The energy usage of parallel chillers systems accounts for 25–40 % of the total energy cost of a building. In light of global warming concerns and the need for energy conservation, it is essential to distribute the load of the parallel chillers systems effectively to achieve energy savings in buildings. Accordingly, this study presents a multi-strategy improved sparrow search algorithm (MSSA) to address optimization of the optimal chillers loading (OCL) problem. The proposed algorithm augments the basic sparrow search algorithm (SSA) by introducing the Sine chaotic map, Levy flight method, and Cauchy variation to enhance diversity, avoid local optima, and increase global and local search capacities. We use 9 benchmark functions to check the performance of the proposed MSSA algorithm, and the results are better than the selected algorithms such as particle swarm algorithm (PSO), harris hawks optimization (HHO), artificial rabbit optimization (ARO) and sparrow search algorithm (SSA). In addition, MSSA is applied to two typical cases to demonstrate its performance to optimal chillers loading and the results indicate that the MSSA outperforms similar algorithms. This study validates that MSSA can provide a promising solution to resolve the OCL problem. This haEnergy system optimization research has become an important field, and in the field of building refrigeration, due to global warming led to building refrigeration energy consumption continues to rise , aim to With the rapid development of artificial intelligence recently, a significant number of scholars are committed to optimal chillers loading using various optimization algorithms. As early as the start of this century, some scholars have adopted precise mathematical methods, such as Chinese scholars Chang et al.  applied SSA can significantly resolve the problem of support vector machine parameter optimization. Wang et al. [Sparrow search algorithm (SSA)  is an ing et al.  integrat(1)Introduction of the Sine chaotic map, Levy flight method, and Cauchy mutation method to enhance algorithm performance based on the original SSA algorithm.(2)Nine benchmark functions are used to test the optimization ability of the improved MSSA, and the results illustrate that the improved MSSA has higher search accuracy, faster convergence speed and stronger robustness.(3)The suitability of MSSA to optimal chillers loading is validated by testing two typical examples, wherein the substantial potential for energy-saving is demonstrated.To overcome the shortcomings of current algorithms in solving OCL problems, such as low accuracy, slow convergence, high computational complexity, and weak robustness, we proposed a multi-strategy modified sparrow search algorithm (MSSA) to optimize chillers loading. The primary contributions are as follows.The remainder of the paper is arranged as follows: Section 2d, c, b, and a represent the performance parameters of the unit itself; Thus, based on the above description, the OCL problems seeks to minimize the entire energy cost of the system through reasonable distribution of the load rate for each chiller unit while ensuring the energy cost demand of terminals. Typically, we can derive the relationship between the part load rate and power cost using the operating data of the chillers, as represented by Equation PLR of the unit is small, it is susceptible to damage, necessitating the constraint of the part load rate. Concurrently, the aggregate load for the system must fulfill the energy demand of the building terminal. The PLR constraints and the power equation constraint are represented by Equation When the PLR of each chiller unit under optimal working conditions. The target equation is expressed as Equation Ultimately, the crux of solving the OCL problem lies in determining the minimum energy consumption of the parallel chillers system and the 33.1The sparrow search algorithm was initially proposed as a meta-heuristic optimization algorithm inspired by sparrows' predation and anti-predation behaviors. It exhibits a robust global search ability and is used extensively to resolve optimization problems across various fields. Sparrows are categorized into producers, scroungers, and scouters.n sparrows are randomly produced to form the initial sparrow population, with its population matrix equation articulated as Equation n denotes the quantity of sparrows and m is decision variable dimension of the problem that needs to be solved.Initially, The initial fitness is calculated as Equation Producers exhibit superior fitness values, granting them priority access to food. They are responsible for foraging throughout the sparrow population, providing foraging direction. The position updates are presented in Equation Scroungers in sparrows’ population continuously monitor producer activities. If producers discover better food, scroungers compete for it, receiving food upon success. In contrast, if they fail, they fly to other territories in search of nourishment. Scrounger position updates are expressed by Equation When a scouter detects them to be at risk, it departs the current area and flies to the best area. Their position updates are shown in Equation 3.2Notwithstanding the potential of SSA to tackle the OCL problem, there is still considerable scope for enhancing search accuracy and convergence speed. Hence, we propose a multi-strategy improved sparrow search algorithm (MSSA), which introduces the Sine chaotic map for population initialization, Levy flight mechanism to improve producer location, and Cauchy mutation to enhance the scroungers' search capabilities in SSA. The details are outlined in this section.3.2.1To amplify the population diversity, the Sine chaotic map is introduced to produce the sparrow population and its population generation mode is expressed by Equation 3.2.2The Levy flight strategy mimics birds' flight trajectory, characterized by random alternating between long and short distances. As the primary optimization group, producers leverage the Levy flight method to enhance the global and local search capabilities of the algorithm, facilitating quick convergence to the optimal value. The updated producer location is expressed in Equation 3.2.3Scroungers serve as the secondary foragers in the sparrow population, with their position update mode impacted by producers' current optimal value. To prevent scroungers from getting trapped in a local optimal value, the Cauchy variation is introduced to perturb their position updating, taking advantage of the characteristics of the Cauchy distribution that it has a small peak value at the origin and long distribution at both ends. Our goal is to improve the local search ability and the quality of the optimal solution . If the 3.2.4(1)Basic algorithm parameters such as the size of population, the maximum number of iterations is initialized.(2)The Sine chaotic map generates the initial population.(3)The fitness value of sparrows is calculated and sorted.(4)When (5)When (6)Scouters' positions are updated according to Equation (7)The optimal fitness value and position of the current population are updated.(8)Determine if the termination criterion is satisfied. When satisfied, output the result.3.3PLR of each chiller.To enhance our understanding of how to use the MSSA algorithm to tackle the OCL problem, 4i7-4790K@4.0 processor and 64 GB of memory and MATLAB R2021a is testing tool.To verify the performance of the proposed MSSA, we tested it applying nine well-known benchmark functions and compared the results with the PSO, HHO, ARO, and SSA algorithms. First, we give the details of benchmark function and the setting of the test algorithms, and then we analyze the search accuracy, convergence speed, robustness in detail. In order to ensure a consistent testing environment, Windows 11 is utilized as the operating system, with an Intel Core (TM) 4.1The nine benchmark functions used in the tests are unimodal functions  5.2In the two cases, the results of MSSA were compared with GA , PSO 4545, SSA , ARO 5959 and HHAs illustrated in (1)MSSA algorithm is better suited for systems with a large number of chillers. Compared to commonly used optimization algorithms such as GA, PSO and SSA, MSSA consistently demonstrates a distinct energy-saving potential.(2)When the building energy consumption demand is at a low level, the energy saving potential of MSSA is greater.Similar to the characteristics of the previous benchmark function test performance, compared with SSA, because MSSA introduces levy flight method to enhances the search capability, MSSA can obtain competitive solutions with higher accuracy in OCL problems compared with SSA. In addition, as many decision variables and complex constraint conditions of OCL problem, it is very difficult for us to obtain the theoretical reference best value by mathematical method. However, the test results of two cases show that the calculation results of MSSA algorithm are equal to or better than many existing algorithms, so we believe that MSSA can provide a relatively optimal solution for OCL problem at present. In a word, as HVAC systems of most public buildings function at low load levels for most of the time, MSSA algorithm has significant energy-saving potential in HVAC systems operation.5.3For the evaluation of OCL algorithm performance, we believe that the search accuracy is the most important, followed by the convergence speed. If an algorithm cannot solve a competitive solution, it means that it is not necessary to compare its convergence speed with other algorithms with higher search accuracy.Compared to the GA, PSO and SSA algorithms, the MSSA algorithm displays unparalleled search accuracy. To further validate the advantages of the MSSA algorithm in tackling OCL problems, we compare it with the ARO  and HHO To obtain a comprehensive understanding of the convergence capability of the MSSA, we selected one high-load demand condition and one low-load demand condition for each case. 5.4In optimization problems, an excellent algorithm should exhibit low computational complexity. We employed the cumulative running time of 30 operations to assess the computational complexity of the three algorithms. 5.5In addition, to conduct a comprehensive analysis of MSSA, In Case 1, 6(1)After testing the benchmark function and comparing the results with PSO, HHO, ARO and SSA algorithms, we believe that the proposed MSSA has significant optimization ability.(2)In the test of solving OCL problem, two classical cases show that the proposed MSSA can provide a competitive energy saving scheme. Compared to GA, PSO and SSA algorithms, for Case 1, when the load demand is 1160RT, MSSA can provide a maximum energy-saving potential of 205.23 kW, which has a significant energy-saving impact. Furthermore, the test results of both cases demonstrate that the lower the end-building energy demand, the greater the energy-saving potential of MSSA.(3)For OCL problems, more test results illustrate that MSSA has significant advantages in convergence speed, computational complexity and robustness, which preliminarily proves the effectiveness of MSSA in solving such problems.To tackle the OCL problem effectively, this paper proposes a multi-strategy improved sparrow search algorithm. Through testing two typical cases, we draw the following conclusions.In summary, the proposed MSSA can effectively to optimal chillers loading, and the use of metaheuristic optimization algorithms provides a new approach to investigate efficient building energy-saving methods. However, we also found that MSSA has a lot of room for improvement in terms of robustness. In our future work, we will utilize new strategy to enhance MSSA to solve OCL problems, and try to apply MSSA to other fields of engineering optimization problems. In addition, we will also be committed to the research of multi-objective MSSA algorithm, and establish a multi-objective energy optimization model based on economy, energy efficiency, carbon emission, etc., to provide new ideas and methods for other energy optimization problems.Data will be made available on request.Xiaodan Shao: Data curation, Formal analysis, Investigation, Methodology, Software, Validation, Visualization, Writing – original draft. Jiabang Yu: Formal analysis, Investigation, Validation, Writing – original draft. Ze Li: Formal analysis, Investigation, Software, Writing – original draft. Xiaohu Yang: Conceptualization, Funding acquisition, Methodology, Project administration, Supervision, Writing – review & editing. Bengt Sundén: Conceptualization, Supervision, Writing – review & editing.The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.


It is no secret that graphene, a two-dimensional single-layered carbon atom crystal lattice, has drawn tremendous attention due to its distinct electronic, surface, mechanical, and optoelectronic properties. Graphene also has opened up new possibilities for future systems and devices due to its distinct structure and characteristics which has increased its demand in a variety of applications. However, scaling up graphene production is still a difficult, daunting, and challenging task. Although there is a vast body of literature reported on the synthesis of graphene through conventional and eco-friendly methods, viable processes for mass graphene production are still lacking. This review focuses on the variety of unwanted waste materials, such as biowastes, coal, and industrial wastes, for producing graphene and its potential derivatives. Among the synthetic routes, the main emphasis relies on microwave-assisted production of graphene derivatives. In addition, a detailed analysis of the characterization of graphene-based materials is presented. This paper also highlights the current advances and applications through the recycling of waste-derived graphene materials using microwave-assisted technology. In the end, it would alleviate the current challenges and forecast the specific direction of waste-derived graphene future prospects and developments. Graphite has an exceptionally anisotropic construction which leads to its in-plane and out-of-plane surface properties being very different . Graphenstrength . Since estrength .Graphene oxide (GO) is not a conductor. However, it can be reduced by heat processes into conductive reduced GO (rGO) . rGO is Graphene, both single layer and multilayer, can now be manufactured in a variety of ways. The layers of graphene union are fabricated through a hierarchical or base methodology . GraphitIt has become challenging in the 21st century to obtain clean, affordable, and reliable energy sources which are essential from both a financial and natural outlook. Biomass has been identified as one of the most favorable sustainable sources of energy . BiomassThermal exfoliation and carbon growth are two methods for the thermal degradation of biomass . The exf3. However, such GO sheets are bound to break into negligible round shapes of nanometers. It is striking that there is an expansion in the utilization of coal-derived nanomaterials for a variety of industrial applications [Coal is a unique carbon material that can be subdivided into lignite, bituminous coal, and anthracite . Ligniteications .2(SO4)3 as a catalyst followed by Hummers’ method oxidation into the corresponding graphite-like carbon oxides  [Currently, the strategy for reprocessing coal into graphene is to initially convert huge molecule coal into an antecedent carbon source prior to synthesizing graphene. The precursor carbon source can be gaseous or a particular form . Primer arately) ,58.Malaysia is an emerging nation that relies on modern efficiency as one of its monetary donors. Different types of wastes are produced in industrial processes, including chemical effluents, industrial plants waste, paper waste, metals, concrete, sludge, electronic devices wastes, etc. . A numbe2SO4) and graphite (30 mL & 1 g) were mixed homogeneously in an ice bath for 30 min during the synthesis cycle. A total of 5 g of potassium permanganate (KMnO4) was added and mixed for another 15 min at temperatures below 10 °C [4 was subsequently increased from 1:3 to 1:5 to speed up the oxidation rate. From that point onward, 8 mL of ultrapure water was added dropwise for 15 min, and the temperature of the mixture was kept under 98 °C for around 60 min. Finally, the oxidation reaction was obtained by adding 60 mL ultrapure water followed by 1 mL H2O2 [A modified Hummers method was utilized to prepare to GO from graphite obtained from modern waste filtering . Concentow 10 °C . The ext mL H2O2 .Microwave radiation is electromagnetic radiation with wavelengths ranging from 0.01 to 1 m and frequencies ranging from 300 MHz to 300 GHz . Modern 2SO4 and KMnO4 [4 and the reaction is stabilized by adding hydrogen peroxide into the solution [2H4) and NaBH4, are normally utilized in substance methodology to reduce GO. Thermal treatment, on the other hand, does not require the utilization of hazardous reducing agents making it a more attractive option [Graphite or GO, is a typical wellspring of GSs, which are made from a conventional or modified Hummer’s method . Hummersnd KMnO4 . Since tsolution . A few he option . The mice option . Microwae option . Figure e option . Furthere option .Furthermore, the reduction degree of GSs was further enhanced, and the functional groups on the surface of GO are successfully lowered . There ad spacing = 0.335 nm). Later, the oxidation of graphite into graphite oxide shows middle basal (002) reflection peak moves to 11.2°, corresponding to a d spacing of 0.79 nm [Firstly, XRD is a reliable technique for the structural analysis of GO. This analysis can be used to assess the pattern/shape and crystallinity of GO . XRD als 0.79 nm . The inc 0.79 nm . Non-cov 0.79 nm . This tr 0.79 nm . Figure  0.79 nm . This di 0.79 nm ,110.XPS is one of the most common techniques used to study the relative amount of carbon, oxygen, and functional groups present in GO and electrochemically rGO (ErGO). It is an accurate technique to determine the amount of carbon and oxygen compared to elemental analysis because it is difficult to fully dehydrate a GO sample . This isThe C(1 s) and (O1 s) peaks, which are located at about 285 and 532 eV, respectively, in the XPS full scan spectra of GO and rGO, are discernible . The bonThe appearance of distinctive peaks in XPS can be used to verify that graphene has been successfully non-covalently functionalized . Accordi−1 and addresses the level of defects in the graphite. The higher the D band, the more defects in the graphite are observed [2 hybridization of carbon atoms which is located near 1580 cm−1. The 2D peak, also known as G’, represents the number of graphene layers and is observed at 2700 cm−1 [Raman spectroscopy detects the transformation in energy connected with the Stokes and anti-Stokes transitions between the scattered photons. It is a non-destructive technique that provides information on chemical structure and molecular interactions by the combination of light within the bond of material . Moreoveobserved . The G b700 cm−1 . Figure 700 cm−1 .−1 which are attributed to the vibration of adsorbed water molecules (the O-H stretching) and the skeletal vibrations from graphite domains, respectively (the sp2 aromatic C=C) [−1), alkoxy stretching vibrations (1040–1170 cm−1), O-H stretching vibrations (3300–3500 cm−1), O-H deformation peaks (1300–1400 cm−1), and carboxyl peaks (1700–1750 cm−1) [−1, the aromatic C=C peak was visible. This peak is a result of the sp2 domains in the unoxidized region of the graphite, and the vibration that is produced there is known as skeletal vibration [Pristine graphene is a carbon allotrope, and no signal can be collected using FTIR. Graphite oxide exfoliation is one of the primary routes for preparing practical graphene which supports catalytic research, and the oxidation step is critical . As a retic C=C) . The oxytic C=C) . These i50 cm−1) . Notablyaks 1300–0 cm−1, aAs a result of the limits of scanning tunneling microscopy (STM), such as the requirement for conductive examples, atomic force microscopy (AFM) was created in 1985 . AFM is Since it can rapidly examine/imagine the morphology of a huge sample, electron microscopy is broadly utilized in everyday schedule examinations . A potenTEM is best known for imaging a specimen’s morphology, a wide variety of other combined techniques are also available in TEM to extract chemical, electrical, and structural data. For instance, local diffraction patterns can be measured using the parallel electron beam of the TEM which can offer precise measurements of the crystal system and parameters . FurtherThe image of the materials’ microstructure is captured using the cutting-edge technology known as FESEM. Gas molecules have a tendency to disturb the electron beam and the emitted secondary and backscattered electrons used for imaging and FESEM is typically carried out in a high vacuum . The difOptimization of process variables and techniques to regulate the size, quality, and morphology of graphene-derived materials from waste materials.Improved synthetic concepts and methods are highly inspiring and necessitate commercial research involving renewable and biodegradable waste materials.Well-ordered oxidation/decrease and functionalization are expected for calibrating material properties, for example, band hole, electrical conductivity, and mechanical properties .Controlled graphite, GO, and rGO adjustment is in this way basic for widening the utilizations of graphene-based materials.To survey the wellbeing risk related with graphene and its subsidiaries, the poisonousness and biocompatibility of these unique carbon structures and their subordinates should be examined .Due to its extensive property, graphene preparation is a crucial area for material scientists. As a result, the scientific community should focus on advanced and novel microwave instruments which would be a great substitute of toxic and harsh chemicalsTo explore more variations that involving novel synthetic techniques, high purity GO for its mass production.There should be more consideration to lessen the cost effects of graphene derivatives.There should be more emphasis on the high yield and purity of graphene derivatives using a variety of wastes through microwave synthesis.This may also lead towards the excellence of functionalization, such as ID, 2D, and 3D graphene members, to fabricate waste materials into graphene-based structures with enhanced functionalities and high surface areas .Improving synthetic ideas and microwave approaches are remarkably motivating and requires further investigations by recycling waste materials for the optimization of parameters, such as time, power, and frequency.Further analysis of microwave synthesis and applications should be explored where the waste-based graphene derivatives can be utilized and, thus, the structures and properties can be modified as per the industrial demands.Even though scientific interest in graphene has increased for a variety of applications, there are still several significant obstacles and challenges that need to be addressed and overcome. One of the critical issues is the reproducibility of waste materials into graphenaceous materials. Improved morphological properties should be combined with procedures that are both scalable and affordable. Excitedly, there is a sustained interest in the synthesis of materials based on graphene and the evaluation of their production and fusion with other materials. Although waste precursors have been the subject of recent studies, none of them have yet been able to be marked into commercially available products. Due to graphene’s industrial significance, there is great concern about its sources and synthesis methods. These variables affect the price of graphene, and its industrial applications are constrained. The current study presents an overview of various types of wastes followed for the synthesis of graphene-based materials. Graphene creates wonders with its intriguing qualities and great attention from researchers all over the world. GO isolation has been established for more than ten years. However, the process is a continual exploration of variations involving novel synthesis techniques, and highly pure GO for its mass production and commercialization. The major attention is to have a process that is cost effective and economical. The literature is rich with important process parameters, their optimizations, and the synthesis of GO from a variety of waste which is useful for a wide range of applications. We have narrated literature that lists the synthetic routes for GO, particularly microwave synthesis, as well as different characterization methods. Although waste biomass-inferred graphene is one more encouraging material with various applications, its synthesis method is still open to be explored. Accordingly, more examination is expected to exhibit the best strategy for creating graphene with the best properties and optimization. In addition, successful and cost-effective planning may lead to the use of graphene in a wide range of applications from energy to the environment. Furthermore, material progress always demonstrates a superior effect in any field. Due to its diverse properties, graphene preparation is an important area for material scientists. As a result, the scientific community will always give attention to effective and efficient graphene preparation. The improvement of graphene through green combination addresses a huge progression in graphene innovation. The cost of producing graphene in large quantities could be reduced in alternative ways using carbonaceous wastes as raw materials. The production of graphene for industrial applications should successfully utilize a variety of environmentally hazardous solid waste precursors. Since waste-derived graphene might have impurities, additional purification procedures are needed. Future research is therefore required to increase graphene production with better yield and properties.


Since the early daysof quantum mechanics, it has been known thatelectrons behave simultaneously as particles and waves, and now quantumelectronic devices can harness this duality. When devices are shrunkto the molecular scale, it is unclear under what conditions does electrontransmission remain phase-coherent, as molecules are usually treatedas either scattering or redox centers, without considering the wave–particleduality of the charge carrier. Here, we demonstrate that electrontransmission remains phase-coherent in molecular porphyrin nanoribbonsconnected to graphene electrodes. The devices act as graphene Fabry–Pérotinterferometers and allow for direct probing of the transport mechanismsthroughout several regimes. Through electrostatic gating, we observeelectronic interference fringes in transmission that are stronglycorrelated to molecular conductance across multiple oxidation states.These results demonstrate a platform for the use of interferometriceffects in single-molecule junctions, opening up new avenues for studyingquantum coherence in molecular electronic and spintronic devices. Most studiesof single molecules in junctions have focused on observing quantumphenomena, such as quantum interference (QI), in two-terminal devicesby comparing transport properties of homologous series of molecules7 and averaging data such that details of different transport mechanismsare lost, rather than manipulating and studying QI within the samemolecule.9 The ability to measure and tune the transportproperties of the same single-molecule device, by changing the gatepotential, magnetic field, and temperature, is necessary to understandhow the different transport mechanisms that arise from molecular,electrode, and molecule–electrode hybrid states all come togetherto contribute to the device conductance. Studies of this nature cananswer questions such as: How can electron transmission be shown tobe phase coherent? What molecule–electrode coupling regime10 is required for this to be the case? The understandinggained from these studies feeds into one of the ultimate goals ofsingle-molecule electronics, which is to integrate molecules and nanoribbons,11 one-by-one, into solid-state devices with somefunctionality that exploits the quantum properties of an individualmolecule.The ability to harness and exploit coherenceat the nanoscale iscrucial for emerging quantum technologies being developed in researchareas across engineering, chemistry, and condensed-matter physics,and it may also play a role in electron transfer in biomolecular systems.14 The spatial confinement and longcoherence lengths of electrons in graphene mean that devices displaya range of quantum-coherent features, such as electronic Fabry–Pérot(FP) interferometry.16 Furthermore, there are establishedroutes to interface molecules, including biomolecules, with graphenethrough π-stacking interactions17 to generate electrostatically gated single-molecule graphene junctions,enabling temperature and magnetic-field-dependent measurements witha high operating frequency.18 This is facilitatedby the weaker screening by graphene of the gate electric field, comparedwith bulk 3D metallic electrodes used in traditional molecular junctions,20 and by the electronic band structure of graphene, which makes itpossible to differentiate between the contributions to transport fromgraphene and molecular states.21Graphene devices are an ideal platform for investigatingphase-coherentphenomena in charge transport.Vsd), gate voltage (Vg), andtemperature exhibit a wide range of QI phenomena such as FP and Kondoresonances. Our results demonstrate that an 8 nm porphyrin octamernanoribbon sustains phase-coherent electron transmission that canbe tuned electrostatically and is highly oxidation-state dependent.Overall, we reveal a comprehensive picture of different quantum-coherentphenomena that arise in molecule–graphene junctions.In this work, we study charge transport through porphyrin nanoribbon–graphenedevices at cryogenic temperatures 1a and deNi-FP8) device  complexes,25 but in this case nickel was used to facilitate the synthesis andto reduce the energy of the HOMO, thus enhancing the chemical stabilityof the ribbon. The porphyrin octamer Ni-FP8 is diamagneticin its neutral oxidation state, because the nickel(II) ions have alow-spin d8 electron configuration. The molecules are solubledue to the presence of bis(trihexylsilyl)phenyl groupsthat decorate the porphyrin units; these groups prevent aggregationand improve the likelihood of obtaining transport data dominated bya single nanoribbon.26 The fused porphyrinoctamer core is functionalized with tridodecyloxypyrene (TDP) anchorgroups at both ends, to bind to the graphene source and drain electrodesvia π-stacking. An idealized view of the junction is displayedin We focus on charge transportthrough an edge-fused nickel(II) porphyrinoctamer  device 1a. Ni-FP2 substrate by electron-beam evaporation and then covered withan ALD-grown dielectric layer of 10 nm of HfO2. The thin,high-κ HfO2 layer provides a large electrostaticcoupling between the chemical potentials of the molecular states andthe gate potential, typically larger than 0.1 eV/V.27 The large coupling, combined with a dielectric breakdownvoltage for the HfO2 of Vg ≈± 5 V, allows multiple molecular charge states to be measured.Platinum source and drain electrodes are deposited next, and thenCVD-grown graphene is transferred onto the whole substrate. The grapheneis patterned into a bow-tie-shaped constriction .30 The source-drain electrode spacing can be determined by fitting the Isd–Vsd trace, measured after electroburning, tothe Simmons model.31 This typically yieldsvalues of 1.0–2.5 nm;17 however,for the Ni-FP8 devices described below, the tunnelingcurrent was too low for Simmons fitting, leading us to estimate alower bound for the gap size of >2.5 nm for these devices.Feedback-controlled electroburning isused to convert the bowtie-shapedconstrictions into nanometer-spaced graphene electrodes (see the Ni-FP8 in toluene(1 μM) directly onto the devices. The tunneling current throughthe devices before and after the drop-casting are compared, and onlywhen there are new regions of high tunneling current after depositionthat are at least an order of magnitude higher than the baseline arethe devices wire-bonded and cooled for detailed measurements .We discuss transport through two Ni-FP8 devices in themain text (device 1 and device 2) and also present data from two shorterporphyrin oligomers shown in Supporting Information Figure S3-6: one with a zinc porphyrin monomer (Zn-P1) one with an edge-fused porphyrin trimer (Zn-FP3) thatdisplay the same evidence for phase-coherent charge transport outlinedbelow.The porphyrin nanoribbons are interfaced with the electroburntnanogaps by drop-casting a solution of Gsd = dIsd/dVsd) of the porphyrinnanoribbon Ni-FP8 device 1, measured at 4.2 K, displaysseveral Coulomb diamonds and associated resonant tunneling regions, as this diamond has a large addition energy, at Eadd = 0.7 eV. There is a zero-bias conductancepeak in the neighboring diamond between Vg ≈−0.7 V and Vg ≈−1.4 V. This peak is consistent with the experimental signatureof a Kondo resonance (discussed in detail below) that results fromscreening of an unpaired spin on the nanoribbon by electrons withinthe graphene electrodes.35 The observation ofa Kondo resonance is consistent with assigning this smaller diamondto the odd N – 1 state , confirming that the larger diamond at Vg = 0 is the even N state,as the number of electrons on the molecule differs by one betweenadjacent Coulomb diamonds.27 The two sequentialtransport regions with broad edges that flank the N – 1 diamond are then the N – 1/N charge transition (Vg ∼−0.7 V) and the N – 2/N – 1 transition (Vg ∼ −1.4V). Finally, the highly conductive region at positive Vg is the N/N + 1 chargetransition.The full differentialconductance map ( regions1c. We cakBT = 0.4 meV), thepoor definition in the boundaries of the Coulomb diamonds  is attributed to lifetime broadening that results fromintermediate molecule–electrode coupling, a regime consistentwith the appearance of the Kondo resonance.37 The broad diamond edges suggest that there are large regions wherethe molecular charge state is not well defined, in contrast with adevice in the weakly coupled regime (commonly observed for moleculesπ-stacked to graphene electrodes38), where conductance occurs only within the sequentialtransport regions separated by Coulomb diamonds. In Vsd = 10 mV (awayfrom the Kondo peak), the conductance is not completely suppressed.Instead, the conductance remains above the noise except within the N diamond, indicating that there are significant contributionsfrom off-resonant phase-coherent transport around the N – 1/N and N – 2/N – 1 transitions of the intermediately coupled nanoribbondevice.36 As the hybridization betweenmolecule and electrode increases with electronic coupling, transportthrough an intermediately coupled molecular junction can only be fullyunderstood by considering the entire graphene–nanoribbon–graphenesystem. We discuss the mixture of effects that arise from this holisticapproach by initially focusing on phase-coherent transport withinthe graphene channel and then on molecular transport.Considering the low measurement temperature . The periodic featuresare more obvious in the second derivative of the current (dIsd2/dVsd2) map, which displays a “checkerboard”pattern . Similar periodicities have been observedin transport measurements through graphene and are attributed to theformation of an electronic FP interferometer within the graphene channel.39 By analogy with an optical FPcavity formed from a pair of partially reflective mirrors, two potentialsteps, induced by doping, define an electronic FP cavity. Consideringthe resonances within a one-dimensional FP cavity, our measured periodicenergy spacings of ∼4 and ∼8 meV correspond to cavitylengths of L = hvF/(2E) = 900 and 450 nm, approximating a Fermi velocity of vF = 1.8 × 106 m/s by adjustingthe value for CVD graphene on SiO2 (vF = 2.49 × 106 m/s) for the larger dielectricconstant of HfO2.41 The length scales of900 and 450 nm are 2 orders of magnitude larger than those that wouldbe associated with reflections within the Ni-FP8 nanoribbonitself, confirming that the QI pattern is dominated by reflectionswithin the graphene leads.The energy spacings betweenadjacent conductance oscillations in 2 is p-doped42  region .The FP cavity is formed within grapheneby doping induced by interactionswith the underlying substrate. Graphene transferred onto HfOp-doped and p+-doped graphene regions generate outer potential steps,and the molecule/electrode interfacial tunnel barriers also act aspotential steps at which electrons are reflected or transmitted. Therefore,there are two FP cavities formed over ∼450 nm (correspondingto 8 meV energy spacing) that occur between a p/p+ potential step and a molecule/electrode tunnel barrier oneach side of the device. The reflections inside these cavities areindicated by red arrows in 15 with reflectionoccurring on the outer p/p+ potentialsteps in the graphene, as indicated by blue arrows in The interfacesbetween Ni-FP8 device  also displaythe same features, with periodic interference patterns that have lowergate couplings than the molecular states. The second Ni-FP8 device has a stronger molecule–graphene coupling Γthan device 1, increasing the conductance, and consistent with a weakercoupling of the more hybridized molecular states to the gate potential. All four devices share the propertyof having stronger conductance fluctuations when the molecule is oxidizedto the N – 1 state , which can be inferred from the regions of resonant transportthat are higher in conductance,28 whereasthe opposite is true for device 2 (ΓD > ΓS).The data for asecond, more strongly coupled, hvF/L (4 meV), the FP interference patterndisappears, probably because the thermal energy becomes larger thanthe FP spacing , and the coherence length within the graphene channeldecreases with increasing temperature.45 Furthermore, the Kondo resonance, clearly visible as a zero-biasconductance enhancement within the N – 1 state(Ni-FP8+) at 5 K, as shown in Ni-FP8 is oxidized to the N – 1 state(Ni-FP8+), an electron is removedfrom the molecular π-system and the resulting spin (S = 1/2) and charge are delocalized over the molecule, formingthe Kondo state with electrons in the graphene electrodes. As thermalenergy increases toward the binding energy of this many-body molecule-electrodestate, the zero-bias conductance will decay. The characteristic temperature-dependenceis parameterized by the Kondo temperature (TK), the temperature when the conductance is half its valueat 0 K, i.e., G(T = TK) = 0.5 × G(T =0). We extract the Kondo temperature of our device at Vg = −0.85 V by fitting the temperature-dependenceof the zero-bias conductance to the usual spin-1/2 models is an empirical parameter.46Next, we describethe temperature and charge-state dependence of device conductancein more depth. As shown in TK = 18 ± 1K and s = 0.30 ± 0.04 from the fit in TK at this gate voltage from the fwhm of theresonance, whichgives a similar value of 17 ± 1 K (see N – 1 transition (0.12eV for Ni-FP8 device 1) and exponentially on the molecule–electrodecoupling, hence its association with the onset of intermediate coupling.10 A typical gate-dependent measurement of a Kondoresonance would give a smooth conductance decay as Vg is detuned from the resonances,described by the Haldane relation.46 However, as TK also depends on the graphene density of states,47 our gate-dependent measurement of the Kondopeak displays conductance oscillations, as opposed to a smooth valley,within the N – 1 state, in line with a previousobservation of transmission through a coupled Kondo/FP system.48 The Kondo resonance is not visible in the N + 1 state, probably because the gate voltages in whichthe molecule is in the singly reduced state are close to the Diracpoint of graphene , or alternativelya weaker molecule–electronic coupling to that charge statedue to transport through more localized orbitals , or a combinationof both. Aside from the Kondo behavior, the conductance of the device,including at the Coulomb peaks, show a weak temperature dependence.We calculate  1 K see 3b, insetNi-FP8 constitutesone ofthe longest molecular systems (8 nm) with well-defined anchor groupsover which phase-coherent transport has been measured, consistentwith previous conductance measurements that demonstrate low or negativeattenuation factors for edge-fused porphyrin oligomers.49 This is a property of the molecule and results from strong couplingbetween the fused porphyrin units and subsequent delocalization ofelectronic wavefunctions over the whole π-system, as corroboratedby measurements of the optical gap.50 Anothermolecular-structure-dependent charge transport property is the presenceof the 2 Coulomb peaks (N – 1/N and N – 1/N – 2)at negative Vg that are due to oxidationof the electron-rich TDP anchor groups, coupled through the porphyrinnanoribbon.51 Thus, the two Ni-FP8 devices share this relatively similar feature with Zn-P1 and Zn-FP3 , with a different addition energy dependent on thelength of the fused porphyrin core.The porphyrin nanoribbon Ni-FP8, as well as depending on Vg within each diamond due to the effect of thegate potential on the level alignments (–7.0G0) of our experimental setup away from the Coulomb peaks,but it is 10–4.7G0 inthe mid-gap of the N – 1 state and 10–5.8G0 in N – 2, indicating that in the oxidized states of the Ni-FP8, off-resonance coherent transport is more efficientthan in the neutral state. The same trend is observed in Ni-FP8 device 2, but with higher conductance values due to stronger molecule–electrodecoupling, with values of 10–3.6G0, 10–3.2G0, and 10–3.4G0 for N, N – 1, and N – 2 charge states, respectively. The difference in magnitudesbetween device 1 and device 2 highlights the continuing need for strategiesto be developed to control Γ so that devices with specific properties(rather than trends) are able to be fabricated on a larger scale.It is striking that theoff-resonance conductance and the appearanceof the phase-coherent FP interference pattern is strongly dependenton the charge state of ignments 3d. For t52 with a similar conductance enhancement of ∼100 reportedfor an edge-fused trimer upon oxidation, similar to the ratio measuredin our device that has a quite different geometry. Furthermore, ourresults are consistent with the general observation in single-moleculeconductance measurements that upon oxidation or reduction, a conductanceenhancement is observed in the odd-electron number state.55 As with these previous studies, we can utilize a combination ofDFT and quantum transport theory58 to calculate the phase-coherenttransmission  (orange curve), when eight Cl atoms are present, isplotted in T(E) is observed overa large energy range close to Fermi energy upon oxidation, primarilydue to a shrinking of the HOMO–LUMO gap . As shown in Supporting Information Figure S5-4, the increasein transmission upon oxidation is robust to changes in the portionof the nanoribbon overlapping with each of the graphene electrodes.The introduction of Cl atoms creates extra features (spikes) in thetransmission that does not affect the increasing trend . The calculatedtransmission spectrum based on coherent transport theory shows excellentagreement with the experimental measurements of Ni-FP8, particularly at higher temperature.Charge-state-dependent conductance has been observed previouslyin STM-BJ measurements of fused porphyrin oligomers,60Ourresults show that phase-coherent electron transport persiststhrough a fused porphyrin nanoribbon even over a molecular lengthof 8 nm. The mechanism is confirmed by measurement of charge-state-dependentinterference fringes from FP resonances that extend over a graphene–molecule–graphenecavity, along with the observation of temperature-independent off-resonantconductance in multiple oxidation states of the molecule. The persistenceof phase-coherent transport behavior across such extended molecularsystems within the intermediate molecule–electrode couplingregime is important for understanding and designing systems for efficientlong-range electron transport. The gate electrode can be used to preciselytune the thermodynamic driving-force of electron-transfer reactions,and therefore this platform could be extended to study long-rangeelectron transmission and transfer in biological systems appropriatelyfunctionalized to interface with graphene.4 therefore providing an interesting research direction inexploring nanoribbon–graphene hybrid devices for quantum informationprocessing.The study of coherent electron transport through a molecularnanoribbonembedded in a FP cavity opens up prospects for all-electrical interferometricmeasurements between graphene and molecular pathways, where both transmissionmagnitude and phase through the device can be determined experimentally.This platform could enable the readout of molecular topological qubits,for which nanoribbons have demonstrated potential,Ni-LP8Br  in dry 1,2-dichloroethane , a suspension of AuCl3  and AgOTf  in dry DCE (2.5mL) was added dropwise, and the reaction mixture was stirred at 25°C for 15 min. After that, a suspension of AuCl3  and AgOTf  in dry DCE (0.5 mL) was added dropwise to the reaction mixture,and the reaction was monitored by UV–vis–NIR spectroscopywith CH2Cl2 + 1% triethylamine as the solvent.After completion, triethylamine (1.0 mL) was added to the reactionmixture. The resulting mixture was purified by flash column chromatographyon silica gel using pentane/CH2Cl2 (9:1) asthe eluent to give product Ni-FP8Br .To a solution of Ni-FP8Br , Pd(PPh3)4 , and CuI  in dry toluene(0.5 mL) and diisopropylamine  was degassed by threefreeze–pump–thaw cycles. A solution of 1,3,6-tris(dodecyloxy)-8-ethynylpyrene in dry toluene (0.5 mL) and DIPA(0.5 mL) was degassed by three freeze–pump–thaw cyclesand transferred to the reaction mixture under argon. After that, themixture was stirred at 50 °C under argon for 2 h. Then, a degassedsolution of 1,3,6-tris(dodecyloxy)-8-ethynylpyrene  in dry toluene (0.5 mL) and DIPA (0.5 mL) was added to thereaction mixture and the mixture was stirred at 50 °C for another20 h. After reaction, the resulting mixture was separated by flashcolumn chromatography on silica gel using pentane/CH2Cl2 (1:1) as eluent, followed by size-exclusion chromatography(Biorad Bio beads SX-1) with toluene/pyridine (99:1) as the eluentto give the crude mixture. The crude mixture was further subjectedto recycling GPC with toluene/pyridine (99:1) as the eluent to separatethe desired product Ni-FP8 . See Supporting Information Section S2 for the reactionsand Supporting Information Section S2 forcharacterization data of intermediate compounds.A mixture of Ni-FP8 devices was fabricatedusing the following procedure.On a degenerately n-doped silicon wafer with a layer (300 nm thick)of thermally grown silicon dioxide (SiO2), a local gateelectrode (3 μm wide) was defined by optical lithography withlift-off resist and electron-beam (e-beam) evaporation of titanium(5 nm thick) and platinum (15 nm thick). A layer (10 nm) of hafniumdioxide (HfO2) was then deposited using atomic layer deposition(ALD). Next, source and drain contact electrodes separated by a 7μm gap  were also defined by optical lithographywith lift-off resist and e-beam evaporation of titanium (5 nm thick)and platinum (45 nm thick). The procedure for the fabrication of substratesused for Zn-P1 and Zn-FP3 has been publishedpreviously.44The substrate used for 4)2S2O8) solution(3.6 g in 60 mL water) for 4 h, after which the PMMA-protected graphenewas transferred three times to Milli-Q water and scooped up usingthe substrate. Air bubbles were removed by partly submerging the samplein 2-propanol (IPA). The sample was dried overnight and baked at 180°C for 1 h. The PMMA was then removed in hot acetone (50 °C)for 3 h.A layer (600 nm) of poly(methyl methacrylate)(PMMA) (with a molecular weight of 495 kDa) was spin-coated onto chemicalvapor deposition (CVD)-grown graphene (purchased from Grolltex) oncopper. The copper was then etched in aqueous ammonium persulfate((NHSupporting Information Section S1.The Z-shaped graphene tape with bow-tie shaped structurewas patterned by e-beam lithography (EBL) with a bi-layer lift-offresist (PMMA495 and PMMA950) and thermal evaporation of aluminum (50nm thick). The Z-shaped graphene pattern was used so the inner grapheneleads are coplanar with the bowtie structure see 1b,c, red–7G0). The empty nanogaps were characterized by measuring a currentmap as a function of bias voltage (Vsd) and gate voltage (Vg) at room temperaturein order to exclude devices containing residual graphene quantum dots;17 only clean devices were selected for furthermeasurement.Graphene nanogaps were preparedby feedback-controlled electroburningof the graphene bow-tie shape until the resistance of the tunnel junctionexceeds 1.3 GΩ (10Supporting Information Section S3.The solution ofthe porphyrin nanoribbon (1 μM in toluene) was drop-cast onelectroburnt graphene electrodes and allowed to dry in air. Only devicesthat showed clean current maps before molecule deposition were selectedfor further measurements. Thus, new signals appearing after moleculedeposition can be attributed to transport through molecular junctions.Then, the chip containing molecular devices was connected to a chipholder via wire bonding, loaded in Oxford Instruments 4K PuckTester,and cooled down to cryogenic temperature for detailed measurements.The current maps and differential conductance maps of before and aftermeasurements can be found in 56 with a local density approximation LDA functional, a double-ζpolarized basis, a cutoff energy of 200 Ry, and a 0.04 eV/Å forcetolerance. From the Hamiltonian and overlap matrices of the DFT calculationof the junction, Gollum57 calculates thetransmission coefficient Tnm(E) between scattering channels n and m in the electrodes, from whichthe transmission coefficient G is the (retarded)Green’s function of the junction and Γi is the imaginary part of the self-energy of electrode i. The electrical conductance is obtained fromEF is theFermi energy of the device, G = G0T(EF). In the presence of Cl atoms, spin-polarized calculations werecarried out to obtain the transmission coefficients T↑, T↓ for the two different spins,from which the total transmission coefficient Geometricaloptimizationswere carried out using the DFT code SIESTA,7of ref , this is


In this work, an enhanced tunable microwave phase shifter is presented. The phase shifter consists of three short circuited stubs and a tapered line. The stubs are connected to graphene pads. Graphene’s tunable conductivity is varied by a DC voltage. This in turn causes a reactance variation at the input of the tapered line, which causes a phase variation. The physical parameters of the stubs are optimized for a maximum reactance variation by the help of analytical models, circuit and full wave simulations. Measurements of an optimized prototype are performed and a dynamic phase variation of  Carbon-based materials have caught significant attention in recent years. This is due to their exciting mechanical, electrical, optical and morphological characteristics ,2. GraphThe prototype is designed on the Rogers 3035 RF dielectric substrate . The dielectric substrate has a dielectric constant, In the phase shifter fabrication, graphene flakes are deposited as thin films. The Raman spectrum is measured in order to estimate the graphene’s quality ,28. The Graphene flakes size and surface morphology was investigated by a field emission scanning electron microscope  . The acqThe phase shifter consists of three short circuited stubs connected to graphene pads and a tapered line. Graphene is modeled as a resistor with resistance equal to An ideal phase shifter should have a maximum phase variation and a minimum insertion loss variation. To this aim, the variation of the imaginary part of the input impedance The impedance at the input of the tapered line section can be simplified by using a transmission line model. The input impedance at the beginning of the tapered line is given by The lengths of the stubs can be optimized for the range of graphene resistance variation in order to maximize Equation . A plot Once the length of the stubs is optimized by this analytical analysis, each stub is connected to a two-port transmission line. The topology is simulated for different values of graphene resistances by the help of AWR Microwave Office . For the sake of simplicity, the transmission lines are modeled as ideal, lossless transmission lines, the tapered line section is modeled as a non-homogeneous linear taper and graphene is modeled as a tunable resistor. The two-port transmission line has a characteristic impedance of The reflection coefficient see  is arounThe measured amplitude and phase of the two port scattering parameters at different DC voltages are shown in An enhanced tunable phase shifter based on graphene is presented. The phase shifter has the capability to dynamically tune its phase by a variable DC voltage. The phase shifter is based on three short circuited stubs connected to a tapered line and a two-port transmission line with the help of graphene pads. In order to maximize the phase shifting capability of the phase shifter and to reduce the insertion loss, the imaginary input impedance at the tapered line section needed to be maximized, keeping the real part as low as possible. Analytical computations are performed in order to choose appropriate lengths and widths of the stub. The values achieved are verified by further analysis, performed with circuit models and finite element simulations. This is followed by the fabrication and measurement of prototypes with the optimized parameters. The optimized design has resulted in enhanced phase-shifting capabilities with minimum insertion loss variation.


Graphene aerogels are of high interest nowadays since they have ultralow density, rich porosity, high deformability, and good adsorption. In the present work, three different morphologies of graphene aerogels with a honeycomb-like structure are considered. The strength and deformation behavior of these graphene honeycomb structures are studied by molecular dynamics simulation. The effect of structural morphology on the stability of graphene aerogel is discussed. It is shown that structural changes significantly depend on the structural morphology and the loading direction. The deformation of the re-entrant honeycomb is similar to the deformation of a conventional honeycomb due to the opening of the honeycomb cells. At the first deformation stage, no stress increase is observed due to the structural transformation. Further, stress concentration on the junctions of the honeycomb structure and over the walls occurs. The addition of carbon nanotubes and graphene flakes into the cells of graphene aerogel does not result in a strength increase. The mechanisms of weakening are analyzed in detail. The obtained results further contribute to the understanding of the microscopic deformation mechanisms of graphene aerogels and their design for various applications. The study of tree-dimensional 3D) porous graphene nanostructures is one of the hot issues in different industrial applications. Such structures as aerogels and crumpled graphene are lightweight and low-cost and have high porosity and high strength ,2. AmongD porous One of the most promising methods is the fabrication of GA from graphene oxide, which is synthesized and dispersed in distilled water ,13. GrapThe mechanical properties of graphene aerogel are of high importance for stretchable electronics, wearable devices, and smart manufacturing. Some of the disadvantages of graphene aerogel are the volume shrinkage and the deformation of porous skeletons, which affect the mechanical properties ,28,29. GDespite the fact that numerous works have been conducted to date on the mechanical behavior of GA, there is still a lack of understanding of the effect of morphology on its strength ,34,35,36In the present work, the mechanical properties of three different morphologies of GA under tension are studied by molecular dynamics simulation. The high stretchability is studied and analyzed for tension along different lattice directions. The effect of high temperatures on the strain and deformation behavior is studied.z direction, while the tension along lamella is equivalent to the tension along the x direction.aerogels ,41,42,43l = 16 Å, and the number of atoms is 2500.The morphology, which will be addressed as a honeycomb is widespread and has been previously fabricated and studied in numerous works ,39,44,45a = 20 Å, b = 11 Å  can also be called bridges, while long graphene nanoribbons  can be called lamellae. The difference with the structure considered in [One of the possible morphologies of GA, which will be addressed as an arrow honeycomb, is also studied for comparison. Here, the main parameters are ussed in ,50. The dered in  is the cAll calculations are performed using the LAMMPS ,52,53 soDespite the slight uncertainty of the AIREBO application for the study of mechanical properties of GA, there is plenty of work devoted to this issue, where simulation is conducted with AIREBO. Based on such works as ,39,63,64In the present work, the tensile behavior of GA is analyzed at two temperatures, 300 and 1000 K, and thus the limitation of the AIREBO potential for the simulation of such effects needs to be discussed. In , ReaxFF Periodic boundary conditions are applied in all directions. A Nose–Hoover thermostat is used with a target temperature, a constant number of atoms, and standard velocity–Verlet time integration with a timestep of 1 fs. OVITO is used to visualize the atomistic simulation data. At first, the energy minimization at 0 K is conducted to reach an equilibrium state of the system.x or z axes, and the corresponding stress components are calculated as it is implemented in the LAMMPS package. Stress and energy per atom are also calculated using LAMMPS. At low temperatures, it has been previously shown that the strain rate has only a slight effect on the fracture strength and fracture strain for graphene [To study the deformation behavior, uniaxial tensile stress is applied along the graphene . Thus, tThe structural changes during the relaxation were analyzed: the structural parameters were changed insignificantly. For example, sharp edges were smoothed for re-entrant and arrow honeycomb. For arrow honeycomb, cells transform to a more square shape.The compressive behaviors of such honeycombs under in-plane uniaxial compression was previously studied ,38. It wx-axes for honeycomb . These bonds are almost aligned with the loading direction and thus mostly strained. These bonds (and others of the same orientation) are continuously changing from 1.41 Å to 1.75 Å, while bond c changes from 1.41 Å to 1.55 Å. Very similar results were obtained previously [Considerable structural changes occur at sed CNTs . Thus, ueviously .As can be seen from Note, in  mechanicFor re-entrant honeycomb, just part of the stress–strain curves are presented with critical points labeled as 1–7. As can be seen from the structure snapshots during tension, re-entrant cells begin to open even at the first deformation steps. At ained in  and furtained in . At ε = mentally . Up to εAs was shown in , for sucz, which was smoothed during relaxation. During tension, the walls of the cells can oscillate, and arrows transform into squares. Cell walls aligned along x are mostly stressed since they are oriented along the loading direction.As can be seen from z-axis. Until z-axis are unstrained . These bonds are almost aligned with the loading direction and thus mostly strained. These bonds (and others of the same orientation) are continuously changing until c almost does not change until d, e, f, which are analogous to a, b, c, are named differently to distinguish bonds on the cell walls that are differently oriented. These bonds are analyzed on the walls in the loading direction. They slightly change between 1.38 and 1.42 Å due to the thermal fluctuations. Both walls of the cell after relaxation are equal to about 19 Å; however, during tension the cell wall along the z-axis did not change, while the length of the cell wall along the x-axis increased to 26 Å (increased to 36%).An analysis of the interatomic bonds showed that on the cell walls along the The deformation mechanisms at 300 K and 1000 K are the same. Strength is decreased just due to the thermal fluctuations.x- and z-axis. x- and z-axes as the function of applied strain for (a) honeycomb; (b) re-entrant honeycomb; and (c) arrow honeycomb.We also have studied the effect of the direction of the applied strain on the deformation behavior of the structure. The uniaxial strain was applied along the x-axis) and zigzag (z-axis) was found. It was also previously shown for the same GA that when stretched along the armchair direction, mechanical strength and failure strain decrease with the increase in sidewall width [x-axis, while the zigzag coincides with the z-axis. Note that for re-entrant honeycomb, only part of the stress–strain curve for tension along the x-direction is presented, which is explained above. However, the continuation of the stress–strain curve for the re-entrant honeycomb is shown by a dashed line for reference.ll width . Moreovez, both structures deformed similarly: the straightening of the walls of honeycomb cells took place with almost no stress concentration; then, stresses over the walls aligned with the tension direction began to increase, which was followed by fracture . For the honeycomb, the length of the wall is about 32 Å, and for the re-entrant honeycomb the length is 22 Å; thus, strength and failure strain for honeycomb GA are higher. Cells for both structures are presented in two structural states for comparison. The same deformation mode with the elongation of the honeycomb cell to the rectangular cell was shown in [For tension along shown in ,67. The shown in  that thex and z directions almost coincide. During relaxation, the cells of the arrow honeycomb change their shape into almost square cells. The size of the cell wall along x and z is 20 Å. Thus, the structure is isotropic in the x is explained solely by thermal fluctuations. The opened cells of arrow GA are very similar to the opened cells of re-entrant GA.For arrow GA, the stress–strain curves for uniaxial tension along the The addition of CNTs and graphene flakes in the present form does not result in an increase in strength or failure strain. Deformation occurs very differently due to the presence of CNTs and graphene: with these elements, honeycomb cells can transform into square, hexagonal, or collapsed shapes. However, additional studies are required to understand how to control morphology to obtain better mechanical properties of honeycomb. However, reinforcing elements increase the structure density, which results in a strength decrease. As was shown in , the strThese morphologies of GA composed of cells not only possess high strength but also show good elasticity during tension. The elasticity of such structures is of significant interest for the materials used in applications where they are expected to deform periodically while still retaining most of their original strength and stiffness. Let us discuss the elastic behaviors of the considered GA in detail.x direction of the arrow honeycomb, the structure remains elastic until the first fracture starts. From the beginning of loading until fracture, there was no change in the atomic configuration, confirming the elasticity of GA. Before z direction, deformation behavior is very close. Until the sharp increase in stress at During tension in the ions see . This inA detailed study has been performed to investigate the mechanical properties of graphene aerogels of a different morphology using MD simulations. The tensile loading of graphene aerogels is analyzed for two temperatures, 300 and 1000 K. Loading is conducted along two lattice directions to overview the structure anisotropy.At the first deformation stages, the walls (bridges) of the honeycomb cells can easily bend or oscillate in the direction normal to the tension direction. With the strain increase, the stress concentration near the junctions of the honeycomb cell and over the walls oriented along tensile direction can be observed. The strength of GA is considerably dependent on the length of the walls of honeycomb cells and their density. The deformation of the re-entrant honeycomb is defined by the opening of the cells during the first deformation stages. Further, the re-entrant honeycomb transforms into the conventional honeycomb. Very similar deformation mechanisms were found for honeycomb under compression . The newFor comparison, aerogel carbon matrices were reinforced by graphene flakes and carbon nanotubes; however, this distribution of the reinforcing elements did not lead to an increase in the strength and failure strain. The search for the new morphologies with special distribution of the reinforcing elements is further required. The obtained results further contribute to the understanding of the microscopic deformation mechanisms of GA and their design for various applications.


Drosophila Kenyon cells of the mushroom body (MB), a circuit with structural characteristics reminiscent of vertebrate cortical networks. This model recapitulates several aspects of dementia-relevant pathological features including age-dependent neuronal loss, nuclear depletion and cytoplasmic accumulation of TDP-43, and behavioral deficits in working memory and sleep that occur prior to axonal degeneration. RNA immunoprecipitations identify several candidate mRNA targets of TDP-43 in MBs, some of which are unique to the MB circuit and others that are shared with motor neurons. Among the latter is the glypican Dally-like-protein (Dlp), which exhibits significant TDP-43 associated reduction in expression during aging. Using genetic interactions we show that overexpression of Dlp in MBs mitigates TDP-43 dependent working memory deficits, conistent with Dlp acting as a mediator of TDP-43 toxicity. Substantiating our findings in the fly model, we find that the expression of GPC6 mRNA, a human ortholog of dlp, is specifically altered in neurons exhibiting the molecular signature of TDP-43 pathology in FTD patient brains. These findings suggest that circuit-specific Drosophila models provide a platform for uncovering shared or disease-specific molecular mechanisms and vulnerabilities across the spectrum of TDP-43 proteinopathies.Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) comprise a spectrum of neurodegenerative diseases linked to TDP-43 proteinopathy, which at the cellular level, is characterized by loss of nuclear TDP-43 and accumulation of cytoplasmic TDP-43 inclusions that ultimately cause RNA processing defects including dysregulation of splicing, mRNA transport and translation. Complementing our previous work in motor neurons, here we report a novel model of TDP-43 proteinopathy based on overexpression of TDP-43 in a subset of The online version contains supplementary material available at 10.1186/s40478-023-01656-0. TARDBP, the gene encoding TDP-43, have been identified as causative of ALS [Characterized by extensive overlap in cellular pathology, genetic mutations, and molecular markers, amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) comprise a neurodegenerative disease continuum affecting multiple subsets of neurons in the spinal cord, motor cortex, cingulate cortex, and frontal and temporal lobes –4. Notabe of ALS –9 while e of ALS , 11. Alte of ALS .Substantiating its significance as a marker of pathology, TDP-43 has also been identified as a component of cytoplasmic aggregates in dementias other than FTD, including Alzheimer’s disease (AD) –15, AD wWild-type TDP-43 is localized primarily to the nucleus where it regulates RNA transcription  and spliDrosophila melanogaster) genome contains a homolog of TARDBP, namely TBPH, which displays sequence similarity and functional overlap with human TDP-43 [Following the identification of TDP-43 as a protein of pathological interest in numerous neurodegenerative diseases, model organisms have been critical to understanding the normal biological functions of TDP-43 and how disruption of these functions leads to disease. Despite having diverged from a common ancestor with vertebrates over 600 million years ago , the frun TDP-43 , 43.Drosophila have recapitulated the locomotor phenotypes, motor neuron dysfunction, and shortened lifespan [In addition to extensive overlap in neurogenetic homology, circuit-level conservation in the central nervous system has also been demonstrated between invertebrates and vertebrates –46, makilifespan , 54, 55 lifespan , 38, 56 lifespan –60, and lifespan , 62.While TDP-43 proteinopathy has been extensively studied in motor neurons and glia as a model of ALS, dementia modeling based on TDP-43 proteinopathy in flies has been limited. This is in part because the mushroom bodies (MBs) of the fly brain, which were originally described as essential to odor learning  have onldlp mRNA, encoding the heparan sulfate proteoglycan (HSPG) Dally-like protein (Dlp)/GPC6, which we previously found to be a target of TDP-43 in fly models of ALS and is altered in human ALS spinal cords [dlp mRNA is enriched in TDP-43 complexes in MBs while Dlp protein is decreased within axons during aging, consistent with TDP-43 dependent axonal transport and/or translation deficits in an age-dependent manner. Notably, Dlp overexpression in MBs rescues TDP-43 dependent deficits in working memory consistent with Dlp being a physiologically significant target of TDP-43 in the MB circuit. These findings demonstrate that TDP-43 proteinopathy in MBs causes dementia-like phenotypes that are mediated at least in part by Dlp/GPC6, a regulator of the Wg/Wnt signaling pathway. Further substantiating the link between Dlp/GPC6 and TDP-43 pathology, we found GPC6 mRNA to be altered in FTD patient brains. Specifically, neuronal nuclei expressing STMN2 and KALRN cryptic exons associated with TDP-43 nuclear depletion show an upregulation of GPC6 mRNA when compared with nuclei that retain TDP-43 as evidenced by the presence of canonically spliced STMN2 and KALRN junctions. Lastly, we report a number of candidate targets of TDP-43 that show overlap between our fly models and patient brains, suggesting these models will be useful for studying molecular pathyways underlying distinct neuronal vulnerabilities across the spectrum of TDP-43 proteinopathies.Here we describe the development of a novel fly model of dementia based on TDP-43 proteinopathy induced by specific over-expression of wild-type or mutant human TDP-43 in a well-defined subset of MB neurons. Our model recapitulates key cellular and behavioral characteristics of human dementias with TDP-43 pathology, including age-dependent loss of nuclear TDP-43, axonal degeneration, and working memory and sleep deficits that parallel those observed in dementia patients. Using RNA immunoprecipitations we show that in MBs, TDP-43 associates with several mRNAs, a subset of which are unique to MBs while others are shared with mRNA targets previously identified in motor neurons . Interesal cords . Here weDrosophila lines used in this study and specific experiments where each line was employed can be found in the Key Resources table  w1118; TDP-43WT::YFP, (2) w1118; TDP-43WT::YFP; dlp OE, (3) w1118; TDP-43WT::YFP mCD8::RFP, and (4) w1118 in triplicate. Heads (N = 15 for each genotype) were decapitated and homogenized in 100 µl 2X Laemmli Sample Buffer (BIO-RAD 1610737) containing 5% of 2-Mercaptoethanol (Sigma-Aldrich M3148). The homogenized protein samples were boiled for 5 min in the digital Heat Block (Benchmark), and spun for 1 min. Supernatants were collected and 10 µl of protein sample was loaded in each well of precast Mini-PROTEAN TGX 4–20% gradient Gel (BIO-RAD 4561096). Following SDS-PAGE, the proteins were transferred to Nitrocellulose membrane (BIO-RAD 1620215). After transfer, the membrane was blocked in 5% nonfat milk in PBST (PBS and 0.1% TWEEN20) and incubated overnight at 4 °C with primary antibodies mouse anti-GFP Living Color  to detect TDP-43 YFP and rabbit anti-Beta-Actin  as loading control, followed by washes in TBST and incubation with secondary antibody  and  for 1 h at RT. The blot was imaged using a LICOR scanner (Odyssey®DLx) and protein bands intensities were quantified with LICOR “Image Studio Lite” , they were pooled to increase power. Where necessary, outliers were removed prior to hypothesis testing. When data did not fit the assumptions of parametric models and non-parametric analyses were used, hypothesis testing proceeded by first using a Kruskal-Wallace to test for a difference among groups, followed by pairwise comparisons using the Wilcoxon Rank Sum Test with p-values adjusted for multiple comparisons using the false discovery rate method [Statistical analyses were conducted in R [v. 4.1.2, R Core e method . SpecifiSS01276 [Mushroom body morphology was evaluated using membrane-targeted RFP (mCD8 RFP) driven by SS01276 . Adult bSS01276 , fixed flm function in R; R Core Team 2017). Residuals were normally distributed  and we therefore continued with pairwise comparisons using Tukey’s Honest Significant Difference corrected for multiple comparisons.TDP-43 cytoplasmic localization and MBN cell loss: Images were acquired using a Zeiss 880 Laser Scanning Confocal inverted microscope with a Plan-Apochromat 63x/1.4 oil DIC M27 lens. TDP-43 YFP signal intensity was quantified per unit area in the nucleus and cell body. Hoechst was used to define the nuclear boundaries, while total cellular TDP-43 signal was measured by tracing a boundary around YFP signal in the entire cell . This polygon was then dragged to an adjacent area of the brain to obtain a mean fluorescence intensity for the background in each section. In instances where the polygon was shaped in such a way that it could not be dragged to adjacent brain area, a comparably-sized polygon was drawn in the adjacent brain region. Intensity was measured in at least three sections to obtain mean fluorescence intensity in each channel (RFP or FITC) for both α/β and γ lobes of each brain. To test for differences in signal intensity within a genotype across age time points, values of signal intensity were normalized to mean signal intensity of young (1–3 days old) brains of the same genotype and lobe. Statistical analysis of change in fluorescence intensity with age was performed by comparing middle aged (~ 30 days) or old (~ 60 days) flies and young (~ 1 day) flies from the same genotype using the Wilcoxon Rank Sum test. Summary statistics can be found in Additional file 2 in size and showed 0.25–1.0 circularity. To measure mean particle size in each sample, a polygon was drawn over the visible portion of each lobe in at least three sections for each lobe. From these subsamples we calculated a mean particle size for each lobe of each brain. Summary statistics can be found in Additional file YFP particle size: The Analyze Particles function in FIJI was used to identify YFP puncta in MB lobes. Images were first contrast enhanced using a brightness/contrast enhancement  followed by thresholding using the Bernsen method in the Auto Local Threshold function with a 25–50 pixel radius depending on image quality. Particles were considered puncta when they were 0.25–3.0 µmDally-like protein expression during aging in MBs: Brains from flies 1–3 days or 50–55 days were dissected out and the tissue fixed, permeabilized and blocked as described above. Brains were then incubated overnight at 4 °C in an anti-Dally-like protein antibody at 1:5 in block , rinsed 3X in 0.1% PBST and incubated in goat anti-mouse Alexa Flour 647 antibody overnight at 4 °C . Images were acquired using a Zeiss 880 Laser Scanning Confocal inverted microscope with a Plan-Apochromat 63×/1.4 oil DIC M27 lens. Signal intensity was traced from maximum intensity projections of the MBLs and normalized to background intensity. Dlp signal intensity declined with age in TDP-43 flies, so the mCD8 RFP channel was used to trace the lobes in aged brains, ensuring the correct lobe area was used for signal intensity measurements.WT expression, in addition to western blots, we also measured mean pixel intensity from maximum intensity projections of the horizontal lobes (β and γ combined in maximum intensity projection) in two genotypes with SS01276-driven transgenes, OR-R;TDP-43WT::YFP and w1118; TDP-43WT::YFP mCD8::RFP . TDP-43WT also showed a bias for arm A (P = 0.0033), while OR-R controls showed a bias for arm C (P = 0.048). Although these analyses suggest that arm biases exist in males and are not unique to TDP-43 overexpression, a single replicate was used to test whether Dlp over-expression could rescue alternation deficits seen in TDP-43 overexpressing flies therefore hypothesis testing proceeded with non-parametric tests as described in Statistical analyses.Y-maze: To measure changes in working memory, we employed a y-maze assay described by Lewis, Negelspach, Kaladchibachi, Cowen and Fernandez . We tracDrosophila Activity Monitors [DAMs; Trikinetics, Waltham, MA; Drosophila, sleep is defined at five minutes of inactivity [Sleep: Adult flies from the three age time points were monitored individually using activity  and heresurvival [survivminer [Newly eclosed virgin males and females were separated and placed on fresh food, then transferred weekly into new food vials. The number of surviving flies was counted every other day for 100 days. Survival analysis and plots were generated using R packages survival  and survvivminer .WT YFP, or TDP-43G298S YFP were collected for each of three biological replicates. Frozen flies were then transferred to 50 mL conical tubes and heads were separated from the bodies using four rounds of vortexing and flash freezing with liquid nitrogen. A sieve was used to filter out the bodies and isolated heads were collected into tubes containing lysis beads (Next Advance Green lysis beads). Heads were homogenized (Next Advance Bullet Blender) in one mL fresh lysis buffer , protease inhibitors , and RNAsin Plus 400 u/mL (Fischer Scientific PRN2615), centrifuged for 10 min at 10,000 rpm, and the lysate was collected. A portion of the lysate was saved for the protein input and RNA input samples. For protein input, the lysate was mixed with 2X Laemmli buffer, boiled at 95–100 °C for 10 min, and stored at − 20 °C. For RNA input, TRIzol reagent (Thermofisher 15596026) was added to the lysate and the sample was stored at − 80 °C. High affinity GFP-Trap magnetic agarose beads (ChromoTek) were added to the remaining lysate and rotated end-over-end for 90 min at 4 °C to allow for binding of YFP. The beads were separated from the lysate with a magnet and washed three times with fresh wash buffer . The beads were resuspended in wash buffer and split into two tubes for the protein IP and RNA IP samples. For the protein IP sample, 2X Laemmli buffer was added to the beads, samples were boiled at 95–100 °C for 10 min, and the beads were removed with a magnet. Western blots (see above) were performed to ensure that the immunoprecipitated complexes contained TDP-43 before processing the RNA IPs. For the RNA IP sample, TRIzol reagent was added to the beads, the solution was pipetted up and down for 60 s, and the beads were removed with a magnet.RNA Immunoprecipitations: Equal numbers of male and female flies, aged one to three days, were pooled in Eppendorf tubes and flash frozen in liquid nitrogen. A minimum of 500 flies expressing YFP, TDP-43futsch was recovered as enriched with TDP-43 in both variants  following manufacturer’s protocol. Determination of cDNA quality and quantity was determined via Agilent Tapestation High Sensitivity D5000 Screentape and Qubit dsDNA HS assay for input into library amplification. Libraries were quantified by Agilent Tapestation High Sensitivity D1000 Screentape and Kapa Library Quantification kit for Illumina platforms. Libraries were pooled equimolarly, pools were quantified by Agilent Tapestation High Sensitivity D1000 Screentape and Kapa Library Quantification kit and loaded on the NovaSeq 6000 S4 flowcell and sequenced to 101 × 11 × 11 × 101 cycles. Trimmed fastqs were aligned to the Dmel genome with STAR v2.6.1d . AlignedFunctional annotation: The Database for Annotation, Visualization, and Integrated Discovery (DAVID) was used to functionally annotated genes enriched with TDP-43 , 93. To RNA Extraction for Validation: RNA input and RNA IP samples, stored in TRIzol reagent (Thermofisher 15596026) at − 80 °C, were allowed to thaw completely on ice and chloroform was added to each sample. Samples were shaken briefly to mix, left to incubate for three minutes at room temperature, and then centrifuged at 12,000 rpm for 15 minutes at 4 °C to allow for separation of the aqueous and organic phases. The RNA-containing aqueous phase was collected and molecular grade isopropyl alcohol was added to each sample. Samples were incubated for 10 min at room temperature for RNA precipitation and then centrifuged at 12,000 rpm for 10 minutes. The pelleted RNA was washed with 75% ethanol (200 proof ethanol & HyPure water) and left to dry in a fume hood. RNA was resuspended in HyPure water and the concentration and 260/280 absorbance ratio were measured using Nanodrop.RT-qPCR: RNA extracted from input and IP samples was used as template RNA for cDNA synthesis by reverse transcription. Total RNA used in the cDNA synthesis reactions was normalized across samples. cDNA was synthesized using the Fisher First Strand cDNA synthesis kit (Thermofisher Scientific K1641). qPCR reactions were prepared in a 96-well qPCR plate, in triplicates, using Taqman Fast Advanced Master Mix (Thermofisher Scientific 4444556) and Taqman probes for Dally-like protein (dlp) (Thermofisher Scientific Dm01798597_m1) and Gpdh1 (Thermofisher Scientific Dm01841185_m1). qPCR was conducted on the qTOWER  qPCR machine. Delta Ct was calculated as the difference in Ct values (dlp—gpdh1). To determine enrichment of dlp mRNA in the IPs, ΔΔCt was calculated as the difference in Delta Ct values (IP—Input). Fold change was calculated as 2^(− ΔΔCt).Dally-like protein target validation: Brains from flies one to three days or 50–55 days were dissected out and the tissue fixed, permeabilized and blocked as described above. Brains were then incubated overnight at 4 °C in an anti-Dally-like protein antibody at 1:5 in block , rinsed 3X in 0.1% PBST and incubated in goat anti-mouse Alexa Flour 647 antibody overnight at 4 °C . Images were acquired using a Zeiss 880 Laser Scanning Confocal inverted microscope with a Plan-Apochromat 63×/1.4 oil DIC M27 lens. Signal intensity was traced from maximum intensity projections of the MBLs and normalized to background intensity. Dlp signal intensity declined with age in TDP-43 flies, so the mCD8 RFP channel was used to trace the lobes in aged brains, ensuring the correct lobe area was used for signal intensity measurements.STMN2 or KALRN cryptic exon or a STMN2 or KALRN canonical junction, as defined in [GPC6 gene expression was compared between neuronal nuclei containing STMN2 and KALRN cryptic exons and neuronal nuclei containing STMN2 and KALRN canonical splice junctions using the independent t-test function (ttest_ind) in python. Violin plots were generated between neuronal nuclei containing a cryptic exon and neuronal nuclei containing a canonical exon using the scanpy violin function.snRNA seq analyses from FTD patient brains: Log normalized gene counts tables from  were filfined in . Human ofined in . GPC6 geSS01276 available from the Janelia FlyLight Split-GAL4 Driver Collection [Drosophila MBs. Previous studies have shown that neurons forming the γ and α′/β′ lobes are embryonic and larval in origin, respectively [The insect MBs are comprised of large, complex neurons called Kenyon cells, referred herein as MB neurons (MBNs), which form a layered network with reticular feedback , structullection  to exprellection  in a subectively , and perectively .SS01276 in the 3rd instar larva (L3) and found that cytoplasmic YFP, YFP tagged TDP-43 (TDP-43::YFP) and membrane targeted RFP (mCD8:: RFP) are limited to MBNs  and TDP-43G298S OE brains . In L3, the MBs comprise approximately 60 γ lobe MBNs and 40 α′/β′ lobe MBNs [SS01276 drives expression in some combination of γ lobe or α′/β′ lobe MBNs. However, since adult expression of SS01276 was determined to be limited to γ and α/β lobe MBNs [SS01276 corresponds to γ lobe MBNs only.We characterized expression driven by obe MBNs  thus at obe MBNs , and givVisual inspection of larval MBNs indicated that TDP-43 YFP was largely confined to the nucleus in L3 brains Fig. a but becTDP-43WT::YFP mCD8::RFP 0.978 P < 0.0001; Additional file TDP-43G298S:: YFP mCD8::RFP 0.967 P = 0.0004; Additional file WT OE model  and females  to measure total daytime and nighttime sleep. Our experiments show that on average, both male and female TDP-43 OE flies appear to be lethargic; they spent a greater proportion of their time sleeping during both the day and night compared to controls (approximately 11% more for TDP-43WT OE and approximately 8% more for TDP-43G298S OE). However, there are subtle differences in how TDP-43 OE affects sleep patterns based on age and sex. While males consistently sleep more during the day, this effect is weaker in females where we found trends towards increased daytime sleep at some but not all age time points in each genotype  and female declines appear less steep  wh, 95% CI ; P < 0.0WT or TDP-43G298S from Drosophila heads overexpressing TDP-43 YFP in MBNs using the SS01276 split GAL4 driver. Next, we used DESeq2 [WT YFP and TDP-43G298S YFP complexes, respectively  , for a total of 1572 mRNAs enriched across both models . Notably, differential expression analyses also recovered previously identified TDP-43 target mRNAs including futsch  [dscam2  [To understand how TDP-43 overexpression influences the MBN proteome, we immunoprecipitated YFP-tagged TDP-43d DESeq2  to asses.57E−10)  and dsca2.42E−3) .Fig. 6mRWT OE and 42% of TDP-43G298S OE highly associated mRNAs in MBNs overlapped with those we identified previously in motor neurons (MNs) when modeling ALS by overexpressing TDP-43 with the D42 GAL4 driver  and frizzled 2 (fz2) [dally-like protein (dlp), a heparan sulfate proteoglycan (HSPG) that interacts with multiple Wnt ligands [dlp mRNA was also found to be enriched with TDP-43 in fly MNs, where TDP-43 proteinopathy causes depletion of Dlp from the neuromuscular junction and simultaneous accumulation of Dlp puncta in cell bodies [dlp mRNA is a target of TDP-43 in MBNs as predicted by RNA seq, we used RNA IPs followed by RT-qPCR and confirmed that dlp mRNA is enriched in immunoprecipitated TDP-43WT and TDP-43G298S complexes compared to input   receptor ligands  were ovel bodies . To valiWT OE we found 461 enriched mRNAs that are shared in both MB and MN, and 594 mRNAs uniquely enriched in MBs  in the context of TDP-43 proteinopathy could mitigate TDP-43 dependent working memory deficits. Using the Y-maze working memory assay we found that the percent alternation deficits displayed by TDP-43WT OE flies could indeed be mitigated by dlp OE in MBNs , the glypican GPC4 , heat shock 70 kDa protein 2, HSPA2 , and the frizzled family receptor protein FZD7 .To validate our findings from ata sets  and founDrosophila with transcripts altered in single neuronal nuclei containing TDP-43 associated cryptic exons  versus neurons in which canonical splice junctions were present (N = 377), we found several human orthologs to be altered including dlp’s ortholog, GPC6 mRNA, which was found to be significantly upregulated  study of FTD patients with TDP-43 pathology and harboring mutations in C9orf72, the most common form of inherited ALS/FTD . When coNeurodegenerative diseases have historically been classified by shared symptomology and post-mortem pathology. More recently, the integration of molecular markers and genetic testing has uncovered evidence that cell-type susceptibility and dysregulation of specific molecular pathways can interact to produce similar clinical presentations across different disease etiologies . These fA common pathological feature of several neurodegenerative diseases including ALS, FTD, and other dementias is the nuclear depletion of the RNA binding protein TDP-43 accompanied by the accumulation of cytoplasmic, ubiquitinated foci , 18, 134Drosophila melanogaster, we leveraged a split GAL4 driver line with limited expression to a subset of Kenyon cells . Notably, this model exhibits nuclear depletion and cytoplasmic accumulation of TDP-43, accompanied by age-dependent axonal degeneration and cell loss, all of which recapitulate key aspects of disease pathology. Importantly, the onset of FTD relevant behavioral symptoms is detectable prior to widespread degeneration, as observed in human disease. Additionally, this model identifies both novel, MB specific, and motor neuron-shared mRNA candidate targets that have previously been associated with TDP-43 pathology. Of the latter, here we chose to focus on the glypican Dlp, a Wg/Wnt signaling regulator. We show that Dlp is a functional target of TDP-43 in the MB circuit that mediates, in part, TDP-43 dependent working memory deficits in the fly model.It has been previously shown that overexpression of TDP-43 in all MBNs using the driver line OK107 GAL4 (RRID:BDSC_854) caused age-dependent cell loss and axonal degeneration in MBs, however degeneration severity varied widely across nearly isogenic individuals , 139 andNeurodegeneration in ALS/FTD is thought to be driven by both loss of nuclear function, as some nuclei become devoid of TDP-43, and cytoplasmic gain of toxic function, as evidenced by TDP-43 accumulation in cytoplasmic puncta. While thus far invertebrate models of TDP-43 proteinopathy have not faithfully recapitulated TDP-43 mislocalization e.g., , 142, heAxonal degeneration is evident only in older adult flies and differentially affects MBNs that form the α/β and γ lobes, with the latter showing the greatest degeneration severity. This is particularly interesting because MBNs that form the γ lobe are embryonic in origin and therefore older than the pupal-born MBNs that form the α/β lobes , 145, hiDrosophila model.Frontotemporal lobar degeneration with TDP-43 proteinopathy (FTLD-TDP) is pathologically classified based on the cortical layers affected, the number of dystrophic neurite, and the extent of neuronal cytoplasmic inclusions . This paIn humans, behavioral symptoms of FTD precede widespread degeneration by years suggesting that behavioral deficits arise prior to neuronal pathology, in the absence of complete loss of nuclear TDP-43 function or cytoplasmic accumulation . TherefoBehavioral variant frontotemporal dementia (bvFTD) is diagnosed based on often subtle behavioral symptoms including a lack of empathy, increased apathy, dysinhibition and deficits in executive function . FTD wasIn addition to observed behavioral deficits, flies with TDP-43 proteinopathy show an approximately 10% reduction in median lifespan. This may be roughly comparable to human patients where FTD exerts a subtle effect on lifespan. In humans, mean age at diagnosis is 61.9 for early onset disease (< 65 years) with mean survival after diagnosis around 8 years . AlthougDrosophila TDP-43  based on UG-richness [Sequestration of mRNAs is one the mechanisms by which TDP-43 is thought to contribute to neurodegeneration and identifying candidate mRNA targets has helped better understand disease etiology , 157. Alrichness . In futuFunctional analyses of the mRNAs associated with TDP-43 identify numerous cellular pathways including Wg and Hippo that have been previously implicated in sleep regulation in flies . Additiodlp, an mRNA target of TDP-43 that we previously identified in the ALS model of TDP-43 proteinopathy, and a known regulator of Wg signaling. Indeed, Wg signaling is dysregulated in ALS [dlp OE in MBNs mitigates TDP-43 dependent working memory deficits, as evidenced by improved alternation in the Y maze assay. Taken together, these results support the notion that Wg/Wnt signaling is altered in TDP-43 associated neurodegeneration and modulating its activity via Dlp mitigates behavioral deficits caused by TDP-43 proteinopathy. Further substantiating these findings, we found that the expression of GPC6 mRNA, a human ortholog of dlp, is altered in FTD patient brains [i.e., cryptic exon inclusion in specific TDP-43 transcriptional targets), consistent with a link between Dlp/GPC6 and TDP-43 pathology. We speculate that this increase in GPC6 mRNA in neurons with TDP-43 nuclear depletion may reflect a compensatory upregulation caused by a cytoplasmic decrease in protein levels that we detected in Drosophila MBN axons. Indeed, the same study by Gittings, Alsop et al. [Drosophila models to identify functionally relevant disease targets. Interestingly, recent GWAS studies identified GPC6 as a risk factor for AD in African Americans [dlp/GPC6 mitigate specific phenotypic aspects of FTD and identify additional targets across the spectrum of TDP-43 proteinopathies.For functional validation we chose to focus on d in ALS , 162, mRd in ALS  and WNT1d in ALS . Despitet brains , specifip et al. , identifp et al. . Althougmericans , 117. InAdditional file 1:Figure 1-supplement 1. Maximum intensity projections showing split Gal4 driver line SS01276 expression in the 3rd instar larva. Figure 1-supplement 2. TDP-43G298S localization in MBN cell bodies. Figure 1-supplement 3. Method for measuring the ratio of nucleus to total cell TDP-43 YFP from mean pixel intensity of MBNs. Figure 2-supplement 1. Mushroom body lobes (MBLs) show age-related, region-specific TDP-43G298S cytoplasmic accumulation and axonal fragmentation. Figure 4-supplement 1. TDP-43G298S overexpression in MBNs reduces arousal, increasing day and night sleep. Figure 5-supplement 1. Mutant TDP-43 overexpression in MBNs is sufficient to reduce lifespan. Figure 6-supplement 1. mRNAs enriched with TDP-43G298S overexpression in Drosophila MBs. Figure 6-supplement 2. Functional annotation of enriched targets in fly models of TDP-43 driven dementia. Figure 7-supplement 1. Dally-like protein is a target of TDP-43G298S in MBNs. Figure 7-supplement 2. TDP-43WT protein expression is not reduced by the presence of a second UAS-driven transgene. Figure 7-supplement 3. TDP-43WT YFP expression levels is not reduced by concomitant expression of a second UAS-driven transgene (mCD8 RFP).Additional file 2:Table S1E Nucleus to whole cell TDP-43 YFP ratios summarized in Figure 1e. Table S1F Summary statistics for cell number data presented in Figure 1f. Table S2A Summary statistics for mCD8RFP intensity presented in Figure 2a. Table S2B Summary statistics for YFP particle size values presented in Figure 2b. Table S3 Summary statistics for y-maze data presented in Figure 3. Table S4A Summary statistics for night sleep presented in Figure 4a. Table S4B Summary statistics for sleep bout length presented in Figure 4b. Table S4C Summary statistics for sleep bout number presented in Figure 4c. Table S6A Wild-type TDP-43 GOterms from differential gene expression presented in Figure 6. Table S6B Mutant TDP-43 GOterms from differential gene expression presented in Figure 6. Table S7CD Y-maze summary statistics for data presented in Figures 7c and 7d. Table S8 mRNAs enriched in nuclei with TDP-43 associated cryptic exons. Table S9 Summary of phenotypes observed in wild-type and mutant fly models.