Haisenleder D J, Ortolano G A, Yasin M, Dalkin A C, Marshall J C
Department of Internal Medicine, University of Virginia Health Sciences Center, Charlottesville 22908.
Endocrinology. 1993 Mar;132(3):1292-6. doi: 10.1210/endo.132.3.7679975.
The present study examined the effect of GnRH pulse amplitude on alpha, LH beta and FSH beta mRNAs using an in vitro perfusion model. Pituitaries from 30-day-old female rats were dissociated and the cells plated for 48 h to allow attachment to collagen-coated microcarrier beads. The beads were loaded into perifusion chambers, preperifused for 1 h, and then given GnRH pulses (17.5-175 pg/ml) every 30 min for 24 h. Perifusate LH was measured after 2 h and 22 h of perifusion and alpha LH and FSH beta messenger RNAs (mRNAs) were determined by hybridization to complementary DNA (cDNA) probes. All doses of GnRH acutely stimulated LH release, and responses were similar after 2 h and 22 h. LH release increased as a function of GnRH pulse dose with maximal increases seen following 70 pg/ml pulses. alpha mRNA levels (control = 0.73 +/- 0.1 fmol cDNA bound/100 micrograms pituitary DNA) were increased 30% and 40% after 24 h of 35 and 70 pg/ml pulses, respectively (P < 0.05 vs. media controls). LH beta mRNA concentrations (control = 0.44 +/- 0.08 fmol cDNA bound) were only elevated after 35 pg/ml GnRH pulses (36% increase). FSH beta mRNA showed the largest responses to GnRH pulses, increasing by 45% and 84% after 35 and 70 pg pulses, respectively (control = 0.14 +/- 0.02 fmol bound). The highest GnRH pulse dose (175 pg/ml) was ineffective in stimulating an increase in FSH beta mRNA levels. These results show that all three gonadotropin subunit mRNA concentrations increase after 24 h of GnRH pulses, but the pattern of individual subunit mRNA responses was dependent upon the amplitude of the GnRH pulse stimulus. These data support earlier results in vivo, in that the subunit responses to GnRH pulse dose were similar. Thus, alterations in the amplitude of pulsatile GnRH secretion from the median eminence may be one mechanism by which the expression of gonadotropin subunit genes are regulated.
本研究使用体外灌注模型，检测了促性腺激素释放激素（GnRH）脉冲幅度对α、促黄体生成素β（LHβ）和促卵泡生成素β（FSHβ）信使核糖核酸（mRNA）的影响。将30日龄雌性大鼠的垂体进行解离，细胞接种48小时，使其附着于胶原包被的微载体珠上。将这些微载体珠装入灌注室，预灌注1小时，然后每30分钟给予GnRH脉冲（17.5 - 175 pg/ml），持续24小时。在灌注2小时和22小时后测量灌注液中的LH，并通过与互补DNA（cDNA）探针杂交来测定α、LHβ和FSHβ信使核糖核酸（mRNA）。所有剂量的GnRH均能急性刺激LH释放，且在2小时和22小时后的反应相似。LH释放量随GnRH脉冲剂量增加而增加，在70 pg/ml脉冲后达到最大增加量。α mRNA水平（对照组 = 0.73 ± 0.1 fmol cDNA结合量/100μg垂体DNA）在35 pg/ml和70 pg/ml脉冲作用24小时后分别增加了30%和40%（与培养基对照组相比，P < 0.05）。LHβ mRNA浓度（对照组 = 0.44 ± 0.08 fmol cDNA结合量）仅在35 pg/ml GnRH脉冲后升高（增加36%）。FSHβ mRNA对GnRH脉冲反应最大，在35 pg和70 pg脉冲后分别增加45%和84%（对照组 = 0.14 ± 0.02 fmol结合量）。最高的GnRH脉冲剂量（175 pg/ml）未能有效刺激FSHβ mRNA水平升高。这些结果表明，在GnRH脉冲作用24小时后，所有三种促性腺激素亚基mRNA浓度均增加，但各个亚基mRNA反应模式取决于GnRH脉冲刺激的幅度。这些数据支持了早期的体内研究结果，即亚基对GnRH脉冲剂量的反应相似。因此，来自正中隆起的GnRH脉冲分泌幅度的改变可能是调节促性腺激素亚基基因表达的一种机制。
