Study of neural networks and control substances for insect photoperiodism

本研究从形态学上考察了土原蝇（protophoria terraenovae）调节生殖滞育的光周期机制中涉及的神经网络和控制物质。通过关注小型PDF神经元（生物钟神经元）与侧部神经元（滞育控制神经元）和对生殖重要的脑间部神经元的神经连接，我们得到了以下结果。光周期病神经网络形态学分析：在5组脑细胞中发现周期免疫反应性。5组中2组包括小型PDF神经元均采用抗PERIOD和抗PDF（色素分散因子）抗血清双标记。手术切除小型PDF神经元的躯体区域破坏了光周期，表明昼夜节律钟神经元参与了光周期。免疫细胞化学和回填将侧部神经元分为2种类型和6种类型。在脑间部神经元中发现FMRFamide-免疫反应性，在侧部神经元中发现FMRFamide-、PDF-、胆固醇收缩素-、corazonin免疫反应性。光周期神经网络中的控制物质：直接MALDI-TOF质谱、ESI-Q-Tof质谱和MS/MS提示，肌抑制素和SIFamide存在于脑间部，corazonin和corazonin相关肽存在于侧部。实时荧光定量PCR分析显示，在光周期条件下，两个生物钟基因的表达每日波动，但长日照和短日照条件下没有差异。必须确定昼夜节律钟基因是如何参与光周期的。

This study morphologically examined neural networks and control substances involved in photoperiodic mechanisms regulating reproductive diapause in the blow fly Protophormia terraenovae. We obtained following results by focusing on neural connections from small type PDF neurons (circadian clock neurons) to pars lateralis neurons (diapause controlling neurons) and pars intercerebralis neurons important for reproduction.Morphological analysis of neural networks for photoperiodism: PERIOD-immunoreactivity was found in 5 groups of cells in the brain. Among the 5 groups two including small type PDF neurons were double-labeled with anti- PERIOD and anti-PDF (pigment-dispersing factor) antisera. Surgical ablation of the somata region of small type PDF neurons disrupted photoperiodism, suggesting that circadian clock neurons were involved in photoperiodism. Immunocytochemistry and backfills classified two types in the pars lateralis and 6 types of pars lateralis neurons. In the pars intercerebralis neurons, FMRFamide-immunoreacrtivity and in the pars lateralis FMRFamide-, PDF-, cholescystokinin-, corazonin-immunoreacrtivity were found.Control substances in neural networks for photoperiodism: Direct MALDI-TOF MS, and ESI-Q-Tof MS and MS/MS suggested that myosuppressin and SIFamide were present in the pars intercerebralis, corazonin and a corazonin-related peptide in the pars lateralis. Real-time PCR analysis revealed daily fluctuations in the expression of two circadian clock genes period and timeless under photoperiodic conditions, but no differences between long-day and short-day conditions were found. It has to be determined how the circadian clock genes are involved in photoperiodism.