동물의 종 유지에 있어서 교미행동은 매우 중요한 위치를 차지함. 교미과정에서 수컷의 경우 모든 암컷에 대해 성행동을 보이는반면, 암컷은 가장 적절한 교미의 시기를 정하며, 적절한 수컷을 선택하고 교미행동을 보이기 때문에 암컷의 교미행동을 유도하는 과정은 생물학에서 매우 중요한 의미를 지님. 본 연구에서는 초파리 (Drosophila melanogaster) 전자현미경자료와 총신경망분석 (Connectomics)을 이용하여 초파리 뇌에서 암컷의 교미행동을 조절하는 것으로 알려진 pC1신경의 하부신경 pC1b,c가 성적 성숙과정에서 교미를 하고자 하는 욕구 (sexual drive)를 증가시키는 기능을 하는 것을 처음으로 밝힘. 본 연구에서는 처음으로 pC1b,c 신경 내의 cAMP 수준이 교미의 욕구변화를 보여주는 중요한 물질이라는 것을 밝혔고 cAMP의 수준이 오르기 위해서는 신경펩티 드인 Dh44와 그 수용체 GPCR인 Dh44R1과 Dh44R2가 필요하다는 것을 확인함. 또한 cAMP의 변화는 신경내의 CREBB를 통하여 하위 유전자인 pyx (pyrexia)의 발현을 유도한다는 것을 밝힘. 본 연구로 종 유지 메커니즘을 좀 더 이해할 수 있음.

From invertebrate to vertebrate, females uptake sperm for a specific duration post-copulation known as the ejaculate holding period (EHP) before expelling un-stored sperm and the mating plug through sperm ejection. Our study uncovered that encountering males or mated females after mating substantially shortens EHP, a phenomenon we term ‘male-induced EHP shortening (MIES)’. MIES requires Or47b+ olfactory and ppk23+ gustatory neurons, activated by 2-methyltetracosane and 7-tricosene, respectively. These odorants raise cAMP levels in pC1b and c neurons, which are responsible for processing male courtship and regulating female receptivity. Elevated cAMP levels in pC1b and c reduce EHP and reinstate their responsiveness to male courtship cues, promoting re-mating with faster sperm ejection. This study establishes MIES as a genetically tractable model of sexual plasticity with a conserved neural mechanism.

난황형성과정(Vitellogenesis)은 발달하는 난모세포에 난황이 축적되는 과정으로, 이 과정의 개시는 알형성과정(oogenesis)을 제어하는 주 요 메커니즘이다. 곤충생리학 모델인 노랑초파리(Drosophila melanogaster)에서 난황형성과정은 성충으로 우화한 직후 시작하여 성적 성숙이 일어 나는 2-3일간 지속된다. 성숙한 난모세포가 충분히 만들어지고 성적 성숙이 종료되면, 짝짓기 후 알형성과정이 다시 시작될 때까지 난황형성과정은 멈춘다. 수컷 초파리의 정액 단백질인 성 펩타이드(Sex peptide, SP)는 짝짓기의 신호로서 알라타체(corpora allata)를 자극해 유약호르몬 (Juvenile hormone, JH) 생합성 및 분비를 유도하며, 혈림프(hemolymph) JH 농도의 증가는 난황형성과정을 자극한다. 최근 연구 결과에 따르 면, SP수용체 뉴런은 자궁 내막의 수상돌기를 통해 교미 중 정액과 함께 자궁으로 전달된 SP를 감지함으로써, 축삭돌기를 통해 중추신경계인 복부 신경절에 짝짓기 신호를 보내는데, 이러한 중추신경계 SP 신호가 JH 생합성 및 분비, 그리고 난황형성과정을 유도하는 것으로 밝혀졌다. 짝짓기 후 암컷에서의 난황형성과정은 일주기 리듬을 보이는데, 노랑초파리의 일주기 리듬은 중추 신경계 뉴런들에 의해 제어된다. 본 종설은 성적 성숙, 짝짓기 신호, 그리고 일주기 리듬에 따라 난황형성과정을 제어하는 신경 메커니즘에 관한 최근 연구 성과를 다룬다.

During insect development from embryo to adult, airway clearance in tracheal system occurs episodically each time the molt is completed by performance of the ecdysis sequence. We found that the neuropeptide Kinin is required for normal respiratory fluid clearance or “tracheal air-filling” in fruit fly Drosophila larvae. Disruption of Kinin signaling leads to defective air-filling during all larval stages. Targeted Kinin receptor silencing in tracheal epithelial cells also shows tracheal air-filling defect. On the other hand, promotion of Kinin signaling in vivo through peptide injection or Kinin neuron activation induces premature tracheal collapse and air-filling. Moreover, direct exposure of epithelial cells in vitro to Kinin leads to calcium mobilization in tracheal epithelial cells. Our findings strongly implicate the neuropeptide Kinin as an important regulator of airway clearance via intracellular calcium mobilization in tracheal epithelial cells of fruit fly Drosophila.

Reproduction is essential for the survival of a species and is under the close coordination of the central nerve system (CNS). In Drosophila melanogaster, juvenile hormones (JHs) from the corpora allatum(CA) are essential for egg-production because they promote vitellogenesis and enhance ecdysteroid biosynthesis. JH biosynthesis is under the control of neuropeptides, such as Drosophila insulin-like peptide (dilp) and allatostatin C (AstC). However, how these neuropeptide pathways coordinate JH production remain elusive. Here, we report a small number of neurons in the brain and ventral nerve cord (VNC) neurons that produce a neuropeptide AstC regulate egg-production negatively. The brain neurons seem to act indirectly by inhibiting secretion of dilp that up-regulates JH production, whereas the VNC neurons seem to release AstC into the hemolymph to suppress the CA activity directly. On the CA, the AstC actions appear mediated by two G-protein coupled receptors, AstC-R1 and AstC-R2. We will discuss our recent efforts to identify agonists for either receptor, which will be potential leads for insect reproduction regulators.

Animals must maintain proper balance between energy intake and expenditure. Recently, we descovered the enzymaticco-factor tetrahydrobiopterin (BH4) inhibits feeding in Drosophila. BH4 biosynthesis requires the sequential action of theconserved enzymes Punch, Purple, and Sepiapterin Reductase (Sptr). Although we observe increased feeding upon lossof Punch and Purple in the adult fat body, loss of Sptr must occur in the brain. We found Sptr expression is requiredin four adult brain neurons that express NPF, the fly homologue of the vertebrate appetite regulator NPY. Mechanistically,we found BH4 deficiency reduces NPF levels, while excess BH4 increases NPF accumulation without altering its expression.

In many insects, female receives a large amount of sperm and ejaculates upon copulation, and keeps them in her uterus for some time, during which she stores sperm. After varying delays, the female removes unused sperm and ejaculates through sperm ejection behavior, which is known as a major mechanism for the post-copulatory sexual selection. In the fruitfly Drosophila melanogaster, timing of sperm ejection behavior is controlled by the diuretic hormone 44 (Dh44) pathway. To identify pathways that regulate Dh44 activities and sperm ejection, here we examine effects of olfactory cues. We will present our recent results suggesting that male odors can be one of upstream signaling pathway that modulates a brain neuropeptide pathway.

The ecdysis behavioral sequence in insects is a classic fixed action pattern (FAP) initiated by hormonal signaling. Ecdysis triggering hormones (ETHs) release the FAP through direct actions on the CNS. Here we present evidence implicating two groups of central ETH receptor (ETHR) neurons in scheduling the first two steps of the FAP: kinin (aka drosokinin, leucokinin) neurons regulate pre-ecdysis behavior and CAMB neurons (CCAP, AstCC, MIP, and Bursicon) initiate the switch to ecdysis behavior. Ablation of kinin neurons or altering levels of ETH receptor (ETHR) expression in these neurons modifies timing and intensity of pre-ecdysis behavior. Cell ablation or ETHR knockdown in CAMB neurons delays the switch to ecdysis, whereas overexpression of ETHR or expression of pertussis toxin in these neurons accelerates timing of the switch. Calcium dynamics in kinin neurons are temporally aligned with pre-ecdysis behavior, whereas activity of CAMB neurons coincides with the switch from pre-ecdysis to ecdysis. Activation of CCAP or CAMB neurons through temperature-sensitive TRPM8 gating is both necessary and sufficient to trigger ecdysis behavior. Our findings demonstrate that kinin and CAMB neurons are direct targets of ETH and play critical roles in scheduling successive behavioral steps in the ecdysis FAP. Furthermore, temporal organization of the FAP is likely a function of ETH receptor density in target neurons.

Internal sperm storage after mating is important for insect reproduction, because it permits delayed fertilisation, and post-copulatory mate choice in polyandrous females. The polyandry is common in many animal taxa including insects, because it increases female fitness by reducing the risk of infertility and providing opportunities for sperm competition and choice. The reproductive success of males, on the other hand, often depends upon avoidance of sperm competition by preventing mated females from copulating and receiving sperm from other male suitors. A widespread strategy used by males is the use of the male seminal fluid proteins (SFPs) that form the mating plug and alter female behaviors, for example by suppressing mating receptivity and elevating egg-laying. Under these circumstances, females are expected to evolve mechanism(s) to control exposure to the male SFPs in order to maximize fitness by balancing the positive and negative impacts of polyandry. Here, we discover that Drosophila melanogaster females eject male ejaculates 1-6 h after mating with a stereotypic behaviour, and that this is regulated by a brain neuropeptide pathway composed of diuretic hormone 44 (Dh44), and its receptor Dh44R1. We showed that suppressing Dh44 or Dh44R1 signals in the brain expedites sperm ejection, whereas enhancing Dh44 or Dh44R1 signals delays sperm ejection. This study uncovers a molecular mechanism by which females can influence sperm competition and selection, and counter actively the negative impact of polyandry.

Alzheimer’s disease (AD) is the most common type of presenile and senile dementia. Human β-amyloid precursor cleavage enzyme (BACE-1) is a key enzyme responsible for amyloid plaque production. We assessed anti-BACE-1 and behavioral activities of curcuminoids from Curcuma longa, curcumin (CCN), demethoxycurcumin (DMCCN), and bisdemethoxycurcumin (BDMCCN) against AD fly models. Neuro-protective ability of curcuminoids was assessed using fly model system overexpressing BACE-1 and its substrate APP in compound eyes and entire neurons. BDMCCN has the strongest inhibitory activity toward BACE-1 with 17 μM IC50, which was 20 and 13 times lower than those of CCN and DMCCN respectively. Expression of APP/BACE-1 resulted in the progressive and measurable defects in morphology of eyes and locomotion. Supplementing diet with either 1 mM BDMCCN or CCN rescued APP/BACE1 expressing flies and kept them from developing both morphological and behavioral defects. Structural characteristics and hydrophobicity appear to play a role in determining inhibitory potency of curcuminoids on BACE-1.

An assessment was made of beta-site amyloid precursor protein (APP) cleaving enzyme (BACE1) inhibitory, feeding, climbing activities and lifespan of the diarylalkyls curcumin (CCN), demethoxycurcumin (DCCN) and bisdemethoxycurcumin (BDCCN) identified in the rhizomes of Curcuma longa. Based on IC50 values, BDCCN (0.024 mM) was the most inhibitory constituent, followed by DCCN (0.31 mM) and CCN (0.59 mM). Overall the three curcuminoids were significantly less inhibitory than BACE1 inhibitor IV isophthalamide (8.5 × 10-5 mM). The expression of human APP and BACE1 in compound eye of Drosophila melangaster presented rough abnormal ommatidial lattice. Co-expression of APP and BACE1 within the developing nervous system of drosophila showed climbing defects. These transgenic flies kept on media containing 1 mM of CCN and BDCCN were observed to ameliorate eye degeneration, significantly suppress locomotive dysfunctions, and increase media life time, as well as isophthalamide. CCN and BDCCN as human BACE1 inhibitory constituents may be used as potential therapeutics or lead molecules to develop Alzheimer's disease treatment drugs.

The aim of this study was to evaluate surface character¬istics and biological properties of the dentin -derived hydroxyapatite (HA) coating on titanium substrate. Dentin-derived HA was obtained from extracted human teeth using a calcination method at 850℃. The commercially pure titanium (cp-Ti, ASTM Grade II) was used as a metallic substrate and a radio frequency magnetron sputtering method was employed as a coating method. Scanning electron microscopy (SEM) and energy dispersive X-ray analysis (EDX) were utilized to investigate the coating aspects and composition. Atomic forced microscopy (AFM) and a surface profiler were used to assess the surface morphology and roughness. Corrosion tests were performed in phosphate- buffered saline at a 36.5 ± 1℃ in order to determine the corrosion behavior of the uncoated and coated specimens. The biocompatibility of dentin-derived HA coated specimens with fetal rat calvarial cells and human gingival fibroblasts was assessed by SEM and cell prolif¬eration analysis. The results showed that the dentin-derived HA coatings appeared to cover thinly and homogeneously the surfaces without changing of the titanium substrate. The EDX analysis of this the coating surface indicated the presence of Ca and P elements. The mean surface roughness of cp-Ti and dentin-derived coating specimens was 0.27 µm and, 1.7 µm, respectively. Corrosion tests indicated a stable passive film of the dentin-derived HA coating specimens. SEM observations of fetal rat calvarial cells and human fibroblast cells on coated surfaces showed that the cells proliferated and developed a network of dense interconnections. The cells on all specimens proliferated actively within the culture period, showing good cell viability. At day 1 and 3, dentin-derived coating specimens showed 89% and 93% cell viability, respectively, when normalized to cp-Ti specimens. These results suggest that dentin-derived HA coating using the RF magnetron sputtering method has good surface characteristics and biocompatibility.

Upon mating, females of many animal species undergo dramatic changes in their behavior. In Drosophila melanogaster, post-mating behaviors are triggered by sex peptide (SP), a key modulatory substance produced in the male seminal fluid and transferred to female during copulation. SP modulates female behaviors by acting on the sex peptide receptor (SPR) located in a small subset of internal sensory neurons that innervate the female uterus and project to the central nervous system (CNS). Interestingly, however, SPR is also expressed broadly in the CNS of both sexes. Moreover, SPR is also encoded in the genomes of insects that lack obvious SP orthologs. Based on these observations, we speculated that SPR may have additional ligands that are only distantly related to SP, if at all. If so, then this also raises questions on the evolution of SP-SPR signaling. To begin to address these questions, we set out to identify additional ligands for SPR. Here, we identify myoinhibitory peptides (MIPs) as a second family of SPR ligands that is conserved across a wide range of invertebrate species. MIPs are potent agonists for Drosophila, Aedes and Aplysia SPRs in vitro, yet are unable to trigger post-mating responses in vivo. In contrast to SP, MIPs are not produced in male reproductive organs, and are not required for post-mating behaviors in Drosophila females. We conclude that MIPs are evolutionarily conserved ligands for SPR, which are likely to mediate functions other than the regulation of female reproductive behaviors. Therefore, we propose that SPR has a different ancestral function, with a role in post-mating behavior arising only recently in Drosophila evolution, concomitant with the emergence of its novel SP ligand.