随后是一个发展较为缓慢的持续增加（9±3微安/平方厘米），这与对N6,2'-O-二丁酰腺苷3',5'-环磷酸（DBcAMP）的反应相似。异丙肾上腺素引起的短暂和持续反应的解离常数分别为2×10⁻⁸和2×10⁻⁹M。缓激肽（在吲哚美辛存在的情况下）、P物质、组胺和凝血酶仅使Isc出现短暂增加。这些短暂变化的时间进程与用fura 2测量的细胞内钙（[Ca²⁺]i）浓度变化密切平行。对于不同的介质，Isc的最大短暂增加与[Ca²⁺]i的最大增加之间存在显著相关性。Isc的短暂变化与腺苷3',5'-环磷酸（cAMP）升高无关，并且不受DBcAMP预处理的影响，DBcAMP可消除对异丙肾上腺素的稳态增加反应。Isc和[Ca²⁺]i的短暂增加均被钙螯合剂1,2-双（2-氨基苯氧基）乙烷N,N,N',N'-四乙酸预处理所抑制。佛波酯12-O-十四酰佛波醇13-乙酸酯消除了对异丙肾上腺素而非缓激肽的[Ca²⁺]i和Isc的短暂增加。这些结果证明：1）异丙肾上腺素和缓激肽通过不同机制升高[Ca²⁺]i；2）钙升高与Isc的短暂增加相关，而cAMP增加与较小的持续增加相关。
