Hurley David L, Birch Derin V, Almond M Camille, Estrada Irma J, Phelps Carol J
Department of Cell and Molecular Biology and Neuroscience Program, Tulane University, New Orleans, Louisiana 70118, USA.
Endocrinology. 2003 Nov;144(11):4783-9. doi: 10.1210/en.2003-0753. Epub 2003 Aug 14.
Neuropeptide Y (NPY)-producing neurons in the hypothalamic arcuate nucleus (ARC) have been implicated in GH feedback in several studies in rats. Ames (df/df) and Snell (dw/dw) dwarf mice carry mutations in transcription factors Prop-1 and Pit-1, respectively, that abrogate detectable expression of GH, prolactin, and TSH. The present study was undertaken to determine whether and to what extent hypothalamic NPY neurons are affected by the lifelong absence of pituitary hormone feedback in hypopituitary Ames and Snell dwarf mice. Total ARC NPY mRNA levels were quantified using in situ hybridization, and numbers of ARC NPY-producing cells were quantified using immunocytochemistry. For in situ hybridization, dwarf and normal coronal brain sections were hybridized with 35S-labeled riboprobe complementary to rat NPY cDNA, and then analyzed for total signal intensity from the entire ARC for each animal as well as for mRNA per neuron. NPY-containing perikarya in ARC were counted in sections of colchicine-treated (intracerebroventricular) dwarf and normal mice. Total ARC NPY mRNA was reduced in df/df mice to 33.6% (P < 0.01) of that in normal littermates, and reduced in dw/dw mice to 46.3% (P < 0.05) of normals, but there was no difference in expression per neuron as determined by reduced silver-grain counting. The decrement in dwarf mice of total ARC NPY mRNA without reduction in mRNA per cell suggested a reduction in NPY-containing neuron number, which was the case as shown by immunocytochemistry. NPY neuronal number in adult Ames dwarf mice (1048 +/- 104) was significantly (P < 0.01) reduced to 68% of that in DF/df littermates (1536 +/- 73), and significantly (P < 0.05) reduced in Snell dwarf mice to 63% of normals (1138 +/- 137 vs. 1726 +/- 205). This study represents the first enumeration of NPY-producing neurons in mouse hypothalamus and the first demonstration of lower NPY neuron number in a hypopituitary model. The reduction in total NPY mRNA was greater than that reported in studies of GH-deficient rats, suggesting that NPY expression may be affected by the lifelong absence of prolactin or TSH or both, as well as GH.
下丘脑弓状核（ARC）中产生神经肽Y（NPY）的神经元在多项大鼠研究中被认为与生长激素（GH）反馈有关。艾姆斯（df/df）侏儒小鼠和斯内尔（dw/dw）侏儒小鼠分别在转录因子Prop-1和Pit-1中携带突变，这些突变消除了GH、催乳素和促甲状腺激素（TSH）的可检测表达。本研究旨在确定垂体功能减退的艾姆斯和斯内尔侏儒小鼠中，下丘脑NPY神经元是否以及在何种程度上受到垂体激素反馈终身缺失的影响。使用原位杂交法定量ARC中总的NPY mRNA水平，使用免疫细胞化学法定量ARC中产生NPY的细胞数量。对于原位杂交，将侏儒小鼠和正常小鼠的冠状脑切片与与大鼠NPY cDNA互补 的35S标记核糖探针杂交，然后分析每只动物整个ARC的总信号强度以及每个神经元的mRNA。在秋水仙碱处理（脑室内注射）的侏儒小鼠和正常小鼠的切片中计数ARC中含NPY的核周体。df/df小鼠ARC中总的NPY mRNA减少至正常同窝小鼠的33.6%（P<0.01），dw/dw小鼠减少至正常小鼠的46.3%（P<0.05），但通过减少银粒计数确定，每个神经元的表达没有差异。侏儒小鼠ARC中总的NPY mRNA减少而每个细胞的mRNA没有减少，提示含NPY的神经元数量减少，免疫细胞化学结果也证实了这一点。成年艾姆斯侏儒小鼠中NPY神经元数量（1048±104）显著（P<0.01）减少至DF/df同窝小鼠（1536±73）的68%，斯内尔侏儒小鼠中显著（P<0.05）减少至正常小鼠的63%（1138±13 vs.1726±205）。本研究首次对小鼠下丘脑产生NPY的神经元进行了计数，并首次在垂体功能减退模型中证明NPY神经元数量减少。总的NPY mRNA的减少幅度大于GH缺乏大鼠研究中的报道，提示NPY表达可能受到催乳素或TSH或两者以及GH终身缺失的影响。
