Freund R K, Palmer M R
Department of Pharmacology, University of Colorado Health Sciences Center, Denver 80262, USA.
J Pharmacol Exp Ther. 1997 Mar;280(3):1192-200.
Previous studies have found that ethanol (EtOH) will consistently potentiate gamma-aminobutyric acid (GABA) receptor function in the cerebellum during beta adrenergic receptor activation. One consequence of beta adrenergic receptor stimulation is to increase cAMP levels, which, in turn, activate protein kinase A (PKA)-mediated phosphorylation of intracellular protein sites. In the present study, we investigated three cAMP analogues, two activators and one inhibitor of PKA to determine whether this cAMP-mediated second-messenger system may be one mechanism involved in the previously observed beta adrenergic interaction of EtOH with the GABA(A) receptor. Furthermore, because the phosphorylation state of the GABA(A) receptor may be an important determinant of function, we investigated the effect of the block of phosphatase activity on EtOH/GABA receptor interactions. We found that similar to the beta adrenergic agonist isoproterenol, local applications of the membrane-permeable cAMP analogues 8-bromo-cAMP and Sp-cAMP could modulate responses to iontophoretically applied GABA and that these modulated GABA responses were sensitized to the potentiative effects of EtOH. EtOH did not facilitate unmodulated GABA effects or GABA responses that were maximally modulated by 8-bromo-cAMP, suggesting that the cAMP mechanism mediates the observed EtOH interaction with GABA mechanisms. Furthermore, the PKA inhibitor Rp-cAMP reversed the EtOH-induced potentiation of the isoproterenol-modulated GABA responses. Finally, microcystin-LR and okadaic acid, which are type I and IIa phosphatase inhibitors, could also modulate and sensitize GABA responses to EtOH. These data suggest that beta adrenergic sensitization of GABA(A) receptors to EtOH involves the intracellular cAMP/PKA second-messenger cascade.
以往的研究发现，在β肾上腺素能受体激活过程中，乙醇（EtOH）会持续增强小脑γ-氨基丁酸（GABA）受体的功能。β肾上腺素能受体刺激的一个后果是提高环磷酸腺苷（cAMP）水平，进而激活蛋白激酶A（PKA）介导的细胞内蛋白位点磷酸化。在本研究中，我们研究了三种cAMP类似物、两种PKA激活剂和一种PKA抑制剂，以确定这种cAMP介导的第二信使系统是否可能是先前观察到的EtOH与GABA(A)受体的β肾上腺素能相互作用所涉及的一种机制。此外，由于GABA(A)受体的磷酸化状态可能是功能的一个重要决定因素，我们研究了磷酸酶活性阻断对EtOH/GABA受体相互作用的影响。我们发现，与β肾上腺素能激动剂异丙肾上腺素类似，膜通透性cAMP类似物8-溴-cAMP和Sp-cAMP的局部应用可以调节对离子导入应用GABA的反应，并且这些调节后的GABA反应对EtOH的增强作用敏感。EtOH并没有促进未调节的GABA效应或由8-溴-cAMP最大调节的GABA反应，这表明cAMP机制介导了观察到的EtOH与GABA机制的相互作用。此外，PKA抑制剂Rp-cAMP逆转了EtOH诱导的异丙肾上腺素调节的GABA反应增强。最后，I型和IIa型磷酸酶抑制剂微囊藻毒素-LR和冈田酸也可以调节并使GABA反应对EtOH敏感。这些数据表明，β肾上腺素能使GABA(A)受体对EtOH敏感涉及细胞内cAMP/PKA第二信使级联反应。
