Elsevier BVActa Medica Okayama2589-00422982026Brain circadian clock neurons drive fitness advantages in Drosophila117125ENSaeAikawaGraduate School of Environmental, Life, Natural Science and Technology, Okayama UniversityShoichiroTamuraGraduate School of Environmental, Life, Natural Science and Technology, Okayama UniversityMakikoMimuraGraduate School of Environmental, Life, Natural Science and Technology, Okayama UniversityTaishiYoshiiGraduate School of Environmental, Life, Natural Science and Technology, Okayama UniversityThe adaptive significance of circadian clocks is widely assumed due to their ubiquity; yet, direct empirical evidence remains scarce. Evaluating these benefits is often confounded by pleiotropic effects in conventional circadian null mutants. To address this, we selectively altered the circadian period exclusively within brain clock neurons in Drosophila melanogaster. Multi-generational competition assays revealed that flies with aberrant rhythms exhibit a significant fitness disadvantage under standard light-dark (LD 12:12) cycles. This disadvantage was abolished under constant light, confirming that the selection pressure is specifically mediated by the circadian clock. Furthermore, paternity assays conducted under LD 12:12 indicated that the timing of brain clock neurons influences male reproductive success, providing a potential mechanistic link between clock-controlled behavior and fitness. Intriguingly, we found that these fitness costs are highly photoperiod-dependent. Under short-day conditions (LD 8:16), the short-period strain (dbtS) maintained a significantly higher overall frequency than the long-period strain (dbtL). Our behavioral observations suggest that this difference may be associated with the quality of activity rhythms; specifically, dbtS lacked defined morning peaks and showed suppressed nocturnal activity, potentially narrowing its window for reproductive interactions compared to dbtL. These findings illustrate that the circadian system does not merely track a 24-h cycle but functions as a flexible feature that enables flies to cope with changing day lengths by aligning their mating behavior with the most favorable time of day.No potential conflict of interest relevant to this article was reported.Springer Science and Business Media LLCActa Medica Okayama2056-306X1212026Knockout analysis of period and timeless and EGFP-based visualization of per-expressing clock cells in the cricket circadian clock12ENKenjiTomiokaGraduate School of Natural Science and Technology, Okayama UniversityShintaroInoueBio-Innovation Research Center, Tokushima UniversityTaroMitoBio-Innovation Research Center, Tokushima UniversityYoshiyukiMoriyamaDepartment of Natural Sciences, Kawasaki Medical SchoolTaishiYoshiiGraduate School of Environmental, Life, Natural Science and Technology, Okayama UniversityIn the present study, we generated crickets with knockout of either period (per) or timeless (tim) gene by CRISPR/Cas9-based genome editing. We also identified a naturally occurring per- mutant lacking a large coding region including PAS domains. To examine possible synergistic effects, a per- and timKO double mutant was generated by applying genome editing to the per- crickets. Under constant darkness (DD), timKO crickets exhibited a locomotor rhythm with a free-running period of 23.06 } 0.20 h (mean } SD), which was significantly shorter than that of the parental strain (23.78 } 0.12 h). By contrast, perKO and per- crickets showed basically similar phenotype of locomotor rhythm: they exhibited an arrhythmic pattern during the first two to three weeks after transfer to DD but subsequently showed a complex rhythmic pattern with one or multiple components with significantly longer free-running periods (33.35 } 10.72 h). In the per-;timKO double mutants, approximately 60% of individuals became arrhythmic, while the remaining 40% exhibited complex rhythms with extremely longer free-running periods (37.0 } 9.17 h) under DD. These results suggest the existence of an underlying oscillatory mechanism that is responsible for regulating locomotor rhythms independently of the canonical per/tim feedback loop. Furthermore, we generated a reporter line on a per| background by knocking egfp into exon 1 of the per gene, allowing egfp expression to report per transcription. EGFP expression was detected in three distinct clusters of cells within the optic lobe: two located along the dorsal and ventral boundaries between the lamina and medulla neuropils, and one situated near the accessory medulla. These findings raise the possibility that these form part of the circadian clock network that governs circadian locomotor rhythms.No potential conflict of interest relevant to this article was reported.Public Library of Science (PLoS)Acta Medica Okayama1553-74042222026Octopamine signaling from clock neurons plays dual roles in Drosophila long-term memorye1012045ENYutoKurataDepartment of Biological Sciences, Tokyo Metropolitan UniversityTaishiYoshiiGraduate School of Environmental, Life, Natural Science and Technology, Okayama UniversityTakaomiSakaiDepartment of Biological Sciences, Tokyo Metropolitan UniversityCircadian clock genes are best known for regulating circadian rhythms, but they also play crucial roles in memory processes. This suggests that memory is modulated by neural networks containing clock neurons, although the underlying mechanisms remain unclear. In Drosophila melanogaster, approximately 240 clock neurons are grouped into at least eight distinct clusters. Among them, the dorsal–lateral neurons (LNds) are required for maintaining long-term memory (LTM). In contrast, the neuropeptide Pigment-dispersing factor (Pdf), expressed in both small and large ventral–lateral neurons (s-LNvs and l-LNvs, respectively), functions as a circadian output signal and is also essential for maintaining LTM. In addition, Pdf-expressing neurons (hereafter, Pdf neurons) release neurotransmitters other than Pdf, which are involved in LTM consolidation. However, the specific transmitters used by LNds and Pdf neurons in LTM processing have remained unknown. Here, we show that octopamine signaling from LNds is essential for LTM maintenance, whereas octopamine in Pdf neurons is essential for LTM consolidation. Temporally restricted knockdown of Tyramine hydroxylase (Tbh), the gene encoding the enzyme required for octopamine synthesis, disrupted LTM maintenance when targeted in LNds, whereas it impaired LTM consolidation when targeted in Pdf neurons. Notably, Tbh knockdown in LNds or Pdf neurons had minimal effects on circadian behavioral rhythms or sleep. These findings reveal that octopamine released from specific subtypes of clock neurons independently regulates distinct phases of LTM in Drosophila.No potential conflict of interest relevant to this article was reported.Springer Science and Business Media LLCActa Medica Okayama2056-306X1112025Effect of temperature cycles on the sleep-like state in Hydra vulgaris2ENAyaSatoFaculty of Arts and Science, Kyushu UniversityManabuSekiguchiGraduate School of Natural Science and Technology, Okayama UniversityKogaNakadaGraduate School of Systems Life Sciences, Kyushu UniversityTaishiYoshiiGraduate School of Natural Science and Technology, Okayama UniversityTaichi Q.ItohFaculty of Arts and Science, Kyushu UniversityBackground Sleep is a conserved physiological phenomenon across species. It is mainly controlled by two processes: a circadian clock that regulates the timing of sleep and a homeostat that regulates the sleep drive. Even cnidarians, such as Hydra and jellyfish, which lack a brain, display sleep-like states. However, the manner in which environmental cues affect sleep-like states in these organisms remains unknown. In the present study, we investigated the effects of light and temperature cycles on the sleep-like state in Hydra vulgaris.<br>
Results Our findings indicate that Hydra responds to temperature cycles with a difference of up to 5 C, resulting in decreased sleep duration under light conditions and increased sleep duration in dark conditions. Furthermore, our results reveal that Hydra prioritizes temperature changes over light as an environmental cue. Additionally, our body resection experiments show tissue-specific responsiveness in the generation ofthe sleep-like state under different environmental cues. Specifically, the upper body can generate the sleep-like state in response to a single environmental cue. In contrast, the lower body did not respond to 12-h light–dark cycles at a constant temperature.<br>
Conclusions These findings indicate that both light and temperature influence the regulation of the sleep-like state in Hydra. Moreover, these observations highlight the existence of distinct regulatory mechanisms that govern patterns of the sleep-like state in brainless organisms, suggesting the potential involvement of specific regions for responsiveness of environmental cues for regulation of the sleep-like state.No potential conflict of interest relevant to this article was reported.Springer Science and Business Media LLCActa Medica Okayama2041-17231512024A high-protein diet-responsive gut hormone regulates behavioral and metabolic optimization in Drosophila melanogaster10819ENYutoYoshinariMetabolic Regulation and Genetics, Institute for Molecular and Cellular Regulation, Gunma UniversityTakashiNishimuraMetabolic Regulation and Genetics, Institute for Molecular and Cellular Regulation, Gunma UniversityTaishiYoshiiGraduate School of Environmental, Life, Natural Science and Technology, Okayama UniversityShuKondoDepartment of Biological Science and Technology, Faculty of Advanced Engineering, Tokyo University of ScienceHiromuTanimotoGraduate School of Life Sciences, Tohoku UniversityTomoeKobayashiDivision of Molecular Genetics, Shigei Medical Research InstituteMakotoMatsuyamaDivision of Molecular Genetics, Shigei Medical Research InstituteRyusukeNiwaLife Science Center for Survival Dynamics, Tsukuba Advanced Research Alliance (TARA), University of TsukubaProtein is essential for all living organisms; however, excessive protein intake can have adverse effects, such as hyperammonemia. Although mechanisms responding to protein deficiency are well-studied, there is a significant gap in our understanding of how organisms adaptively suppress excessive protein intake. In the present study, utilizing the fruit fly, Drosophila melanogaster, we discover that the peptide hormone CCHamide1 (CCHa1), secreted by enteroendocrine cells in response to a high-protein diet (HPD), is vital for suppressing overconsumption of protein. Gut-derived CCHa1 is received by a small subset of enteric neurons that produce short neuropeptide F, thereby modulating protein-specific satiety. Importantly, impairment of the CCHa1-mediated gut-enteric neuronal axis results in ammonia accumulation and a shortened lifespan under HPD conditions. Collectively, our findings unravel the crosstalk of gut hormone and neuronal pathways that orchestrate physiological responses to prevent and adapt to dietary protein overload.No potential conflict of interest relevant to this article was reported.SAGE PublicationsActa Medica Okayama0748-73042025Neurotransmitter and Receptor Mapping in Drosophila Circadian Clock Neurons via T2A-GAL4 ScreeningENAyumiFukudaGraduate School of Natural Science and Technology, Okayama UniversityAikaSaitoGraduate School of Environmental, Life, Natural Science and Technology, Okayama UniversityTaishiYoshiiGraduate School of Environmental, Life, Natural Science and Technology, Okayama UniversityThe circadian neuronal network in the brain comprises central pacemaker neurons and associated input and output pathways. These components work together to generate coherent rhythmicity, synchronize with environmental time cues, and convey circadian information to downstream neurons that regulate behaviors such as the sleep/wake cycle. To mediate these functions, neurotransmitters and neuromodulators play essential roles in transmitting and modulating signals between neurons. In Drosophila melanogaster, approximately 240 brain neurons function as clock neurons. Previous studies have identified several neurotransmitters and neuromodulators, including the Pigment-dispersing factor (PDF) neuropeptide, along with their corresponding receptors in clock neurons. However, our understanding of the neurotransmitters and receptors involved in the circadian system remains incomplete. In this study, we conducted a T2A-GAL4-based screening for neurotransmitter and receptor genes expressed in clock neurons. We identified 2 neurotransmitter-related genes and 22 receptor genes. Notably, while previous studies had reported the expression of 6 neuropeptide receptor genes in large ventrolateral neurons (l-LNv), we also found that 14 receptor genes\including those for dopamine, serotonin, and -aminobutyric acid\are expressed in l-LNv neurons. These findings suggest that l-LNv neurons serve as key integrative hubs within the circadian network, receiving diverse external signals.No potential conflict of interest relevant to this article was reported.The Endocrine SocietyActa Medica Okayama1945-717016682025Neuromedin U Deficiency Disrupts Daily Testosterone Fluctuation and Reduces Wheel-running Activity in Ratsbqaf102ENMaiOtsukaGraduate School of Natural Science and Technology, Okayama UniversityYuTakeuchiGraduate School of Natural Science and Technology, Okayama UniversityMahoMoriyamaGraduate School of Natural Science and Technology, Okayama UniversitySakuraEgoshiDepartment of Biology, Faculty of Science, Okayama UniversityYukiGotoGraduate School of Natural Science and Technology, Okayama UniversityTingtingGuGraduate School of Environmental, Life, Natural Science and Technology, Okayama UniversityAtsushi PKimuraDepartment of Biological Sciences, Faculty of Science, Hokkaido UniversityShogoHaraguchiDepartment of Biochemistry, Showa University School of MedicineTaishiYoshiiGraduate School of Natural Science and Technology, Okayama UniversitySakaeTakeuchiGraduate School of Natural Science and Technology, Okayama UniversityMakotoMatsuyamaDivision of Molecular Genetics, Shigei Medical Research InstituteGeorge EBentleyDepartment of Integrative Biology and Helen Wills Neuroscience Institute, University of California at BerkeleySayakaAizawaGraduate School of Natural Science and Technology, Okayama UniversityThe objective of this study was to elucidate the role of endogenous Neuromedin U (NMU) in rats by performing NMU knockout (KO). Male, but not female NMU KO rats exhibited decreased wheel-running activity vs wildtype (WT), although overall home cage activity was not affected. Plasma testosterone in WT rats varied significantly over the course of a day, with a peak at ZT1 and a nadir at ZT18, whereas in NMU KO rats testosterone remained stable throughout the day. Chronic administration of testosterone restored wheel-running activity in NMU KO rats to the same level as in WT rats, suggesting that the decrease in wheel-running activity in NMU KO rats is due to the disruption of the diurnal change of testosterone. Accordingly, expression of the luteinizing hormone beta subunit (Lhb) mRNA in the pars distalis of anterior pituitary was significantly lower in NMU KO rats; immunostaining revealed that the size of luteinizing hormone (LH)–expressing cells was also relatively small in those animals. In the brain of male WT rats, Nmu was highly expressed in the pars tuberalis, and the NMU receptor Nmur2 was highly expressed in the ependymal cell layer of the third ventricle. This study reveals a novel function of NMU and indicates that endogenous NMU in rats plays a role in the regulation of motivated activity via regulation of testosterone.No potential conflict of interest relevant to this article was reported.SAGE PublicationsActa Medica Okayama0748-73043952024A Detailed Re-Examination of the Period Gene Rescue Experiments Shows That Four to Six Cryptochrome-Positive Posterior Dorsal Clock Neurons (DN1p) of Drosophila melanogaster Can Control Morning and Evening Activity463483ENManabuSekiguchiGraduate School of Natural Science and Technology, Okayama UniversityNilsReinhardNeurobiology and Genetics, Theodor-Boveri Institute, Biocenter, University of WürzburgAyumiFukudaGraduate School of Natural Science and Technology, Okayama UniversityShunKatohGraduate School of Environmental, Life, Natural Science and Technology, Okayama UniversityDirkRiegerNeurobiology and Genetics, Theodor-Boveri Institute, Biocenter, University of WürzburgCharlotteHelfrich-FörsterNeurobiology and Genetics, Theodor-Boveri Institute, Biocenter, University of WürzburgTaishiYoshiiGraduate School of Natural Science and Technology, Okayama UniversityAnimal circadian clocks play a crucial role in regulating behavioral adaptations to daily environmental changes. The fruit fly Drosophila melanogaster exhibits 2 prominent peaks of activity in the morning and evening, known as morning (M) and evening (E) peaks. These peaks are controlled by 2 distinct circadian oscillators located in separate groups of clock neurons in the brain. To investigate the clock neurons responsible for the M and E peaks, a cell-specific gene expression system, the GAL4-UAS system, has been commonly employed. In this study, we re-examined the two-oscillator model for the M and E peaks of Drosophila by utilizing more than 50 Gal4 lines in conjunction with the UAS-period16 line, which enables the restoration of the clock function in specific cells in the period (per) null mutant background. Previous studies have indicated that the group of small ventrolateral neurons (s-LNv) is responsible for controlling the M peak, while the other group, consisting of the 5th ventrolateral neuron (5th LNv) and the three cryptochrome (CRY)-positive dorsolateral neurons (LNd), is responsible for the E peak. Furthermore, the group of posterior dorsal neurons 1 (DN1p) is thought to also contain M and E oscillators. In this study, we found that Gal4 lines directed at the same clock neuron groups can lead to different results, underscoring the fact that activity patterns are influenced by many factors. Nevertheless, we were able to confirm previous findings that the entire network of circadian clock neurons controls M and E peaks, with the lateral neurons playing a dominant role. In addition, we demonstrate that 4 to 6 CRY-positive DN1p cells are sufficient to generate M and E peaks in light-dark cycles and complex free-running rhythms in constant darkness. Ultimately, our detailed screening could serve as a catalog to choose the best Gal4 lines that can be used to rescue per in specific clock neurons.No potential conflict of interest relevant to this article was reported.Springer Science and Business Media LLCActa Medica Okayama0340-75942023A four-oscillator model of seasonally adapted morning and evening activities in Drosophila melanogasterENTaishiYoshiiGraduate School of Natural Science and Technology, Okayama UniversityAikaSaitoGraduate School of Natural Science and Technology, Okayama UniversityTatsuyaYokosakoGraduate School of Natural Science and Technology, Okayama UniversityThe fruit fly Drosophila melanogaster exhibits two activity peaks, one in the morning and another in the evening. Because the two peaks change phase depending on the photoperiod they are exposed to, they are convenient for studying responses of the circadian clock to seasonal changes. To explain the phase determination of the two peaks, Drosophila researchers have employed the two-oscillator model, in which two oscillators control the two peaks. The two oscillators reside in different subsets of neurons in the brain, which express clock genes, the so-called clock neurons. However, the mechanism underlying the activity of the two peaks is complex and requires a new model for mechanistic exploration. Here, we hypothesize a four-oscillator model that controls the bimodal rhythms. The four oscillators that reside in different clock neurons regulate activity in the morning and evening and sleep during the midday and at night. In this way, bimodal rhythms are formed by interactions among the four oscillators (two activity and two sleep oscillators), which may judiciously explain the flexible waveform of activity rhythms under different photoperiod conditions. Although still hypothetical, this model would provide a new perspective on the seasonal adaptation of the two activity peaks.No potential conflict of interest relevant to this article was reported.Informa UK LimitedActa Medica Okayama0742-05284032023Pigment-dispersing factor and CCHamide1 in the Drosophila circadian clock network284299ENRikoKuwanoGraduate School of Natural Science and Technology, Okayama UniversityMakiKatsuraGraduate School of Natural Science and Technology, Okayama UniversityMaiIwataGraduate School of Natural Science and Technology, Okayama UniversityTatsuyaYokosakoGraduate School of Natural Science and Technology, Okayama UniversityTaishiYoshiiGraduate School of Natural Science and Technology, Okayama UniversityAnimals possess a circadian central clock in the brain, where circadian behavioural rhythms are generated. In the fruit fly (Drosophila melanogaster), the central clock comprises a network of approximately 150 clock neurons, which is important for the maintenance of a coherent and robust rhythm. Several neuropeptides involved in the network have been identified, including Pigment-dispersing factor (PDF) and CCHamide1 (CCHa1) neuropeptides. PDF signals bidirectionally to CCHa1-positive clock neurons; thus, the clock neuron groups expressing PDF and CCHa1 interact reciprocally. However, the role of these interactions in molecular and behavioural rhythms remains elusive. In this study, we generated Pdf (01) and CCHa1(SK8) double mutants and examined their locomotor activity-related rhythms. The single mutants of Pdf (01) or CCHa1(SK8) displayed free-running rhythms under constant dark conditions, whereas approximately 98% of the double mutants were arrhythmic. In light-dark conditions, the evening activity of the double mutants was phase-advanced compared with that of the single mutants. In contrast, both the single and double mutants had diminished morning activity. These results suggest that the effects of the double mutation varied in behavioural parameters. The double and triple mutants of per (01), Pdf (01), and CCHa1(SK8) further revealed that PDF signalling plays a role in the suppression of activity during the daytime under a clock-less background. Our results provide insights into the interactions between PDF and CCHa1 signalling and their roles in activity rhythms.No potential conflict of interest relevant to this article was reported.WileyActa Medica Okayama0179-16131282022Artificial selections for death-feigning behavior in beetles show correlated responses in amplitude of circadian rhythms, but the period of the rhythm does not453460ENTakahisaMiyatakeGraduate School of Environmental and Life Science, Okayama UniversityMasatoS. AbeCenter for Advanced Intelligence Project, RIKENKentarouMatsumuraLaboratory of entomology, Faculty of AgricultureTaishiYoshiiGraduate School of Natural Science and Technology, Okayama UniversityOne of the most important survival strategies of organisms is to avoid predators. Studying one of such strategies, namely, death-feigning behavior, has recently become more common. The success or failure of this antipredator strategy will be affected by the circadian rhythms of both prey and predator because death feigning sometimes has a diurnal rhythm. However, few studies have analyzed the effects of differences in circadian rhythms on predator-avoidance behavior at the genetic level. Recently, the relationship between genes relating to circadian rhythm and death-feigning behavior, an antipredator behavior, has been established at the molecular level. Therefore, in this study, we compared three circadian rhythm-related traits, the free-running period of rhythms, amplitude of circadian rhythms, and total activity of strains of three Tribolium species that were artificially selected for the death-feigning duration: short (S-strains) and long (L-strains) durations. As a result, the amplitude of circadian rhythms and total activity were significantly different between S- and L-strains, but there was no difference in the free-running periods of the rhythm between the strains in T. castaneum, T. confusum, and T. freemani. Although the relationship between death-feigning behavior and activity has been reported for all three species, a genetic relationship between the duration of death feigning and the amplitude of circadian rhythms has been newly found in the present study. It is important to investigate the relationship between antipredator strategies and circadian rhythms at the molecular level in the future.No potential conflict of interest relevant to this article was reported.Public Library of ScienceActa Medica Okayama1932-62031612021Amplitude of circadian rhythms becomes weaken in the north, but there is no cline in the period of rhythm in a beetlee0245115ENMasato S.AbeCenter for Advanced Intelligence Project, RIKENKentarouMatsumuraGraduate School of Environmental and Life Science, Okayama UniversityTaishiYoshii Graduate School of Natural Science and Technology, Okayama UniversityTakahisaMiyatakeGraduate School of Environmental and Life Science, Okayama UniversityMany species show rhythmicity in activity, from the timing of flowering in plants to that of foraging behavior in animals. The free-running periods and amplitude (sometimes called strength or power) of circadian rhythms are often used as indicators of biological clocks. Many reports have shown that these traits are highly geographically variable, and interestingly, they often show latitudinal or longitudinal clines. In many cases, the higher the latitude is, the longer the free-running circadian period (i.e., period of rhythm) in insects and plants. However, reports of positive correlations between latitude or longitude and circadian rhythm traits, including free-running periods, the power of the rhythm and locomotor activity, are limited to certain taxonomic groups. Therefore, we collected a cosmopolitan stored-product pest species, the red flour beetle Tribolium castaneum, in various parts of Japan and examined its rhythm traits, including the power and period of the rhythm, which were calculated from locomotor activity. The analysis revealed that the power was significantly lower for beetles collected in northern areas than southern areas in Japan. However, it is worth noting that the period of circadian rhythm did not show any clines; specifically, it did not vary among the sampling sites, despite the very large sample size (n = 1585). We discuss why these cline trends were observed in T. castaneum.No potential conflict of interest relevant to this article was reported.Oxford University PressActa Medica Okayama0024-406613012020Genetic variation and phenotypic plasticity in circadian rhythms in an armed beetle, Gnatocerus cornutus (Tenebrionidae)3440ENKentarouMatsumuraGraduate School of Environmental and Life Science, Okayama UniversityMasato SAbeCenter for Advanced Intelligence Project, RIKENManmohan DSharmaCentre for Ecology and Conservation, School of Biosciences, University of ExeterDavid JHoskenCentre for Ecology and Conservation, School of Biosciences, University of ExeterTaishiYoshiiGraduate School of Natural Science and Technology, Okayama UniversityTakahisaMiyatakeGraduate School of Environmental and Life Science, Okayama UniversityCircadian rhythms, their free-running periods and the power of the rhythms are often used as indicators of biological clocks, and there is evidence that the free-running periods of circadian rhythms are not affected by environmental factors, such as temperature. However, there are few studies of environmental effects on the power of the rhythms, and it is not clear whether temperature compensation is universal. Additionally, genetic variation and phenotypic plasticity in biological clocks are important for understanding the evolution of biological rhythms, but genetic and plastic effects are rarely investigated. Here, we used 18 isofemale lines (genotypes) of Gnatocerus cornutus to assess rhythms of locomotor activity, while also testing for temperature effects. We found that total activity and the power of the circadian rhythm were affected by interactions between sex and genotype or between sex, genotype and temperature. The males tended to be more active and showed greater increases in activity, but this effect varied across both genotypes and temperatures. The period of activity varied only by genotype and was thus independent of temperature. The complicated genotype–sex–environment interactions we recorded stress the importance of investigating circadian activity in more integrated ways.No potential conflict of interest relevant to this article was reported.SAGE PublicationsActa Medica Okayama0748-73042019A Catalog of GAL4 Drivers for Labeling and Manipulating Circadian Clock Neurons in Drosophila melanogasterENManabuSekiguchiMatching Program Course, Okayama UniversityKotaroInoueGraduate School of Natural Science and Technology, Okayama UniversityTianYangGraduate School of Natural Science and Technology, Okayama UniversityDong-GenLuoGraduate School of Natural Science and Technology, Okayama UniversityTaishiYoshiiGraduate School of Natural Science and Technology, Okayama University Daily rhythms of physiology, metabolism, and behavior are orchestrated by a central circadian clock. In mice, this clock is coordinated by the suprachiasmatic nucleus, which consists of 20,000 neurons, making it challenging to characterize individual neurons. In Drosophila, the clock is controlled by only 150 clock neurons that distribute across the fly's brain. Here, we describe a comprehensive set of genetic drivers to facilitate individual characterization of Drosophila clock neurons. We screened GAL4 lines that were obtained from Drosophila stock centers and identified 63 lines that exhibit expression in subsets of central clock neurons. Furthermore, we generated split-GAL4 lines that exhibit specific expression in subsets of clock neurons such as the 2 DN2 neurons and the 6 LPN neurons. Together with existing driver lines, these newly identified ones are versatile tools that will facilitate a better understanding of the Drosophila central circadian clock.No potential conflict of interest relevant to this article was reported.Society for NeuroscienceActa Medica Okayama0270-647435152015Cryptochrome-dependent and -independent circadian entrainment circuits in Drosophila.61316141ENTaishiYoshiiChristianeHermann-LuiblChristaKistenpfennigBenjaminSchmidKenjiTomiokaCharlotteHelfrich-Förster@Entrainment to environmental light/dark (LD) cycles is a central function of circadian clocks. In Drosophila, entrainment is achieved by Cryptochrome (CRY) and input from the visual system. During activation by brief light pulses, CRY triggers the degradation of TIMELESS and subsequent shift in circadian phase. This is less important for LD entrainment, leading to questions regarding light input circuits and mechanisms from the visual system. Recent studies show that different subsets of brain pacemaker clock neurons, the morning (M) and evening (E) oscillators, have distinct functions in light entrainment. However, the role of CRY in M and E oscillators for entrainment to LD cycles is unknown. Here, we address this question by selectively expressing CRY in different subsets of clock neurons in a cry-null (cry0) mutant background. We were able to rescue the light entrainment deficits of cry0 mutants by expressing CRY in E oscillators but not in any other clock neurons. Par domain protein 1 molecular oscillations in the E, but not M, cells of cry0 mutants still responded to the LD phase delay. This residual light response was stemming from the visual system because it disappeared when all external photoreceptors were ablated genetically. We concluded that the E oscillators are the targets of light input via CRY and the visual system and are required for normal light entrainment.No potential conflict of interest relevant to this article was reported.Okayama UniversityActa Medica Okayama122015Molecular features of the circadian clock system in fruit fliesENTaishiYoshiiNo potential conflict of interest relevant to this article was reported.