Abuhagr Ali M, Blindert Jennifer L, Nimitkul Sukkrit, Zander Ian A, Labere Stefan M, Chang Sharon A, Maclea Kyle S, Chang Ernest S, Mykles Donald L
Department of Biology, Colorado State University, Fort Collins, CO 80523, USA.
J Exp Biol. 2014 Mar 1;217(Pt 5):796-808. doi: 10.1242/jeb.093385. Epub 2013 Nov 6.
In decapod crustaceans, regulation of molting is controlled by the X-organ/sinus gland complex in the eyestalks. The complex secretes molt-inhibiting hormone (MIH), which suppresses production of ecdysteroids by the Y-organ (YO). MIH signaling involves nitric oxide and cGMP in the YO, which expresses nitric oxide synthase (NOS) and NO-sensitive guanylyl cyclase (GC-I). Molting can generally be induced by eyestalk ablation (ESA), which removes the primary source of MIH, or by multiple leg autotomy (MLA). In our work on Carcinus maenas, however, ESA has limited effects on hemolymph ecdysteroid titers and animals remain in intermolt at 7 days post-ESA, suggesting that adults are refractory to molt induction techniques. Consequently, the effects of ESA and MLA on molting and YO gene expression in C. maenas green and red color morphotypes were determined at intermediate (16 and 24 days) and long-term (~90 days) intervals. In intermediate-interval experiments, ESA of intermolt animals caused transient twofold to fourfold increases in hemolymph ecdysteroid titers during the first 2 weeks. In intermolt animals, long-term ESA increased hemolymph ecdysteroid titers fourfold to fivefold by 28 days post treatment, but there was no late premolt peak (>400 pg μl(-1)) characteristic of late premolt animals and animals did not molt by 90 days post-ESA. There was no effect of ESA or MLA on the expression of Cm-elongation factor 2 (EF2), Cm-NOS, the beta subunit of GC-I (Cm-GC-Iβ), a membrane receptor GC (Cm-GC-II) and a soluble NO-insensitive GC (Cm-GC-III) in green morphs. Red morphs were affected by prolonged ESA and MLA treatments, as indicated by large decreases in Cm-EF2, Cm-GC-II and Cm-GC-III mRNA levels. ESA accelerated the transition of green morphs to the red phenotype in intermolt animals. ESA delayed molting in premolt green morphs, whereas intact and MLA animals molted by 30 days post treatment. There were significant effects on YO gene expression in intact animals: Cm-GC-Iβ mRNA increased during premolt and Cm-GC-III mRNA decreased during premolt and increased during postmolt. Cm-MIH transcripts were detected in eyestalk ganglia, the brain and the thoracic ganglion from green intermolt animals, suggesing that MIH in the brain and thoracic ganglion prevents molt induction in green ESA animals.
在十足目甲壳动物中，蜕皮的调节由眼柄中的X器官/窦腺复合体控制。该复合体分泌蜕皮抑制激素（MIH），它抑制Y器官（YO）产生蜕皮类固醇。MIH信号传导涉及YO中的一氧化氮和环鸟苷酸，YO表达一氧化氮合酶（NOS）和对NO敏感的鸟苷酸环化酶（GC-I）。蜕皮通常可通过眼柄切除（ESA）诱导，这会去除MIH的主要来源，或者通过多条腿自切（MLA）诱导。然而，在我们对海蟹的研究中，ESA对血淋巴蜕皮类固醇滴度的影响有限，动物在ESA后7天仍处于蜕皮间期，这表明成年动物对蜕皮诱导技术不敏感。因此，在中间（16天和24天）和长期（约90天）间隔下，测定了ESA和MLA对海蟹绿色和红色形态型蜕皮及YO基因表达的影响。在中间间隔实验中，处于蜕皮间期动物的ESA在最初2周内使血淋巴蜕皮类固醇滴度短暂升高了两倍至四倍。在处于蜕皮间期的动物中，长期ESA在处理后28天使血淋巴蜕皮类固醇滴度升高了四倍至五倍，但没有出现蜕皮前晚期动物特有的晚期蜕皮前峰值（>400 pg μl(-1)），并且动物在ESA后90天没有蜕皮。ESA或MLA对绿色形态型中Cm-延伸因子2（EF2）、Cm-NOS、GC-I的β亚基（Cm-GC-Iβ）、膜受体GC（Cm-GC-II）和可溶性对NO不敏感的GC（Cm-GC-III）的表达没有影响。长期的ESA和MLA处理影响了红色形态型，表现为Cm-EF2、Cm-GC-II和Cm-GC-III mRNA水平大幅下降。ESA加速了处于蜕皮间期动物中绿色形态型向红色表型的转变。ESA延迟了处于蜕皮前阶段的绿色形态型的蜕皮，而完整动物和MLA处理的动物在处理后30天蜕皮。完整动物的YO基因表达有显著变化：Cm-GC-Iβ mRNA在蜕皮前增加，Cm-GC-III mRNA在蜕皮前下降，在蜕皮后增加。在绿色蜕皮间期动物的眼柄神经节、脑和胸神经节中检测到了Cm-MIH转录本，这表明脑和胸神经节中的MIH阻止了绿色ESA动物的蜕皮诱导。
