Kamegai J, Unterman T G, Frohman L A, Kineman R D
Department of Medicine, University of Illinois at Chicago, 60612, USA.
Endocrinology. 1998 Aug;139(8):3554-60. doi: 10.1210/endo.139.8.6136.
In this study, the spontaneous dwarf rat (SDR) has been used to examine GHRH production and action in the selective absence of endogenous GH. This dwarf model is unique in that GH is not produced because of a point mutation in the GH gene. However, other pituitary hormones are not obviously compromised. Examination of the hypothalamic pituitary-axis of SDRs revealed that GHRH messenger RNA (mRNA) levels were increased, whereas somatostatin (SS) and neuropeptide Y (NPY) mRNA levels were decreased, compared with age- and sex-matched normal controls, as determined by Northern blot analysis (n = 5 animals/group; P < 0.05). The elevated levels of GHRH mRNA in the SDR hypothalamus were accompanied by a 56% increase in pituitary GHRH receptor (GHRH-R) mRNA, as determined by RT-PCR (P < 0.05). To investigate whether the up-regulation of GHRH-R mRNA resulted in an increase in GHRH-R function, SDR and control pituitary cell cultures were challenged with GHRH (0.001-10 nM; 15 min), and intracellular cAMP concentrations were measured by RIA. Interestingly, SDR pituitary cells were hyperresponsive to 1 and 10 nM GHRH, which induced a rise in intracellular cAMP concentrations 50% greater than that observed in control cultures (n = 3 separate experiments; P < 0.05 and P < 0.01, respectively). Replacement of GH, by osmotic minipump (10 microg/h for 72 h), resulted in the suppression of GHRH mRNA levels (P < 0.01), whereas SS and NPY mRNA levels were increased (P < 0.05), compared with vehicle-treated controls (n = 5 animals/treatment group). Consonant with the fall in hypothalamic GHRH mRNA was a decrease in pituitary GHRH-R mRNA levels. Although replacement of insulin-like growth factor-I (IGF-I), by osmotic pump (5 microg/h for 72 h), resulted in a rise in circulating IGF-I concentrations comparable with that observed after GH replacement, IGF-I treatment was ineffective in modulating GHRH, SS, or NPY mRNA levels. However, IGF-I treatment did reduce pituitary GHRH-R mRNA levels, compared with vehicle-treated controls (P < 0.05). These results further validate the role of GH as a negative regulator of hypothalamic GHRH expression, and they suggest that SS and NPY act as intermediaries in GH-induced suppression of hypothalamic GHRH synthesis. These data also demonstrate that increases in circulating IGF-I are not responsible for changes in hypothalamic function observed after GH treatment. Finally, this report establishes modulation of GHRH-R synthesis as a component of GH autofeedback regulation.
在本研究中，自发性侏儒大鼠（SDR）被用于在选择性缺乏内源性生长激素（GH）的情况下检测生长激素释放激素（GHRH）的产生及作用。该侏儒模型的独特之处在于，由于GH基因中的一个点突变，导致无法产生GH。然而，其他垂体激素并未受到明显影响。通过Northern印迹分析（每组5只动物；P < 0.05）发现，与年龄和性别匹配的正常对照相比，SDR下丘脑-垂体轴中GHRH信使核糖核酸（mRNA）水平升高，而生长抑素（SS）和神经肽Y（NPY）的mRNA水平降低。通过逆转录聚合酶链反应（RT-PCR）测定，SDR下丘脑GHRH mRNA水平的升高伴随着垂体GHRH受体（GHRH-R）mRNA增加56%（P < 0.05）。为了研究GHRH-R mRNA的上调是否导致GHRH-R功能增加，用GHRH（0.001 - 10 nM；15分钟）刺激SDR和对照垂体细胞培养物，并用放射免疫分析（RIA）测量细胞内环磷酸腺苷（cAMP）浓度。有趣的是，SDR垂体细胞对1和10 nM GHRH反应过度，诱导细胞内cAMP浓度升高，比对照培养物中观察到的高50%（3个独立实验；分别为P < 0.05和P < 0.01）。通过渗透微型泵（10微克/小时，持续72小时）替代GH，与给予赋形剂处理的对照相比，导致GHRH mRNA水平受到抑制（P < 0.01），而SS和NPY mRNA水平升高（P < 0.05）（每组5只动物）。与下丘脑GHRH mRNA下降一致的是垂体GHRH-R mRNA水平降低。虽然通过渗透泵（5微克/小时，持续72小时）替代胰岛素样生长因子-I（IGF-I）导致循环中IGF-I浓度升高，与GH替代后观察到的相当，但IGF-I处理在调节GHRH、SS或NPY mRNA水平方面无效。然而，与给予赋形剂处理的对照相比，IGF-I处理确实降低了垂体GHRH-R mRNA水平（P < 0.05）。这些结果进一步证实了GH作为下丘脑GHRH表达负调节因子的作用，并表明SS和NPY在GH诱导的下丘脑GHRH合成抑制中起中介作用。这些数据还表明，循环中IGF-I的增加与GH治疗后观察到的下丘脑功能变化无关。最后，本报告确定GHRH-R合成的调节是GH自身反馈调节的一个组成部分。
