Department of Psychiatry, University of Illinois at Chicago, Chicago, Illinois 60612, USA.
Alcohol Clin Exp Res. 2010 Mar 1;34(3):451-61. doi: 10.1111/j.1530-0277.2009.01109.x. Epub 2009 Dec 18.
The neuropeptide Y (NPY) system of the central nucleus of amygdala (CeA) has been shown to be involved in anxiety and alcoholism. In this study, we investigated the molecular mechanisms by which NPY in the CeA regulates anxiety and alcohol drinking behaviors using alcohol-preferring (P) rats as an animal model.
Alcohol-preferring rats were bilaterally cannulated targeting the CeA and infused with artificial cerebrospinal fluid (aCSF) or NPY. Alcohol drinking and anxiety-like behaviors were assessed by the 2-bottle free-choice paradigm and light/dark box (LDB) exploration test, respectively. The levels of NPY and related signaling proteins were determined by the gold immunolabeling procedure. The mRNA levels of NPY were measured by in situ RT-PCR. Double-immunofluorescence labeling was performed to observe the colocalization of NPY and Ca(2+)/calmodulin-dependent protein kinase IV (CaMK IV).
We found that NPY infusion into the CeA produced anxiolytic effects, as measured by the LDB exploration test, and also decreased alcohol intake in P rats. NPY infusion into the CeA significantly increased levels of CaMK IV and phosphorylated cAMP responsive element-binding (pCREB) protein and increased mRNA and protein levels of NPY, but produced no changes in protein levels of CREB or the catalytic alpha-subunit of protein kinase A (PKA-Calpha) in the CeA. We also observed that alcohol intake produced anxiolytic effects in P rats in the LDB test and also increased NPY expression and protein levels of pCREB and PKA-Calpha without modulating protein levels of CREB or CaMK IV, in both the CeA and medial nucleus of amygdala. In addition, we found that CaMK IV-positive cells were co-localized with NPY in amygdaloid structures of P rats.
These results suggest that NPY infusion may increase the expression of endogenous NPY in the CeA, which is most likely attributable to an increase in CaMK IV-dependent CREB phosphorylation and this molecular mechanism may be involved in regulating anxiety and alcohol drinking behaviors of P rats.
杏仁中央核（CeA）的神经肽 Y（NPY）系统已被证明参与焦虑和酗酒。在这项研究中，我们使用酒精偏好（P）大鼠作为动物模型，研究了 CeA 中的 NPY 调节焦虑和饮酒行为的分子机制。
酒精偏好大鼠双侧靶向 CeA 进行套管，并注入人工脑脊液（aCSF）或 NPY。通过 2 瓶自由选择范式和明暗箱（LDB）探索测试评估酒精摄入和焦虑样行为。通过金免疫标记程序测定 NPY 及相关信号蛋白水平。通过原位 RT-PCR 测量 NPY 的 mRNA 水平。进行双免疫荧光标记以观察 NPY 和钙/钙调蛋白依赖性蛋白激酶 IV（CaMK IV）的共定位。
我们发现，CeA 中 NPY 的输注产生了焦虑样作用，如 LDB 探索测试所测量的，并且还减少了 P 大鼠的酒精摄入量。CeA 中 NPY 的输注显著增加了 CaMK IV 和磷酸化 cAMP 反应元件结合（pCREB）蛋白的水平，并增加了 NPY 的 mRNA 和蛋白水平，但 CeA 中的 CREB 或蛋白激酶 A（PKA-Calpha）的催化α亚单位的蛋白水平没有变化。我们还观察到，酒精摄入在 P 大鼠的 LDB 测试中产生了焦虑样作用，并且增加了 NPY 的表达以及 pCREB 和 PKA-Calpha 的蛋白水平，而没有调节 CREB 或 CaMK IV 的蛋白水平，无论是在 CeA 还是杏仁核中间核。此外，我们发现 P 大鼠杏仁核结构中 CaMK IV 阳性细胞与 NPY 共定位。
这些结果表明，NPY 的输注可能会增加 CeA 中内源性 NPY 的表达，这很可能归因于 CaMK IV 依赖性 CREB 磷酸化的增加，这种分子机制可能参与调节 P 大鼠的焦虑和饮酒行为。
