Kimura M, Yu W H, Rettori V, McCann S M
Institute for Medical and Dental Engineering, Tokyo Medical and Dental University, Japan.
Neuroimmunomodulation. 1997 Sep-Dec;4(5-6):237-43. doi: 10.1159/000097342.
Several cytokines produced by immune cells act within the hypothalamus and/or on the pituitary to produce the pattern of pituitary hormone secretion that characterizes infection. Granulocyte-macrophage colony stimulating factor (GMCSF) was first described as a hematopoietic cytokine; however, its synthesis is also stimulated during infection, and it has been found in glia in the brain. Previous research indicates that interleukin-1 inhibits release of luteinizing hormone-releasing hormone (LHRH) both in vivo and in vitro. In the present study, we determined that GMCSF inhibited the release of LHRH in vitro and evaluated the mechanisms involved. After a 1-hour preincubation in Krebs-Ringer bicarbonate glucose buffer (KRB), medial basal hypothalamic explants were incubated in KRB together with recombinant murine GMCSF for 0.5 h in a Dubnoff metabolic shaker (50 cycles/min) in an atmosphere of 95% O2/5% CO2. LHRH release into the media was determined by radioimmunoassay. At concentrations of 10(-12) and 10(-11) M, GMCSF significantly inhibited LHRH release. There was a U-shaped dose-response curve and LHRH release was not inhibited at lower or higher cytokine concentrations. The inhibition was specific since it was completely blocked by GMCSF antiserum. Since sodium nitroprusside (NP; 300 microM), a releaser of nitric oxide (NO), stimulates LHRH, presumably by acting within the LHRH neurons, we examined the effect of GMCSF (10(-11) M) on NP-induced LHRH release. It completely suppressed NP-induced release of LHRH. Bicuculline (10(-5) M), a gamma-aminobutyric acid (GABA) receptor antagonist, partially reversed the inhibitory effects of GMCSF on LHRH release. This dose completely reversed the suppression of LHRH release induced by GABA. The present results indicate that the inhibitory effects of GMCSF on LHRH release are partially caused by blockade of NO-induced LHRH release by its activation of GMCSF receptors on GABAergic neurons. The stimulated release of GABA acts on the GABA-a receptors on the LHRH terminals to inhibit their response to NO. At the end of the experiment, NO synthase (NOS) activity was measured in the tissue homogenate by the citrulline method. NOS activity was highly significantly reduced by GMCSF (10(-11) M) indicating that part of its suppressive action on LHRH release is mediated by reduction in NOS activity in the medial basal hypothalamus.
免疫细胞产生的几种细胞因子在下丘脑内和/或作用于垂体，以产生表征感染的垂体激素分泌模式。粒细胞-巨噬细胞集落刺激因子（GMCSF）最初被描述为一种造血细胞因子；然而，其合成在感染期间也会受到刺激，并且已在大脑中的神经胶质细胞中发现。先前的研究表明，白细胞介素-1在体内和体外均抑制促黄体生成素释放激素（LHRH）的释放。在本研究中，我们确定GMCSF在体外抑制LHRH的释放，并评估其中涉及的机制。在Krebs-Ringer碳酸氢盐葡萄糖缓冲液（KRB）中预孵育1小时后，将内侧基底下丘脑外植体在KRB中与重组鼠GMCSF一起在Dubnoff代谢摇床（50次/分钟）中于95% O2/5% CO2气氛中孵育0.5小时。通过放射免疫测定法测定培养基中LHRH的释放。在浓度为10^(-12)和10^(-11) M时，GMCSF显著抑制LHRH的释放。存在一条U形剂量反应曲线，在较低或较高的细胞因子浓度下LHRH的释放未受到抑制。这种抑制是特异性的，因为它被GMCSF抗血清完全阻断。由于硝普钠（NP；300 microM），一种一氧化氮（NO）释放剂，大概通过在LHRH神经元内起作用来刺激LHRH，我们研究了GMCSF（10^(-11) M）对NP诱导的LHRH释放的影响。它完全抑制了NP诱导的LHRH释放。荷包牡丹碱（10^(-5) M），一种γ-氨基丁酸（GABA）受体拮抗剂，部分逆转了GMCSF对LHRH释放的抑制作用。该剂量完全逆转了GABA诱导的LHRH释放的抑制。目前的结果表明，GMCSF对LHRH释放的抑制作用部分是由于其激活GABA能神经元上的GMCSF受体阻断了NO诱导的LHRH释放。受刺激释放的GABA作用于LHRH终末上的GABA-A受体，以抑制它们对NO的反应。在实验结束时，通过瓜氨酸法在组织匀浆中测量一氧化氮合酶（NOS）活性。GMCSF（10^(-11) M）使NOS活性高度显著降低，表明其对LHRH释放的部分抑制作用是由内侧基底下丘脑NOS活性的降低介导的。
