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【Biology of Reproduction】GNRH脉冲可调节大鼠脑垂体
http://www.biolreprod.org/cgi/content/abstract/biolreprod.108.070987v1?papetoc
Regulation of Intracellular Signaling Cascades by GNRH Pulse Frequency in the Rat Pituitary: Roles for CaMK II, ERK, and JNK Activation
GNRH脉冲可调节大鼠脑垂体细胞内信号转导的级联反应：CaMK II、ERK和JNK激活的作用
Laura L. Burger *, Daniel J. Haisenleder , Kevin W. Aylor , and John C. Marshall
* To whom correspondence should be addressed. E-mail: lburger@virginia.edu .
Abstract
Pulsatile GnRH (GNRH) differentially regulates LH and FSH subunit genes, with faster frequencies favoring Lhb transcription and slower favoring Fshb. Various intracellular pathways mediate the effects of GNRH, including CaMK II (CAMK2), ERK and JNK. We examined if activation of these pathways are regulated by GNRH pulse frequency in vivo. GNRH deficient rats received GNRH pulses (25ng iv every 30 or 240 min for 8 h, vehicle to controls). Pituitaries were collected 5 min after the last pulse, bisected and one half processed for RNA (to measure beta-subunit primary transcripts [PT]) and the other for protein. Phosphorylated (phospho) CAMK2, ERK (MAPK1/3, also known as p42 ERK2 and p44 ERK1 respectively) and JNK (MAPK8/9, also known as p46 JNK1 and p54 JNK2 respectively) were determined by western blotting. 30 min pulses maximally stimulated Lhb PT (8-fold), whereas 240 min was optimal for Fshb PT (3-fold increase). Both GNRH pulse frequencies increased phospho-CAMK2 4-fold. Activation of MAPK1/3 was stimulated by both 30 and 240 min pulses, but phosphorylation of MAPK3 was significantly greater following slower GNRH pulses (240 min = 4-fold, 30 min = 2-fold). MAPK8/9 activation was unchanged by pulsatile GNRH in this paradigm, but as previous results showed that GNRH induced activation of MAPK8/9 is delayed, 5 min post GNRH may not be optimal to observe MAPK8/9 activation. These data show that CAMK2 is activated by GNRH but not in a frequency dependant manner, whereas MAPK3 is maximally stimulated by slow frequency GNRH pulses. Thus, the ERK response to slow pulse frequency is part of the mechanisms mediating Fhb transcriptional responses to GNRH.
Key words: Follicle-stimulating hormone • Follistatin • Gonadotropin-releasing hormone • Luteinizing hormone • Signal transduction Pulsatile GnRH (GNRH) differentially regulates LH and FSH subunit genes, with faster frequencies favoring Lhb transcription and slower favoring Fshb.Various intracellular pathways mediate the effects of GNRH, including CaMK II (CAMK2), ERK and JNK.
GnRH脉冲对于LH和FSH亚基的基因表达的调控存在差异，快脉冲倾向激活LHβ转录，而较慢速的脉冲则激活FSHβ的转录。细胞内多种信号通路都可以传递GnRH信号，包括CaMK II(CAMK2)、ERK和JNK。
We examined if activation of these pathways are regulated by GNRH pulse frequency in vivo. GNRH deficient rats received GNRH pulses (25ng iv every 30 or 240 min for 8 h, vehicle to controls). Pituitaries were collected 5 min after the last pulse, bisected and one half processed for RNA (to measure beta-subunit primary transcripts [PT]) and the other for protein.
我们观察了是否体内这些信号通路的激活是由GnRH脉冲调控的。我们人为给予缺乏GnRH脉冲的大鼠静脉注射GnRH （8小时内每隔30分钟或者240分钟静脉注射GnRH25ng，设立载体组为对照组）。在最后注射后的5分钟收集脑垂体，将脑垂体一分为二，一半用于RNA实验（检测β亚单位一级转录水平），另一半用于蛋白质分析。
Phosphorylated (phospho) CAMK2, ERK (MAPK1/3, also known as p42 ERK2 and p44 ERK1 respectively) and JNK (MAPK8/9, also known as p46 JNK1 and p54 JNK2 respectively) were determined by western blotting.
应用免疫印迹测定磷酸化CAMK2，（ MAPK1/3，也分别称为p42 ERK2和p44 ERK1）和JNK （ MAPK8/9，也分别称为p46 JNK1和p54 JNK2）。
30 min pulses maximally stimulated Lhb PT (8-fold), whereas 240 min was optimal for Fshb PT (3-fold increase). Both GNRH pulse frequencies increased phospho-CAMK2 4-fold. Activation of MAPK1/3 was stimulated by both 30 and 240 min pulses, but phosphorylation of MAPK3 was significantly greater following slower GNRH pulses (240 min = 4-fold, 30 min = 2-fold). MAPK8/9 activation was unchanged by pulsatile GNRH in this paradigm, but as previous results showed that GNRH induced activation of MAPK8/9 is delayed, 5 min post GNRH may not be optimal to observe MAPK8/9 activation.
30分钟 GNRH脉冲，可最大限度的激发LHβ初级转录物，使之增加8倍，而240分钟间隔最适用于激发FSHβ初级转录物，使之增加3倍。这两个脉冲频率都可以使磷酸化CAMK2 增加4倍，都可以激活MAPK1/3，但慢频率显著增加磷酸化MAPK3（ 240分钟= 4倍， 30分钟= 2倍）。 MAPK8/9的激活不变，但正如前面的结果所示，GNRH脉冲激活MAPK8/9有延迟效应，GNRH脉冲后5分钟可能不是观察MAPK8/9激活的最佳时机。
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作者:admin@医学,生命科学 2011-08-22 06:36
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