Datta S, Siwek D F, Stack E C
Laboratory of Sleep and Cognitive Neuroscience, Boston University School of Medicine, 85 East Newton Street, Suite M-902, Boston, MA 02118, USA.
Neuroscience. 2009 Sep 29;163(1):397-414. doi: 10.1016/j.neuroscience.2009.06.035. Epub 2009 Jun 18.
Recent studies have shown that in the pedunculopontine tegmental nucleus (PPT), increased neuronal activity and kainate receptor-mediated activation of intracellular protein kinase A (PKA) are important physiological and molecular steps for the generation of rapid eye movement (REM) sleep. In the present study performed on rats, phosphorylated cyclic AMP response element-binding protein (pCREB) immunostaining was used as a marker for increased intracellular PKA activation and as a reflection of increased neuronal activity. To identify whether activated cells were either cholinergic or noncholinergic, the PPT and laterodorsal tegmental nucleus (LDT) cells were immunostained for choline acetyltransferase (ChAT) in combination with pCREB or c-Fos. The results demonstrated that during high rapid eye movement sleep (HR, approximately 27%), significantly higher numbers of cells expressed pCREB and c-Fos in the PPT, of which 95% of pCREB-expressing cells were ChAT-positive. With HR, the numbers of pCREB-positive cells were also significantly higher in the medial pontine reticular formation (mPRF), pontine reticular nucleus oral (PnO), and dorsal subcoeruleus nucleus (SubCD) but very few in the locus coeruleus (LC) and dorsal raphe nucleus (DRN). Conversely, with low rapid eye movement sleep (LR, approximately 2%), the numbers of pCREB expressing cells were very few in the PPT, mPRF, PnO, and SubCD but significantly higher in the LC and DRN. The results of regression analyses revealed significant positive relationships between the total percentages of REM sleep and numbers of ChAT+/pCREB+ (Rsqr=0.98) cells in the PPT and pCREB+ cells in the mPRF (Rsqr=0.88), PnO (Rsqr=0.87), and SubCD (Rsqr=0.84); whereas significantly negative relationships were associated with the pCREB+ cells in the LC (Rsqr=0.70) and DRN (Rsqr=0.60). These results provide evidence supporting the hypothesis that during REM sleep, the PPT cholinergic neurons are active, whereas the LC and DRN neurons are inactive. More importantly, the regression analysis indicated that pCREB activation in approximately 98% of PPT cholinergic neurons, was caused by REM sleep. Moreover the results indicate that during REM sleep, PPT intracellular PKA activation and a transcriptional cascade involving pCREB occur exclusively in the cholinergic neurons.
近期研究表明，在脚桥被盖核（PPT）中，神经元活动增加以及红藻氨酸受体介导的细胞内蛋白激酶A（PKA）激活是快速眼动（REM）睡眠产生的重要生理和分子步骤。在本项对大鼠进行的研究中，磷酸化环磷酸腺苷反应元件结合蛋白（pCREB）免疫染色被用作细胞内PKA激活增加的标志物以及神经元活动增加的反映。为了确定被激活的细胞是胆碱能还是非胆碱能的，对PPT和外侧背盖核（LDT）细胞进行胆碱乙酰转移酶（ChAT）与pCREB或c-Fos的联合免疫染色。结果表明，在高快速眼动睡眠期间（HR，约27%），PPT中表达pCREB和c-Fos的细胞数量显著更多，其中95%表达pCREB的细胞是ChAT阳性。在HR时，脑桥内侧网状结构（mPRF）、脑桥网状核嘴侧部（PnO）和蓝斑下背核（SubCD）中pCREB阳性细胞数量也显著更高，但在蓝斑（LC）和中缝背核（DRN）中很少。相反，在低快速眼动睡眠期间（LR，约2%），PPT、mPRF、PnO和SubCD中表达pCREB的细胞数量很少，但在LC和DRN中显著更高。回归分析结果显示，REM睡眠的总百分比与PPT中ChAT+/pCREB+细胞数量（Rsqr = 0.98）以及mPRF（Rsqr = 0.88）、PnO（Rsqr = 0.87）和SubCD（Rsqr = 0.84）中pCREB+细胞数量之间存在显著正相关；；；而与LC（Rsqr = 0.70）和DRN（Rsqr = 0.60）中的pCREB+细胞数量存在显著负相关。这些结果提供了证据支持以下假设：在REM睡眠期间，PPT胆碱能神经元活跃，而LC和DRN神经元不活跃。更重要的是，回归分析表明，约98%的PPT胆碱能神经元中的pCREB激活是由REM睡眠引起的。此外，结果表明在REM睡眠期间，PPT细胞内PKA激活以及涉及pCREB的转录级联反应仅发生在胆碱能神经元中。
