Heterologous sensitization of adenylate cyclase is protein kinase A-dependent in Cath.a differentiated (CAD)-D2L cells
Heterologous sensitization of adenylate cyclase is protein kinase A-dependent in Cath.a differentiated (CAD)-D2L cells
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DOI：
10.1046/j.1471-4159.2002.01033.x
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发表时间：
2002-09-01
影响因子：
4.7
通讯作者：
Watts, VJ
中科院分区：
文献类型：
--
作者：
Johnston, CA;Beazely, MA;Watts, VJ
持续激活Ga（i/o）偶联受体导致随后药物刺激的腺苷酸环化酶活性的矛盾增强。环AMP信号通路中这种上调的确切机制（称为异源致敏）仍不明确。本研究旨在探讨环腺苷酸依赖性蛋白激酶参与的D-2L受体介导的敏化在神经元细胞环境。目前的研究中进行的Cath.a分化（CAD）细胞系稳定转染的D-2L多巴胺受体（CAD-D-2L）。长期18小时的治疗与D-2受体激动剂，quinpirole，导致毛喉素刺激的环AMP积累的两倍增强。类似地，用PKA抑制剂H89或Rp-8Br-cAMP长期治疗也增强腺苷酸环化酶活性。与此相反，长期激活蛋白激酶A（PKA）的毛喉素，异丁基甲基黄嘌呤（IBMX），或二丁酰环AMP引起的后续毛喉素刺激的环AMP的积累显着减少，并减少喹吡罗和H89诱导的异源致敏。PKA抑制剂和激活剂的作用不涉及PKA亚基表达的变化。RT-PCR分析腺苷酸环化酶同工型表达模式显示CAD-D-2L细胞中ACVI和ACIX的mRNA表达。ACVI被PKA负调控的能力与PKA抑制导致CAD-D-2L细胞中腺苷酸环化酶活性的异源致敏的观察结果一致。
Persistent activation of Galpha(i/o) -coupled receptors results in a paradoxical enhancement of subsequent drug-stimulated adenylate cyclase activity. The exact mechanism of this up-regulation in the cyclic AMP signaling pathway, known as heterologous sensitization, remains undefined. The present study was designed to investigate the involvement of cyclic AMP-dependent protein kinase in D-2L receptor-mediated sensitization in a neuronal cellular environment. The current studies were conducted in the Cath.a differentiated (CAD) cell line transfected stably with the D-2L dopamine receptor (CAD-D-2L ). Long-term 18 h treatment with the D-2 receptor agonist, quinpirole, resulted in a two-fold enhancement of forskolin-stimulated cyclic AMP accumulation. Similarly, long-term treatment with the PKA inhibitors, H89 or Rp-8Br-cAMP, also enhanced adenylate cyclase activity. In contrast, long-term activation of protein kinase A (PKA) by forskolin, isobutylmethylxanthine (IBMX), or dibutyryl cyclic AMP caused a significant reduction in subsequent forskolin-stimulated cyclic AMP accumulation and reduced both quinpirole- and H89-induced heterologous sensitization. The effects of PKA inhibitors and activators did not involve changes in PKA subunit expression. RT-PCR analysis of adenylate cyclase isoform expression patterns revealed the expression of mRNA for ACVI and ACIX in CAD-D-2L cells. The ability of ACVI to be negatively regulated by PKA is consistent with the observation that inhibition of PKA results in heterologous sensitization of adenylate cyclase activity in CAD-D-2L cells.