Role of the cAMP sensor Epac as a determinant of KATP channel ATP sensitivity in human pancreatic β-cells and rat INS-1 cells
Role of the cAMP sensor Epac as a determinant of KATP channel ATP sensitivity in human pancreatic β-cells and rat INS-1 cells
复制标题
DOI：
10.1113/jphysiol.2007.143818
复制
发表时间：
2008-03-01
影响因子：
5.5
通讯作者：
Holz, George G.
中科院分区：
文献类型：
--
作者：
Kang, Guoxin;Leech, Colin A.;Holz, George G.
3 '，5'-环磷酸腺苷（cAMP）的蛋白激酶A（PKA）非依赖性作用由Epac介导，Epac是胰腺β细胞中表达的cAMP传感器。Epac可能介导β细胞ATP敏感性K+通道（K-ATP）的cAMP依赖性抑制的证据由先前对人β细胞和大鼠胰岛素分泌细胞系（INS-1细胞）的一项研究提供，其中证明Epac选择性cAMP类似物（ESCA）抑制在全细胞记录条件下测量的磺酰脲敏感性K+电流。使用来自人β细胞和大鼠INS-1细胞的质膜的切除的补丁，我们现在报告，2 '-O-Me-cAMP，激活Epac而不是PKA的ESCA，使单个K-ATP通道对ATP的抑制作用敏感，从而降低通道活性。在2 ′-O-Me-cAMP（50 μ M）存在下，描述K-ATP通道活性（NPo）的ATP依赖性抑制的剂量-反应关系左移，使得产生50%抑制的ATP浓度（IC 50）对于人β细胞从22 μ M降低至1 μ M，对于大鼠INS-1细胞从14 μ M降低至4 μ M。相反，当贴片暴露于固定浓度的ATP（10 μ M）时，给予2 ′-O-Me-cAMP以剂量依赖性和可逆的方式抑制通道活性（两种细胞类型的IC 50均为12 μ M）。抗环核苷酸磷酸二酯酶的ESCA（Sp-8-pCPT-2 '-O-Me-cAMPS）也抑制K-ATP通道活性，从而证明本文报道的ESCA的抑制作用不太可能由于其水解为腺苷的生物活性衍生物而产生。基于这些发现，可以得出结论，在人β细胞和大鼠INS-1细胞中存在一种新形式的离子通道调节，其中K-ATP通道的ATP敏感性由Epac调节。
Protein kinase A (PKA)-independent actions of adenosine 3',5'-cyclic monophosphate (cAMP) are mediated by Epac, a cAMP sensor expressed in pancreatic beta-cells. Evidence that Epac might mediate the cAMP-dependent inhibition of beta-cell ATP-sensitive K+ channels (K-ATP) was provided by one prior study of human beta-cells and a rat insulin-secreting cell line (INS-1 cells) in which it was demonstrated that an Epac-selective cAMP analogue (ESCA) inhibited a sulphonylurea-sensitive K+ current measured under conditions of whole-cell recording. Using excised patches of plasma membrane derived from human beta-cells and rat INS-1 cells, we now report that 2'-O-Me-cAMP, an ESCA that activates Epac but not PKA, sensitizes single K-ATP channels to the inhibitory effect of ATP, thereby reducing channel activity. In the presence of 2'-O-Me-cAMP (50 mu M), the dose-response relationship describing ATP-dependent inhibition of K-ATP channel activity (NPo) is left-shifted such that the concentration of ATP producing 50% inhibition (IC50) is reduced from 22 mu M to 1 mu M for human beta-cells, and from 14 mu M to 4 mu M for rat INS-1 cells. Conversely, when patches are exposed to a fixed concentration of ATP (10 mu M), the administration of 2'-O-Me-cAMP inhibits channel activity in a dose-dependent and reversible manner (IC50 12 mu M for both cell types). A cyclic nucleotide phosphodiesterase-resistant ESCA (Sp-8-pCPT-2'-O-Me-cAMPS) also inhibits K-ATP channel activity, thereby demonstrating that the inhibitory actions of ESCAs reported here are unlikely to arise as a consequence of their hydrolysis to bioactive derivatives of adenosine. On the basis of such findings it is concluded that there exists in human beta-cells and rat INS-1 cells a novel form of ion channel modulation in which the ATP sensitivity of K-ATP channels is regulated by Epac.