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标题：A New Enzymatic Assay for Guanosine 3′:5′-Cyclic Monophosphate and Its Application to the Ductus Deferens of the Rat
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作者：G. Schultz ; J. G. Hardman ; K. Schultz 等
期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：1973
卷号：70
期号：6
页码：1721-1725
DOI：10.1073/pnas.70.6.1721
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：A sensitive enzymatic procedure has been developed for the determination of guanosine 3':5'-cyclic monophosphate (cyclic GMP). It is based on the conversion of cyclic GMP to GMP by cyclic nucleotide phosphodiesterase and on the transfer of 32P from [{gamma}-32P]ATP to GMP by the action of a specific ATP:GMP phosphotransferase (EC 2.7.4.8 ). The [32P]GDP is separated from the remaining [32P]ATP by enzymatic degradation of ATP by myosin and by precipitation of the 32Pi formed. The reaction blank, which is mostly caused by the nucleotide content of the enzymes, is doubled by about 0.1 pmol of cyclic GMP. The procedure has advantages in speed and/or accuracy over other methods in current use. Cyclic nucleotide concentrations were studied in the ductus deferens of the rat; two agents were used, carbachol and norepinephrine, which cause contraction. Incubation with 0.1 mM carbachol caused a 3-fold increase in cyclic GMP content, which was maximal about 2 min after carbachol addition. Cyclic AMP concentrations were not significantly changed. Addition of 0.01 mM norepinephrine increased cyclic GMP content by about 25% within 1 min and by 40% within 3 min; cyclic AMP concentrations were only slightly increased. A 3-min incubation with the phosphodiesterase inhibitor 1-methyl-3-isobutylxanthine (0.1 mM) doubled the cyclic GMP content and increased cyclic AMP concentration by 50%.
关键词：[³²P]GDP formation ; cholinergic agents ; adrenergic agents ; phosphodiesterase inhibitors ; smooth muscle