犬中枢神经不同部位磷酸二酯酶基因表达和活性测定

Determination of Gene Expression and Activity of Phosphodiesterase in Various Fractions of Canine Central Nerve

为探明磷酸二酯酶(phosphodiesterase,PDE)在犬中枢神经活动中的作用,采用反转录聚合酶链反应检测18个PDE亚型在犬中枢神经不同部位中(大脑、小脑、延髓和脊髓各段)的表达分布,以高效液相色谱法检测环核苷酸在酶反应前后的含量变化,计算PDE活性。结果显示:在所测18个PDE亚型中,犬中枢神经的8个测定部位均有PDE1A,PDE1B,PDE1C,PDE2A,PDE3A,PDE3B,PDE4A,PDE4B,PDE4C,PDE4D,PDE5A,PDE7A,PDE7B,PDE8A,PDE8B,PDE9A,PDE11A表达,PDE10A均呈阴性表达,不同部位之间表达量差异较大,其中PDE4在各部位均呈优势表达;8个测定部位cAMP-PDE活性均高于cGMP-PDE活性,且不同部位PDE活性存在差异,其中脑组织cAMP-PDE活性和cGMP-PDE活性均高于脊髓各段。结果提示PDE可能在由cAMP和cGMP介导的中枢神经经信号传导中发挥着重要作用。

To explore the function of PDE in canine central nerve, gene expression of 18 PDE subtypes in different fractions of canine central nerve （cerebrum, cerebellum, myelencephalon and spinal cord） were detected by RT-PCR,and PDE activity was calculated by the content changes of cyclic nucleotides before and after the PDE reaction examined by high performance liquid chromatography. The results showed that. In 18 PDE subtypes determined, mRNAs of PDE1A, PDE1B, PDE1C, PDE2A, PDE3A, PDE3B, PDE4A, PDE4B, PDE4C, PDE4D, PDE5A, PDETA, PDETB, PDE8A, PDE8B, PDE9A, PDE11A were expressed in all 8 determined fractions of canine central nerve, PDE 10A was negatively expressed and PDE4 was the dominant subtype in all 8 determined fractions, and expressions within different subtypes were located in different fractions; cAMP-PDE activity was higher than cGMP-PDE activity in 8 determined fractions,and the activities of both cAMP-PDE and cGMP-PDE in brain was higher than that in spinal cord. The results above indicated PDE might play an important role in central nervous information transfer mediated by cyclic nucleotides.