Al Kahtane A, Chaiseha Y, El Halawani M
Department of Animal Science, University of Minnesota, St Paul, Minnesota 55108, USA.
J Mol Endocrinol. 2003 Aug;31(1):185-96. doi: 10.1677/jme.0.0310185.
It is well documented that prolactin (PRL) release and PRL gene expression in birds are controlled by the tonic stimulation of hypothalamic vasoactive intestinal peptide (VIP). However, there is good evidence that dopamine (DA) exerts both stimulatory (at the hypothalamic level) and inhibitory (at the pituitary level) effects on PRL secretion. The interactions between VIP and DA in the regulation of PRL gene transcription are not known. This study was designed to examine the effects of a D(2) DA receptor agonist (D(2)AG; R(-)-propylnorapomorphine HCl) on basal and VIP-stimulated PRL gene transcription rate, PRL mRNA steady-state levels, PRL mRNA stability and PRL release from cultured turkey anterior pituitary cells. The D(2)AG (10(-)(10) M) completely inhibited the stimulatory effect of VIP (10(-)(7) M) upon nascent PRL mRNA as determined utilizing a nuclear run-on transcription assay. To examine further the effect of the D(2)AG on PRL mRNA post-transcriptional events, anterior pituitary cells were treated with different concentrations of D(2)AG (10(-)(12)-10(-)(4) M). Semi-quantitative RT-PCR and RIA were performed to determine the levels of PRL mRNA and PRL content in the medium respectively. The results show that D(2)AG inhibited VIP-stimulated PRL mRNA steady-state levels as well as basal and VIP-stimulated PRL release, effects which were diminished by the D(2) DA receptor antagonist, S(-)-eticlopride HCl (10(-)(10) M). Actinomycin D (5 microg/ml), an inhibitor of mRNA synthesis, was used to assess the effect of D(2)AG on PRL mRNA stability in response to VIP. The stimulatory effect of VIP on PRL mRNA stability was completely negated by the D(2)AG (from a half-life of 53.0+/-2.3 h in VIP-treated cells to 25.5+/-1.6 h in D(2)AG+VIP-treated cells, P<or=0.05). These results support the hypothesis that VIP and DA play a major role in the regulation of PRL gene expression in avian species, at both the transcriptional and post-transcriptional levels. In addition, these findings suggest that the DAergic system inhibits PRL release and synthesis by antagonizing VIP at the pituitary level via D(2) DA receptors.
有充分的文献记载，鸟类中催乳素（PRL）的释放和PRL基因表达受下丘脑血管活性肠肽（VIP）的紧张性刺激控制。然而，有充分证据表明多巴胺（DA）对PRL分泌具有刺激作用（在下丘脑水平）和抑制作用（在垂体水平）。VIP和DA在PRL基因转录调节中的相互作用尚不清楚。本研究旨在检测D（2）DA受体激动剂（D（2）AG；R（-）-丙基去甲阿扑吗啡盐酸盐）对基础和VIP刺激的PRL基因转录速率、PRL mRNA稳态水平、PRL mRNA稳定性以及培养的火鸡垂体前叶细胞PRL释放的影响。利用核转录分析确定，D（2）AG（10（-）（10）M）完全抑制了VIP（10（-）（7）M）对新生PRL mRNA的刺激作用。为了进一步检测D（2）AG对PRL mRNA转录后事件的影响，用不同浓度的D（2）AG（10（-）（12）-10（-）（4）M）处理垂体前叶细胞。分别进行半定量RT-PCR和RIA以确定培养基中PRL mRNA水平和PRL含量。结果显示，D（2）AG抑制了VIP刺激的PRL mRNA稳态水平以及基础和VIP刺激的PRL释放，D（2）DA受体拮抗剂S（-）-依托必利盐酸盐（10（-）（10）M）可减弱这些作用。用放线菌素D（5μg/ml）（一种mRNA合成抑制剂）来评估D（2）AG对VIP刺激下PRL mRNA稳定性的影响。VIP对PRL mRNA稳定性的刺激作用被D（2）AG完全抵消（从VIP处理细胞中的半衰期53.0±2.3小时降至D（2）AG+VIP处理细胞中的25.5±1.6小时，P≤0.05）。这些结果支持了以下假说：VIP和DA在鸟类PRL基因表达的调节中在转录和转录后水平均起主要作用。此外，这些发现表明多巴胺能系统通过D（2）DA受体在垂体水平拮抗VIP来抑制PRL释放和合成。
