Self-administration was characterized under Progressive Ratio (PR) and Fixed Ratio (FR) 1 schedules of reinforcement in 3 h sessions. THC decreased IVSA of oxycodone in a FR procedure but increased reward seeking in a PR procedure. CBD decreased the IVSA of oxycodone in the FR but not the PR procedure. The results are consistent with an anti-reward effect of CBD but suggest THC acts to increase the reinforcing efficacy of oxycodone in this procedure.Epidemiological evidence suggests that the legalization of cannabis may reduce opioid-related harms. Preclinical evidence of neuropharmacological interactions of endogenous cannabinoid and opioid systems prompts further investigation of cannabinoids as potential therapeutics for the non-medical use of opioids. In these studies female rats, previously trained to self-administer oxycodone (0.15 mg/kg/infusion) intravenously in 6 h sessions, were allowed to self-administer oxycodone after exposure to cannabidiol (CBD) and Δ It is difficult to experimentally determine if cannabis reduces opioid use in humans, but animal models can assist in distinguishing pro-reward from anti-abuse effects.Reduction of opioid use concomitant with cannabis use  may aris9-tetrahydrocannabinol (THC) reinstates beer- and sucrose-seeking behavior in rats, for example  reduces alcohol or cocaine seeking behavior in rats when applied via a dermal gel , aged 10–11 weeks on arrival in the laboratory in a single cohort of 20. The vivarium was kept on a 12:12 hour reversed light-dark cycle, and behavior studies were conducted during the vivarium dark period. The animals were pair housed and food and water were provided ad libitum in the home cage and in the self-administration chambers. Experimental procedures were conducted in accordance with protocols approved by the Institutional Animal Care and Use Committee of the University of California, San Diego and consistent with recommendations in the NIH Guide .Two animals were lost due to mechanical failure of vivarium equipment prior to the start of intravenous self-administration (IVSA) training. Three other animals were lost to the study prior to the start due to catheter/implantation complications . One animal was discontinued for self-injurious behavior (paw chewing) after Session 27. Five animals were discontinued due to loss of catheter port integrity after Sessions 22, 31, 53, 54, and 64, respectively. The nine at the end of the study included 5 original THC group animals and 4 original Vehicle group animals.2.2Vapor inhalation was conducted as previously described  with the2.3Female rats were implanted with intravenous catheters and trained to self-administer oxycodone (0.15 mg/kg/infusion) on a Fixed Ratio (FR) 1 schedule of reinforcement using methods previously described . In this2.42.4.1j)–5  before the 34th and 35th sessions in a counter-balanced order. The 36th and 37th sessions were conducted under PR, without any pre-session treatment. The per-infusion dose of oxycodone was changed to 0.06 mg/kg/inf for Sessions 38–41 and the rats were injected with either Veh or THC  before the 39th and 40th sessions in a counter-balanced order. For analysis, the day following the injection days was used as the no-injection comparison.On sessions 33–37 animals were tested on a Progressive Ratio (PR) procedure with 0.15 mg/kg/infusion oxycodone available. In the PR paradigm, the required response ratio was increased after each reinforcer delivery, within a session  as deterj)–5 . In this2.4.2For Sessions 42–64, rats were permitted to self-administer oxycodone in 3 h sessions under a FR1 contingency. In Sessions 43–45 the oxycodone dose  was varied in a counter-balanced order.The study next evaluated injection of 0, 5, 10 and 20 mg/kg of THC, i.p., prior to oxycodone (0.06 mg/kg/infusion) self-administration sessions, assessed in a counter-balanced order in Sessions 48–51.IVSA was then suspended for a week between Sessions 53 and 54 to permit assessment of the acute effects of THC  on rectal temperature and nociception assays.Self-administration of oxycodone was returned to the original training dose (0.15 mg/kg/infusion) in 3 h sessions (FR1) for Sessions 56–64. Rats were exposed to cannabidiol (CBD) vapor (400 mg/mL in the PG) or vehicle (PG) vapor for 30 minutes immediately prior to IVSA sessions in a counter balanced order for Sessions 59–60. Two additional animals were discontinued due to catheter failure, thus N=10 . Vapor inhalation was selected because we have previously reported effects of CBD on body temperature via this route  that wer2.4.3Then rats were returned to the PR response contingency (0.15 mg/kg/infusion) for Sessions 65–69 and exposed to vehicle (PG) vapor, CBD vapor (400 mg/mL in the PG) or THC vapor (100 mg/mL in the PG) for 30 minutes immediately prior to IVSA sessions in a counter balanced order for Sessions 66–68. Thereafter the oxycodone dose was increased to 0.30 mg/kg/infusion for Sessions 70–74 and the PG, CBD and THC pre-exposure conditions repeated in a counterbalanced order in Sessions 71–73. For Sessions 75–77 the dose of oxycodone  was altered in a counterbalanced order, with no pre-treatment. One individual’s catheter exhibited resistance to daily flushing in this experiment and was non-patent by the final experiment and was therefore excluded from analysis. Rats completed PR sessions with no pre-treatment at the 0.03 mg/kg/inf dose for Sessions 78–83, and at the 0.06 mg/kg/infusion dose for Session 84.2.4.4Rats (N=9) were returned to FR1 in 3 h sessions with the oxycodone dose 0.06 mg/kg/infusion for Sessions 85–88. Rats were exposed to PG vapor, or CBD vapor  for 30 minutes immediately prior to IVSA sessions in a counter balanced order for Sessions 86–88. One rat had a catheter that failed patency during this study and was therefore excluded.2.5Infusions obtained, the percentage of responses directed at the drug-associated lever, total correct responses and breakpoints (for Progressive Ratio) were analyzed by ANOVA, or by mixed-effect models where there were missing values. Within-subjects factors of Dose or Pre-treatment were included where relevant. In our approach a single priming infusion is delivered if no responses have been made within 30 minutes of session initiation under FR (this is not included for PR). These are rare after the initial few sessions of acquisition and are thus reported qualitatively where present, but not formally analyzed. In all analyses, a criterion of P<0.05 was used to infer that a significant difference existed. Any significant main effects were followed with post-hoc analysis using Tukey (multi-level factors), Sidak (two-level factors) or Dunnett (to assess change relative to one of the Treatment Conditions.) correction. All analysis used Prism for Windows .3.3.1This experiment showed that THC  increases breakpoints reached by female rats in the progressive ratio procedure , confirmThe impact of THC was statistically indistinguishable across oxycodone unit doses  as there was no significant effect of oxycodone Dose nor an interaction of Dose with Treatment Condition. The post-hoc Dunnett test of the marginal mean confirmed significantly lower breakpoints were reached in the No-Injection condition compared with the THC condition, across oxycodone Dose.3.2The lack of difference between breakpoints generated under the different oxycodone unit doses prompted a comparison under FR1 conditions (3 h sessions to match the maximum duration of the PR sessions), to determine if these were functionally different doses in IVSA. This demonstrated that the 0.15 mg/kg/infusion dose resulted in fewer infusions obtained  reduced the number of infusions obtained in comparison with PG vapor inhalation or a session in which there was no exposure prior to the IVSA session  under FR1 because the oxycodone dose did not alter self-administration of oxycodone under a PR procedure. The one-way ANOVA confirmed a significant impact of oxycodone Dose  on infusions obtained and the Tukey post hoc further confirmed significant differences between all doses .Because CBD did not alter oxycodone IVSA under a PR procedure, we re-determined the impact of CBD vapor under FR1 to rule out a potential development of tolerance to the CBD. For this we tested two CBD vapor doses (100 and 400 mg/mL) and the 0.06 mg/kg/infusion dose of oxycodone. The one-way ANOVA confirmed a significant impact of Pre-Treatment condition  on infusions obtained and the Tukey post-hoc further confirmed significantly fewer infusions were obtained after CBD 400 exposure compared to PG exposure .4.9-tetrahydrocannabinol (THC) increases female rats’ oxycodone seeking under a Progressive Ratio schedule, while it reduces oxycodone self-administration under a Fixed Ratio 1 (FR1) contingency. Our previous study was conducted in male rats and while several conditions of oxycodone IVSA were evaluated, it was all conducted with a FR1 schedule of reinforcement  and the present study found that the effect of FR vs PR reinforcement schedule on the impact of THC extended to oxycodone IVSA in female rats. The increased behavioral output  in the PR procedure associated with THC  in a PR procedure, nor any interaction of the effect of THC with heroin unit dose, in that study. Therefore, the prior results were quite similar to the impact of unit dose of oxycodone across FR1 and PR procedures in the current study.It was somewhat unexpected that rats reached similar breakpoints for 0.06 and 0.15 mg/kg/infusion in the PR under control conditions  given an apparent difference across this dose range that was observed in prior work with male rats trained in sessions of 1 h or 12 h duration . In partOne of the strengths of this study was that animals were originally trained in extended access (6 h) sessions, examined for this study in 3 h sessions under both FR1 and PR response contingencies and exhibited a high degree of behavioral stability across an extended number of sessions. This enhances comparison of findings across the experiments. In general, this confirms ongoing work from multiple laboratories showing that oxycodone is a typical opioid reinforcer in rat intravenous self-administration, and that stable self-administration patterns are produced in short-access IVSA in male and female rats . SimilarIn contrast with THC, the impact of CBD was to reduce oxycodone seeking under FR1 and to leave it unaffected under the PR1 contingency. CBD has been previously shown to have anti-drug-seeking effects in rats  when delInterestingly, the inhalation of vapor from cannabis extracts with high CBD content (64.2% CBD and 7.1% THC) decreased fentanyl IVSA under a FR, but not a PR, schedule in rats with and without induced neuropathic pain . InteracIn conclusion these studies provided additional evidence that THC reduces oxycodone seeking under easy-access conditions because it enhances the rewarding value of a unit dose of oxycodone. When access requires more work in a PR procedure, THC increases oxycodone seeking responding. In contrast, the effect of CBD was to decrease oxycodone self-administration in easy access conditions but to have no effect in harder access conditions. This cautions that cannabis is not necessarily reducing addiction liability in association with any observed reduction in oxycodone intake.


The deep subsurface is, including subsurface groundwater, one of the last unknown frontiers to humankind. A significant part of life on Earth resides in the deep subsurface, hiding the great potential of microbial life of which we know only a little. The extreme conditions in deep terrestrial subsurface, such as anoxia and oligotrophy, high salinity, lack of primary production, are thought to resemble those of early Earth, which makes this environment an analog for studying early life in addition to possible extraterrestrial life in ultra-extreme conditions. Furthermore, deep subsurface microorganisms offer possibilities for novel biotechnological applications.Much new knowledge has been gained over the last decade describing the microbial communities in different terrestrial deep biospheres. The previous Research Topic . Giovannelli et al. presents a new view of sampling strategy to study ecological features over a greater geological gradient for a broad-scale analysis of the subsurface microbial ecosystem in contrast to studying very specific sites. However, laboratory incubations should not be forgotten, as shown by Nuppunen-Puputti et al. and Mandal et al., where different microbial physiologies were revealed by enrichment cultivation, which might not have been detected otherwise. The challenge now is to elucidate the activity and functional properties of the microbial communities and to connect the biological data to that of the geological to link microbial diversity to microbial functions in the very inaccessible deep terrestrial subsurface. With this Research Topic, we aimed to collect new knowledge about the roles and functions of microbial communities in the undisturbed or disturbed subsurface and called upon our colleagues to step up to the task. The deep subsurface is of interest worldwide and the present Research Topic include samplings from Asia, South America, North America, and Europe!The present Research Topic covers the important field of geomicrobiological research of constructed deep geological sites, such as spent nuclear fuel repositories containing metals and bentonite clay  and long-term microcosms studies that enable us to manipulate the environmental conditions  would allow us to close the gap in our current knowledge of the activity and functional properties of the microbial communities, and to connect the biological data to that of the geological to link microbial diversity to microbial functions in the very inaccessible deep terrestrial subsurface. The slow turnover time of deep biosphere microorganisms is reflected also in the studies of this Research Topic, with experimental times ranging from several months  to several years .The deep subsurface biosphere is one of the largest microbial ecosystems on Earth, but many fundamental questions about how life exists and thrives in this habitat are still unanswered. We know only little about the biochemical processes of the deep Earth's crust, or the microbial communities interacting with biotic and abiotic factors in the isolated aquifers or the ability of the microbial communities to adapt to changing environments. This is due much to the high costs, inaccessibility, and challenges of direct sampling of deep environments and to the fact that many deep biosphere microbial populations grow very slowly or are so-called resistant to laboratory culturing techniques. To overcome the access and cost challenges, Nuppunen-Puputti et al. Sessile and planktic bacterial communities were studied in microcosms containing Finnish groundwater  and mica schist from the same site. The microcosms were spiked with different carbon substrates, and it was shown that the microbial communities differed between microcosm types and sessile and planktic communities. Especially, sulfate-reducing bacteria were frequent occupiers of the rock surfaces. Although the sessile and planktic bacterial communities differed in their community structure, some sort of metabolic cooperation likely exists between them and with the fungal communities.The importance of studying both sessile and planktic microbial communities in the deep biosphere was demonstrated by Mandal et al.; Nuppunen-Puputti et al.). Mandal et al. noted a better response of rock-hosted community toward carbon dioxide vs. bicarbonate amendment. They propose that these endolithic microorganisms could show great potential for answering the fundamental questions of deep life and their exploitation in carbon dioxide capture and conversion to useful products.Inorganic carbon or low-molecular-weight carbon  links multiple biogeochemical pathways and thus represents an important carbon and energy source for microorganisms in the deep crystalline bedrock . The microcosms without additional substrates developed the lowest bacterial community richness compared to acetate, methane, or methanol-amended mesocosms . Methanol supported the richest bacterial and fungal communities. Especially the Hydrogenophaga/Serpentinomonas attached to the rock surface had a significant response to methanol.A pulse of low-molecular-weight carbon compounds in the beginning of a long-term microcosm incubation supported epilithic community development on mica schist . Corrosion of metallic waste container materials by anaerobic microorganisms, most commonly sulfate-reducing bacteria, originating from porewater or groundwater has long been considered a major risk factor for long-term safety. Burzan et al. didn't find direct evidence of sulfate reduction within bentonite or of microbially induced corrosion of the carbon steel, unlike previously reported in the case of the crystalline bedrock environment.Deep geological repositories are considered in many countries as the most promising solution for the safe and long-term disposal of hazardous wastes. The ability of deep biosphere microorganisms to influence the environmental conditions or material integrity in deep geological repositories has remained a topic of interest due to the potential impact on the long-term safety of these repositories. In many repository concepts the spent nuclear fuel is packed in metal containers immersed in bentonite clay buffer, which when fully expanded will contain an internal pressure that will hinder microbial growth and movement toward the waste containers. There remains still much to learn about the size of the microbial communities, the genetic variation and metabolic functions in the deepest part of the biosphere despite the advancements during the past decade. Our understanding of how deep biosphere microbes react to changing environments, be it nutrient availability or constructed environment, has greatly advanced. Shifting the focus from only the planktic community to cover also the sessile community and studying the long-overlooked fungal community in addition to better-known bacterial and archaeal communities have advanced our understanding of the diversity and functionality of the deep biosphere. Advancement in sampling techniques and access to geographically and geologically diverse sites has been crucial in building new knowledge. Still, studying the deep biosphere remains a privilege of few and more accessible and innovative sampling and laboratory scale techniques are needed.Both authors listed have made a substantial, direct, and intellectual contribution to the work and approved it for publication.


Flash-induced porous graphene (FPG) was synthesized via a broad-spectrum flash lamp that induced synergistic photo-effects between ultraviolet and visible-near-infrared wavelengths, resulting in large-area synthesis in just a few milliseconds.−1 was produced, exhibiting low density (0.0354 g cm−3) and outstanding absolute electromagnetic interference shielding effectiveness of 1.12 × 105 dB cm2 g−1.A hollow pillar graphene with low sheet resistance of 18 Ω sqA lightweight, flexible, and high-throughput FPG is applied for electromagnetic interference shielding of a drone radar system and the human body.The online version contains supplementary material available at 10.1007/s40820-023-01157-8. 2-size porous graphene with a hollow pillar structure in a few milliseconds. The resulting material demonstrated low density (0.0354 g cm−3) and outstanding absolute EMI shielding effectiveness of 1.12 × 105 dB cm2 g−1. The FPG was characterized via thorough material analyses, and its mechanical durability and flexibility were confirmed by a bending cycle test. Finally, the FPG was utilized in drone and wearable applications, showing effective EMI shielding performance for internal/external EMI in a drone radar system and reducing the specific absorption rate in the human body.Porous 2D materials with high conductivity and large surface area have been proposed for potential electromagnetic interference (EMI) shielding materials in future mobility and wearable applications to prevent signal noise, transmission inaccuracy, system malfunction, and health hazards. Here, we report on the synthesis of lightweight and flexible flash-induced porous graphene (FPG) with excellent EMI shielding performance. The broad spectrum of pulsed flashlight induces photo-chemical and photo-thermal reactions in polyimide films, forming 5 × 10 cmThe online version contains supplementary material available at 10.1007/s40820-023-01157-8. With the advent of future mobility and wearable electronics, various electronic components including sensors, actuators, processors, and controllers are integrated with cloud computing, big data, and 5G wireless communication for autonomous driving, intelligent robotics, and real-time healthcare –3. The hA variety of composites have been developed for soft and lightweight EMI shielding materials by incorporating conductive fillers such as metal nanomaterials , 10, MXeRecently, light-material interactions have attracted a great deal of attention because they can activate transient, multi-physical, and spatiotemporally controlled reactions –23 to st2, N2 and CO gases were released from the PI film surface, resulting in large area (5 × 10 cm2) hollow pillared graphene with a low density of 0.0354 g cm−3 in a few milliseconds (ms). The synthesized FPG showed outstanding absolute EMI SE of 1.12 × 105 dB cm2 g−1 due to its low sheet resistance (18 Ω sq−1) and multiple internal scattering. To confirm the FPG formation mechanism, the chemical/structural changes were thoroughly analyzed by using various characterization methods, such as scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS), Fourier transform infrared spectroscopy (FTIR), Raman spectroscopy, transmission electron microscopy (TEM) and X-ray diffraction (XRD). The feasibility of the FPG growth was verified by theoretically calculating the heat distributions of the PI film, which exhibited sufficient surface temperature (up to 2300 ℃) for graphene synthesis. The synthesized FPG was applied for aerospace and wearable applications, including drones and the human body. In a drone radar system, the lightweight FPG film successfully replaced conventional heavy metal jigs by shielding EMI from multi-stacked printed circuit boards (PCBs) and external noise signals. For wearable applications, the FPG effectively reduced the specific absorption rate (SAR) of the human body by 80.3% per 1 g of tissue and also lowered the magnitude of EM waves penetrating the human body by ~ 9.88 dB.Herein, we report a flash-induced porous graphene (FPG) synthesis method to achieve a high-throughput, outstanding performance, lightweight, and flexible EMI shielding film for drones and wearable applications. The ultraviolet (UV) range of the broad spectrum flashlight led to the breaking of chemical bonds in polyimide (PI) molecules which subsequently created defects for effective light absorption in the visible-near-infrared (Vis–NIR) wavelengths. The photothermal heat generated by the Vis–NIR region caused graphene synthesis while CO−2 and 1–10 ms, respectively. The fluences were measured using an energy meter for different lamp power and pulse widths. The single irradiation area was set at 5 cm × 10 cm with a square mask for edge clearance. The stage was moved along the Y-axis to obtain a large area FPG of 10 cm × 10 cm. The lamp power, pulse width, irradiation frequency, and stage moving speed were adjusted to be 9 W cm−2, 4 ms, 1 Hz, and 100 mm s−1, respectively.Commercial polyimide films  were used for the experiments. PI films were cleaned using ethanol and de-ionized water in an ultrasonic bath. Photo-pyrolysis of the PI film was performed using a flash lamp  under ambient conditions. The wavelengths of the flash lamp range from 250 to 1100 nm  and the wavelengths are tuned using an optical pass filter . The lamp power and pulse width were adjusted in the ranges of 1–16 W cm2) used in the simulation were given by the COMSOL material library. The heat generation per volume from absorbed light was calculated by Eq. (−1. X-ray diffraction (XRD) analysis was conducted using a RIGAKU Ultima IV with Cu Kɑ radiation (λ = 1.54 Å). The sheet resistance was characterized using a four-point probe . A vector network analyzer  and waveguides were used to measure scattering parameters (S11 and S21) of the material in the 18–26.5 GHz (K-band) frequency range at room temperature. The EMI SE value was calculated using the given equations :–36:34–36entclass1pt{minimaquations –40:3\docR, SEA, and SET are reflection, absorption, and total EMI SE, respectively. The effective absorbance (Aeff) is calculated to evaluate the absorption potential of materials. The home-built moving stage was used for bending the FPG.T, R, and A represent the transmission, reflection, and absorption coefficients, respectively. SEA K-band frequency-modulated continuous wave (FMCW) radar on a drone (Pixhwak) operating from 23 to 25 GHz was used to verify the EMI shielding properties for a machine . As show2 area by controlling the position of the probe antenna, as shown in Fig. S2 [The internal EMI from the RF transceiver board was observed using an open-ended probe antenna OEW-42). The 2-dimensional electric fields radiated from the RF transceiver PCB were measured in a 20 × 20 cm. The 2-d Fig. S2 . The antTo verify the EMI shielding performance for a human body, the SAR was measured . As show2, N2, and CO gases are released by dissociation of C=O, C–N, and C–O bonds [sp3) transforms into stacked, ordered graphene sheets (sp2). This process results in the formation of porous graphene, a crystalline structure of carbon atoms partially arranged in a hexagonal lattice [Figure –O bonds , 55. Car–O bonds –58, the  lattice . The syn−2. As displayed in Figs. To confirm the synergistic photo-effects, the full wavelength, and the Vis–NIR wavelength by blocking UV of the flash lamp were individually irradiated on the PI film with lamp fluence of pristine, 8, 12, 14, 18, and 22 J cm2 FPG manufactured by irradiation of a flash twice with a beam area of 5 × 10 cm2. The SEM image in the inset of Fig. sp2 carbons. These are more likely to form the hexagonal graphene structure than other precursors, which primarily generate amorphous carbon.To achieve effective EMI shielding in various applications such as future mobility and wearable devices, a large-area EMI film should be fabricated with high productivity. Figure Due to its high thermal stability, PI is not greatly affected by temperature conditions. However, if the temperature is artificially raised to preheat the PI, it could potentially enhance the speed of the photo-reaction, which could in turn affect the rate of gas emission and the pore expansion time . The fla2. The difference in production time was negligible in the small area of 10 × 10 mm2, while a significant decrease of more than 66.5 times was confirmed in large areas of 100 × 100 and 200 × 200 mm2. Figure S7 shows the calculation method and Table S1 presents the detailed calculation results. Note that the flash can dramatically increase productivity by increasing the size of the lamps and chamber, or by application to roll-to-roll processes. Table S2 shows comparison of porous graphene formation method of this work with previously reported light-induced method. These results suggest that the flash lamp is a suitable light source to fabricate a porous graphene-based EMI film with a large area and high productivity.Figure −2. As depicted in Fig. −2, with heat transfer occurring at roughly 100 µm depth within the PI. These results indicate that the necessary temperature for graphene formation can be achieved using lamp irradiation. Figure −2. As the lamp fluence increased, ~ 5 µm-sized micro-pores began to form and were finally synthesized into porous structural walls that were hundreds of nanometers wide. To further characterize the porous structure, we verified the pore size by lamp fluence, as shown in Fig. S8. As the lamp fluence increased from 18 to 24 J cm−2, the pore size increased from approximately 1.4 to 19.2 µm.To confirm whether a temperature of 1700 °C, necessary for graphene formation, can be achieved using a lamp, we conducted a COMSOL simulation to evaluate the maximum temperature and temperature distribution at various depths. Theoretical calculations were performed to determine the thermal distribution within a 125 µm-thick PI under various flash fluences ranging from 6 to 26 J cms, 397 eV for N 1s, and 532.5 eV for O 1s, which correspond to the main elements of carbon, nitrogen, and oxygen on the surface of the FPG [−1 indicating the stretching and bending modes of C–O, C–N, and C=O bonds. After lamp irradiation, a broad absorption spectrum ranging from 1000 to 1700 cm−1 was observed. At a fluence of 22 J cm−2, the peak associated with C=O, C–O, and C–N noticeably decreases. The XPS and FTIR results demonstrate that the carbon content becomes dominant due to the gas release accompanied by a decrease in nitrogen and oxygen content [The chemical changes and structural characteristics were analyzed via XPS, FTIR, Raman spectrum, TEM and XRD to demonstrate the synthesis mechanism of micro-holes and porous materials. Figure  the FPG . As the  the FPG . Results content .−1, respectively [−2, the 2D peak was observed. The inset graph shows the ratios of ID/IG and I2D/IG Raman peaks, which indicate defects in graphene materials and the number of layers in graphene [ID/IG decreases from 1 to 0.8, and the ratio of I2D/IG increased from 0 to 0.38. The I2D/IG ratio of 0.38 indicates the formation of 4–5 layers of graphene [Figure ectively . In partectively –74. As tgraphene , 76. As graphene . Figure graphene . The crygraphene . The Ramgraphene , 79. Tab−2. Vertically aligned porous hollow pillar structures were formed because the pores expanded and grew in the direction perpendicular to the surface where the gas was released. The thickness of the hollow pillar FPG was formed to ~ 62 μm from the 125 μm thick PI film at a fluence of 22 J cm−2. Figure S11 shows cross-sectional images for thickness measurement. The SEM image in Fig. Figure −1 while the thickness increased from 26 to 92 µm. As the lamp fluence increased, the electrical conductivity also increases, as shown in Fig. S12. Based on these results, the EMI SET of the FPG was evaluated at a frequency range from 18 to 26.5 GHz, the K-band mainly used for 5G/6G communications, as shown in Fig. T was continuously enhanced over the entire frequency range. Figure S13 depicts the EMI SE measured in the C-band (4 to 8 GHz), with the measured results confirming SE performance of over 20 dB, comparable to the K-band.High EMI shielding properties of porous graphene can be achieved by low sheet resistance and high thickness which enables enhancement of the surface reflection and internal absorption, respectively –82. FiguT of the FPG, the EMI shielding performance of SER and SEA as well as the power coefficient were measured. Figure R, SEA, and SET of the FPG under various fluences of 12, 14, 18, 22 and 24 J cm−2. As the fluence increased, SET rose from 12.9 to 34 dB, and SEA also elevated from 9.1 to 27.8 dB, while SER consistently remained below 7 dB. These results confirm that SEA primarily drove the increase in SET. Figures Aeff and power coefficients , which are used to explain the EMI shielding mechanism of the FPG. Throughout the fluence range from 12 to 24 J cm−2, the R values were always higher than the A values. As the fluence increased, R values increased gradually while A and T values decreased, indicating that surface reflection plays a more significant role than absorption and that lower resistance induced by lamp fluence leads to higher surface reflection. The absorption and reflection are the two main mechanisms influencing EMI shielding. Among these, reflection is enhanced by high electrical conductivity, leading to increased reflection of EM waves and, consequently, a larger R coefficient. In shielding materials where reflection dominates [R significantly increases and the T decreases, the A may decrease according to Eq.  and a high absolute EMI SE of 1.12 × 105 dB cm2 g−1 by the porous structure. The FPG retained 1.4 and 0.86 times its initial sheet resistance and SE performance, respectively, even after 10,000 bending tests. The FPG, effectively shields internal and external EMI of a drone radar system, reduces the radiated electric field (− 10.8 dB) and antenna gain (− 12.2 dB), and detects radar signals without saturation. In wearable applications, the FPG reduced SAR by 80.3% and 77.8% for indoor and outdoor frequencies, respectively, satisfying the IEEE standard. We believe that the proposed lightweight, flexible, and highly productive FPG will be effective in resolving diverse EMI shielding issues for future mobility and wearable applications.In summary, we developed a novel high-throughput synthesis technique of flash-induced porous graphene via synergistic photo-effects for EMI shielding. Flash irradiation with a broad spectrum causes a series of photoreactions in the PI film by simultaneously absorbing UV and Vis–NIR wavelengths, leading to the synthesis of porous graphene over a large area in a few milliseconds (ms). The UV wavelengths within the full spectrum induce defects in the PI film, which enhance the absorption of Vis–NIR wavelengths and promote photothermal reactions, resulting in the synthesis of porous graphene. The XPS and FTIR results demonstrate that the carbon content becomes dominant due to the gas release accompanied by a decrease in nitrogen and oxygen content. Raman spectra, XRD and TEM images show the formation of multilayer graphene through the rearrangement of carbon atoms in a disordered carbon matrix. At a lamp fluence of 22 J cmSupplementary file1 (PDF 2048 kb)Below is the link to the electronic supplementary material.


Two-dimensional (2D) materials and their vertically stacked heterostructures have attracted much attention due to their novel optical properties and strong light-matter interactions in the infrared. Here, we present a theoretical study of the near-field thermal radiation of 2D vdW heterostructures vertically stacked of graphene and monolayer polar material (2D hBN as an example). An asymmetric Fano line shape is observed in its near-field thermal radiation spectrum, which is attributed to the interference between the narrowband discrete state (the phonon polaritons in 2D hBN) and a broadband continuum state (the plasmons in graphene), as verified by the coupled oscillator model. In addition, we show that 2D van der Waals heterostructures can achieve nearly the same high radiative heat flux as graphene but with markedly different spectral distributions, especially at high chemical potentials. By tuning the chemical potential of graphene, we can actively control the radiative heat flux of 2D van der Waals heterostructures and manipulate the radiative spectrum, such as the transition from Fano resonance to electromagnetic-induced transparency (EIT). Our results reveal the rich physics and demonstrate the potential of 2D vdW heterostructures for applications in nanoscale thermal management and energy conversion. Radiative heat transfer between two objects in far-field contexts is bound by Stefan Boltzmann’s law of the black body. However, when the separation distance is less than the characteristic wavelength predicted by Wien’s law, the near-field radiative heat transfer is significantly enhanced due to photon tunneling of evanescent waves, exceeding the blackbody limit by several orders of magnitude, especially when the material supports surface wave resonances (such as surface plasmon polaritons and surface phonon polaritons) ,2,3. In Two-dimensional (2D) materials provide an excellent platform for the enhancement and manipulation of near-field thermal radiation due to their strong light-matter interactions in the infrared and have been attracting considerable attention in the last decade, including graphene ,23,24,25Graphene and 2D hBN, with the same crystal structure and naturally small lattice mismatch, are considered fundamental building blocks for 2D heterostructures. Graphene supports surface plasmon polaritons ,23,24 anIn this paper, we investigate the near-field thermal radiation of 2D vdW heterostructure vertically stacked by these two single-layer materials, focusing on the radiative spectral properties. We have demonstrated the contribution of the coupling between broadband plasmons in graphene and narrowband phonon polaritons in 2D hBN to the near-field radiative heat transfer and found an asymmetric line shape in the spectra, and confirmed it to be a Fano resonance arising from the interference between the continuum state (plasmons) and the discrete state (phonon polaritons). The Fano resonance has been widely discovered in spectra of various photonic nanostructures ,46 as weWe assume that the 2D vdW heterostructures are suspended, i.e., free-standing without a supporting substrate, that the lateral dimensions are much larger than the characteristic thermal wavelength so that the finite size effect can be neglected, and that the thicknesses are negligible. Let us assume that they lie on the  formula :(1)QT1,Tp- or s-polarization) of the incident and reflected electromagnetic waves. In this paper, only symmetric structures of cold and hot sides are considered, so Here, given by ,50:(2)TωIn this paper, all materials are non-magnetic, then The 2D vdW heterostructure in this study is considered as a random stack of monolayer graphene and 2D hBN, and neglecting thickness, its effective conductivity can be obtained by simply adding the conductivities of the two components : σeff=σGitten as :(5)σD=iωHere, given by :(7)σ2D-hken from ,43: τ−1=The real and imaginary parts of the optical conductivity of graphene and 2D hBN as a function of frequency are plotted in Based on the previous theoretical formalism, we performed numerical calculations of the near-field thermal radiation using MATLAB code.To understand the near-field thermal radiation of 2D vdW heterostructures, we first separately calculated the near-field radiative characteristics of graphene and 2D hBN. In For the case of graphene, two bright bands shown in the low-frequency region σE=D2The optical property and plasmonic behavior of graphene can be modulated by changing the chemical potential through chemical doping or electrical gating. In As the chemical potential of graphene increases PWhen the detuning frequency is 0, i.e., The preceding section primarily addressed the spectra of energy transfer functions In To conclude, 2D vdW heterostructures can achieve the same high radiative heat flux as graphene, yet with a markedly different spectral distribution, particularly at high chemical potentials.The previous considerations are mainly for suspended 2D vdW heterostructures; although it can be synthesized experimentally , it is mFor simplicity, we first consider non-dissipative substrates with constant dielectric constants For practical dissipative substrates, each substrate requires separate analysis because the optical response and the physical mechanism that interacts with the 2D vdW heterostructure are different for various materials . As an example, we use the widely used SiC as a substrate for illustration, as shown in , three parts of the bright band give rise to three peaks, with a valley at the frequency The plasmons in graphene are coupled to the surface phonon polaritons of SiC and the 2D phonon of 2D hBN. Hence, in the contour plot of 3 and LiNbO3 [In this paper, we have shown that near-field thermal radiation of 2D vdW heterostructure can be significantly enhanced and modulated by tuning the chemical potential of graphene . In d LiNbO3 , which hIn this paper, we investigate the near-field thermal radiation of 2D vdW heterostructures vertically stacked of graphene and monolayer polar material (2D hBN as an example). We show an asymmetric Fano resonance in the radiation spectrum resulting from the interference between a broad continuum of surface plasmon polaritons (SPPs) in graphene and a narrow discrete state of 2D phonon polaritons in 2D hBN. In addition, we show that 2D van der Waals heterostructures can achieve nearly the same high radiative heat flux as graphene but with markedly different spectral distributions, especially at high chemical potentials. By tuning the chemical potential of graphene, we can actively control the radiative heat flux of 2D van der Waals heterostructures and manipulate the radiative spectrum, such as the transition from Fano resonance to electromagnetic-induced transparency (EIT). Our results highlight the great potential of 2D vdW heterostructures to enhance and manipulate near-field thermal radiation, which may pave the way for highly efficient thermal management or design and optimization of energy conversion devices, such as thermophotovoltaics.


After the original 15-item Drive for Muscularity Scale developed by McCreary et al. in 2004, a more theoretically based scale that replicates the original DMS subscales with a better conceptual clarity and a shorter number of items, i.e., the DMS-9, has recently been developed by Chaba et al. in 2018. We sought to contribute to the literature especially under the Arab context, by investigating the psychometric properties of an Arabic translation of the DMS-9 in a sample of Arabic-speaking Lebanese university students of both genders.University students  from multiple universities in Lebanon were invited to fill the survey in this cross-sectional designed study (December 2022 and January 2023). Our sample was chosen using the snowball technique; a soft copy of the questionnaire was created using google forms software, and an online approach was conceived to proceed with the data collection.Using an Exploratory Factor Analysis-to- Confirmatory Factor Analysis strategy, we found that the original two-factor model of the DMS proposed in the parent study was adequately replicated in our sample. The two DMS-9 factor scores showed very good McDonald’s omega values (ω > 0.8). Findings also showed that gender invariance was achieved at the configural, metric, and scalar levels. Additionally, drive for muscularity scores correlated in the expected way with other study variables, providing support for the convergent and divergent validity of the Arabic DMS-9. Specifically, we found that greater drive for muscularity attitudes and behaviors significantly correlated with more severe muscle dysmorphic symptoms, inappropriate eating attitudes, muscle bias internalization, and lower body appreciation.Findings preliminarily suggest that the Arabic DMS-9 is psychometrically sound and suitable tool to assess the drive for muscularity construct among Arabic-speaking community adults. Making the Arabic DMS-9 available will hopefully benefit the scientific community working in Arab settings, promote local and international research in this area, and offer descriptive data on how drive for muscularity may interfere with health indicators in the general Arab population. Drive for muscularity is defined as a desire to develop a visible muscular physique by both increasing muscle mass and decreasing body fat, in order to achieve a muscular upper body and a narrow waist –3. StrivNo gold-standard measure of the drive for muscularity exists . SeveralSince its development, the DMS has been translated in different languages and adapted to different countries and languages, including Spanish , 24, ItaAlthough the 15-item DMS has been widely validated and extensively used in diverse research and clinical settings, and the findings that it relates to different relevant constructs  , this orTo date, no Arabic validation of the DMS exists to the best of our knowledge. Although research on disordered eating and body image disturbances has been widely developed in the Western world, the generalized globalization and westernization contributed to a rise in prevalence rates of these manifestations in people from non-Western cultures even exceeding those seen in Western people , 45. ResThrough the present study, we sought to contribute to the literature especially under the Arab context, by investigating the psychometric properties of an Arabic translation of the short 9-item DMS in a sample of Arabic-speaking Lebanese university students of both genders. As mentioned above, we chose the DMS-9 because of its better conceptual clarity and good psychometric qualities . Besides2 (SD = 4.12), ranging from 14.52 to 50.78 kg/m2. Most participants had a university level of education (88.8%).University students  from multiple universities in Lebanon filled the survey. Participants had a mean age of 24.46 years (SD = 6.60), ranging from 18 to 60 years and had a mean self-reported body mass index (BMI) 23.68 kg/mOther sample characteristics are displayed in Table Our sample was chosen using the snowball technique; a soft copy of the questionnaire was created using google forms software, and an online approach was conceived to proceed with the data collection (December 2022 and January 2023). The study’s main aims and goals, in addition to instructions for filling the questionnaire, were conveyed online for the participants, prior to their participation. Later, initial participants were asked to recruit other participants they know, preferably as diverse as possible regarding place of habitat within the Lebanese governorates. The questionnaire was anonymous and took between 15 and 20 min to complete. There were no credits received for participation. Inclusion criteria for participation included being of a resident and citizen of Lebanon of adult age. The “remove duplicates” option in excel ensured that the same answers were not submitted more than once. After providing digital informed consent, participants were asked to complete the instruments described below, which were presented in a pre-randomized order to control for order effects.The short form of the DMS-9 scale (9 items) was used in this study . ParticiThis scale is composed of 14 items scored on a 7-point Likert Scale . Higher scores indicate higher levels of muscularity bias internalization . This scnever) to 5  (score range 10–50) [Validated in Arabic , 64, thie 10–50) . Higher never) to 3  (score range 0–21). Higher total scores reflect greater disordered eating attitudes. In the present study, McDonald’s ω was 0.80 in the total sample.Participants were asked to complete the EAT-7, which has recently been validated in Arabic . This 7-Validated in the Arabic language , this scParticipants were asked to provide their demographic details consisting of age, gender, marital status, highest education level, self-reported height and weight to calculate the BMI, household crowding index  and phyThe DMS scale was translated to the official Arabic language, which is written and spoken across the Middle East and North Africa (MENA). The translation was performed with the purpose of achieving semantic equivalence between measures in their original and Arabic versions following international norms and recommendations . To thisThere were no missing responses in the dataset. To examine the factor structure of the DMS, we tested the original models proposed by McCreary et al.   was needed to perform the EFA according to Comrey and Lee . EFA wasCFA was conducted via the SPSS AMOS v.29 software using the maximum Likelihood estimation. The minimum sample size to conduct a CFA ranges from 3 to 20 times the number of the scale’s variables . TherefoTo examine gender invariance of DMS-9 scores, we conducted multi-group CFA  using SPComposite reliability in both subsamples was assessed using McDonald’s ω, with values greater than 0.70 reflecting adequate composite reliability . To asseThe fit indices of the one- and two-factor structure of the DMS-15 conducted in males and one-factor structure in females did not show appropriate fit Table .Consequently, we decided to conduct the EFA-CFA strategy to the DMS-9 items.2(36) = 1941.4, p < .001, and KMO (0.865) indicated that the DMS items had adequate common variance for factor analysis. None of the items had an MSA value < 0.5 and none of the items appeared repeatedly in different doublets; therefore, all items were kept in the analysis. The results of the EFA revealed two factors, which explained 81.44% of the common variance (item-factor loadings ≥ 0.62). The WRMR value was also adequate , indicating good fit of the model.Bartlett’s test of sphericity, χThe fit indices of the two-factor model of the DMS-9 scale  showed gMcDonald’s omega values were 0.90 for Factor 1 (Muscularity Behaviors) and 0.92 for Factor 2 (Muscularity body Dissatisfaction).The fit indices in Table Higher Muscularity behaviors and Muscularity body dissatisfaction scores were significantly associated with more muscle bias internalization, muscle dysmorphic disorder and inappropriate eating attitudes, and lower body appreciation Table .The present study was conducted with the aim of making available an Arabic psychometrically sound measure to assess drive for muscularity, i.e. the short 9-item DMS. The Arabic version was found to have excellent psychometric properties in terms of factorial structure, internal consistency, gender invariance, and convergent/divergent validity. These findings preliminarily suggest that the Arabic DMS is a simple, easy to use, and economic self-report scale for the reliable and valid assessment of drive for muscularity among Arabic-speaking community people.Using an EFA-to-CFA strategy as recommended in the literature , we founThe two DMS-9 factor scores showed very good McDonald’s omega values (ω > 0.8), higher than the 0.70 threshold value of good internal consistency suggested by previous researchers , 97, thuFinally, DMS-9 scores correlated in the expected way with other study variables, providing support for the convergent and divergent validity of the Arabic version of the scale. Specifically, we found that greater drive for muscularity attitudes and behaviors significantly correlated with more severe muscle dysmorphic symptoms, inappropriate eating attitudes, muscle bias internalization, and lower body appreciation. These results align with previous evidence. Similar evidence for validity of the DMS-9 through the same patterns of correlations with these variables has previously been reported in other validation studies . In addition, future cross-national validations in samples from different Arab countries are required to provide support to the cross-cultural validity of the scale. Finally, other important psychometric properties have not been addressed in this paper  and should be verified in future research.Through this study, we provide a brief, valid, economic and useful tool to evaluate drive for muscularity in Arabic-speaking men and women. Making the Arabic DMS-9 available will hopefully benefit the scientific community working in Arab settings, promote local and international research in this area, and offer descriptive data on how drive for muscularity may interfere with health indicators in the general Arab population.


Pitx3−/− mice, a genetic model of PD with early onset dopaminergic depletion in the dorsolateral striatum (DLS). This dysregulated BDNF/TrkB signaling might contribute to the pathophysiology of direct and indirect pathway striatal projection neurons in PD. Motor dysfunction is among the main symptoms of Parkinson’s disease (PD). This is closely linked to the loss of the neurotransmitter dopamine (DA) in the midbrain and in the striatum. Additionally, the ability of striatal neurons to undergo adaptive cellular alterations and synaptic plasticity is impaired. Dopamine receptor D1 (DRD1) stimulation is needed for the establishment of long-term potentiation (LTP) at synapses of striatal spiny projection neurons (SPNs), which leads to enhanced neurotransmission. In contrast, dopamine receptor D2 (DRD2) stimulation is needed for the formation of long-term depression (LTD) in SPNs, which leads to the opposite effect. The tropomyosin receptor kinase B (TrkB) and its ligand brain-derived neurotrophic factor (BDNF) are centrally involved in plasticity regulation at the corticostriatal neurons synapses. There are two populations of striatal SPNs, with different projection targets in the brain. DRD1 is expressed in direct pathway spiny projection neurons (dSPNs) and its activation enhances TrkB sensitivity for BDNF by increasing the levels of TrkB at the cell surface. In this study, we showed that the activation of DRD2 in cultured striatal indirect pathway spiny projection neurons (iSPNs) and cholinergic interneurons causes the retraction of TrkB from the plasma membrane. This provides an explanation for the opposing synaptic plasticity changes observed upon DRD1 or DRD2 stimulation. In addition, TrkB was found within intracellular structures in dSPNs and iSPNs in Pitx3−/− mice, a genetic model of PD with early onset dopaminergic depletion in the dorsolateral striatum (DLS). This dysregulated BDNF/TrkB signaling might contribute to the pathophysiology of direct and indirect pathway striatal projection neurons in PD.Motor dysfunction in Parkinson’s disease (PD) is closely linked to the dopaminergic depletion of striatal neurons and altered synaptic plasticity at corticostriatal synapses. Dopamine receptor D1 (DRD1) stimulation is a crucial step in the formation of long-term potentiation (LTP), whereas dopamine receptor D2 (DRD2) stimulation is needed for the formation of long-term depression (LTD) in striatal spiny projection neurons (SPNs). Tropomyosin receptor kinase B (TrkB) and its ligand brain-derived neurotrophic factor (BDNF) are centrally involved in plasticity regulation at the corticostriatal synapses. DRD1 activation enhances TrkB’s sensitivity for BDNF in direct pathway spiny projection neurons (dSPNs). In this study, we showed that the activation of DRD2 in cultured striatal indirect pathway spiny projection neurons (iSPNs) and cholinergic interneurons causes the retraction of TrkB from the plasma membrane. This provides an explanation for the opposing synaptic plasticity changes observed upon DRD1 or DRD2 stimulation. In addition, TrkB was found within intracellular structures in dSPNs and iSPNs from  Parkinson’s disease (PD) is the second most common neurodegenerative disorder and is characterized by a progressive loss of dopaminergic neurons in the substantia nigra (SN) ,2,3,4. TDefective dopaminergic signaling from the SN to the basal ganglia plays a central role for these altered motor functions. Dopamine (DA) released from the nigrostriatal terminals onto spiny projection neurons (SPNs) in the dorsolateral striatum (DLS) modulates circuit activity and plasticity, including long-term potentiation (LTP) and long-term depression (LTD) at the glutamatergic synapses between the corticostriatal afferences and striatal SPNs ,14,15,16s/olf subunit and, upon DA binding, activates adenylyl cyclase 5 (AC5), which facilitates cyclic AMP (cAMP) production ; RRID: MMRRC_000230-UNC). DRD2-eGFP mice were bred on a FVB/NJ genetic background.All experiments were approved by a license for animal testing  and performed in accordance with supervision through the local veterinary authority (Veterinaeramt der Stadt Wuerzburg) and Committee on the Ethics of Animal Experiments, i.e., Regierung von Unterfranken, Wuerzburg, Germany.Pitx3−/− mice (RRID: IMSR_JAX:000942), and brain tissue was dissected from adult (P56-P84) male and female mice. DRD1-td-Tomato x Pitx3−/− mice were bred on a C57Bl6/J genetic background.For IHC analyses, BAC-transgenic DRD1-td-Tomato mice (Tg(Drd1a-tdTomato)6Calak; RRID: IMSR_JAX:016204) were crossed with wild-type or with Pitx3−/−mice were provided by Dr. Rosario Moratalla, Instituto Cajal, Consejo Superior de Investigaciones Cientificas (CSIC), 28002 Madrid, Spain. The BAC transgenic mice were used to identify the striatal direct and indirect pathway neurons. All experimental procedures were approved by the Cajal Institute Bioethics Committee and by the CSIC Ethics Committee and fulfilled the requirements of Spanish (RD 53/2013) and European Union (63/2010/EU) legislation.DRD1-td-Tomato x wildtype/All mice were genotyped via PCR analysis. Mice were housed in groups of 4–6 per cage, with a constant room temperature (21–22 °C) and 12 h light/dark cycle (lights on at 7:00 h) and were given free access to food and water. The studies with rats were performed as previously reported . In brieFor this procedure, 6-OHDA injections were performed as previously described . BrieflyStriatal primary cultures were prepared from P1–3 newborn pups, as described previously . BrieflyEnrichment of primary DRD2-eGFP neurons was accomplished via flow cytometry-based cell sorting using the MACSQuant Tyto Cell Sorter . First, the sample was prepared through dissection and dissociation of the striatum, as described above. Prior to sorting, around 2–2.5 mL of cell suspension was loaded into the cartridge. For this procedure, the cartridge  was prepared by wetting all the channels interconnecting the chambers with NB media supplemented with serum. Additionally, 170 µL of medium was added to the positive collection chamber to provide a suitable and nurturing environment to the sorted DRD2-SPNs. Before adding the cell suspension into the input chamber, the cells were filtered using a 20 µm pre-separation filter  in order to remove cell aggregates and large particles that could clog the channels and hinder the sorting. Gating was set up to detect the GFP-positive neuronal population. The sorting procedure was performed at 25 °C, using a mixing speed of 100 rpm and the automatic pressure adjustment. Thereafter, the purified neurons were resuspended in an appropriate volume of media supplemented with horse serum and immediately plated on polyornithine/laminin-111-coated cell culture dishes. The cell culture medium was completely replaced with a serum-deprived cell culture medium after 24 h to prevent any serum-related effect on the dopamine receptors or TrkB stimulation. Cell cultures were maintained for 3–14 days in vitro (DIV) before preparation for ICC and Western blotting (including surface biotinylation). Enriched DRD2-eGFP neurons were pharmacologically treated with 500 nM sumanirole maleate  and/or stimulated with 10 ng/mL BDNF (homemade). After stimulation, the cells were washed with 1 × PBS before further processing. Primary striatal neurons (DIV5-DIV10) were washed once with 1× PBS and fixed with 4% paraformaldehyde  in 1 × PBS  for 15 min at RT. Subsequently, they were washed twice with 1 × PBS for 5 min. Permeabilization and blocking was performed with 0.1% Triton X-100  and 10% donkey serum  in 1 × PBS for 1 h. For immunostaining, the following antibodies were applied in a blocking solution overnight at 4 °C: chicken anti-GFP , goat-anti-mTrkB , rabbit anti-DARPP-32 , mouse anti-ß-III-tubulin , goat anti-ChAT , rat anti-Lamp-1 , and sheep anti-mouse SORCS-2 . After washing three times with 1XPBS for 10 min, primary antibodies were detected with the following secondary antibodies (incubation: 60–90 min at RT): donkey anti-chicken Alexa Fluor 488 , donkey anti-mouse Alexa Fluor 647 , donkey anti-rabbit Cy3 , donkey anti-rabbit DyLight550 , donkey anti-goat Alexa Fluor 647 , donkey anti-rat Alexa 405 , and donkey anti-sheep Alexa Fluor 647 . Nuclei were stained with 0.4 mg/mL 4′,6-diamidino-2-phenylindole (DAPI). Coverslips were again washed three times for 10 min with 1× PBS and the cells were mounted on Superfrost_Plus glass slides  using MERCK-FluorSave reagent . For cell surface staining, after stimulation, the media was discarded and cold media with the goat anti-mTrkB  antibody was added to the wells. The cells were incubated for 30 min at 4 °C. Afterwards, the cells were washed twice with ice-cold PBS and blocked for 1 h at RT. Then, incubation with the following other primary antibodies was performed: chicken anti-GFP , rabbit anti-DARPP-32 , mouse anti-GFAP , and rabbit anti-GFAP .The mice were anesthetized with pentobarbital  and then transcardially perfused with 4% paraformaldehyde dissolved in PB , dissected, and the brains were immersed overnight in the same fixative solution. The brains were washed and kept in PBS until they were used. The brains were then embedded in 6% agarose  and 40 mm free-floating, coronal brain sections were obtained using a LEICA Vibratome VT1000S (RRID:SCR_016495). Sections were stored in “Cryoprotection Anti-Freeze Buffer” , and ethylene glycol  at −20 °C. 2O, and mounted on SuperfrostPlus glass slides  with FluorSave mounting reagent .Immunohistochemistry was performed using a free-floating staining protocol previously described . In brieCell surface proteins were biotinylated using the Pierce Cell Surface Protein Isolation kit , according to the manufacturer’s instructions. Between 150,000 and 300,000 cultured neurons were required per condition. The neurons were plated on 12 mm glass coverslips and biotinylated on either DIV7 or DIV8. The cells were washed with 1× cold PBS and incubated with EZLINK Sulfo-NHS-SS-biotin at 4 °C for 30 min, followed by the addition of a quenching solution. The cells were then lyzed with lysis buffer containing a protease inhibitor cocktail , sonicated one time, and centrifuged. The cell lysate was applied to the NeutrAvidin agarose gel. The flow through (intracellular protein fraction) was saved for Western blotting. The biotinylated cell surface proteins in the NeutrAvidin agarose gel were pulled down with sodium dodecyl sulfate (SDS) sample buffer (pH 6.8) with 50 mM dithiothreitol (DTT). Each sample was mixed with Laemmli buffer, heated to 95–99 °C for 5 min, and centrifuged shortly before loading onto 5–11% polyacrylamide gel. Polyacrylamide gel electrophoresis was performed in an electrophoresis cell chamber  at 70 V for the stacking gel; then, the voltage was changed to 120 V once the samples reached the separating gel. The proteins were then transferred via wet blotting onto polyvinyl difluoride (PVDF) membranes  in a blotting cell  for 1 h at 4 °C with 135 V and 35 mA in blotting buffer . The membranes were blocked for 1–2 h with 5% milk powder in 1× TBS-T at RT and then incubated with either primary goat polyclonal antibodies , rabbit polyclonal anti-phosphoTrkB (Tyr816) , and rabbit anti-phospho TrkB ), chicken polyclonal anti-Calreticulin , or goat-anti DARPP-32  overnight at 4 °C on a rocker. Following this step, incubation with horseradish peroxidase (HRP)-conjugated secondary antibodies was performed ). For the relative quantification of the effects of sumanirole treatment on the surface distribution of TrkB, the signal densities in the intracellular fractions, the membrane protein fractions, and the intracellular protein fractions were normalized to those of the marker proteins for their respective fractions.Image acquisition was performed with an Olympus FluoView 1000 confocal laser microscope . The different color channels were visualized using lasers with the wavelengths of 405 nm, 488 nm, 559 nm, and 633 nm. The microscope was equipped with the following objectives: HC PL APO CS2 40× (oil NA: 1.30) and HC PL APO CS2 60× . Acquisition was performed using the Olympus FV10-ASW 3.0 (RRID: SCR_014215) software ; z-stacks of different focal planes were obtained; images were saved in the Olympus.oib format and analyzed with ImageJ image processing software version 1.53t  (RRID: SCR_003070). For visualization, the z-stacks were projected as a maximum intensity projection. The line scan colocalization analysis was performed using ImageJ. Brightness and contrast modifications were performed according to the rules of good scientific practice and the changes were documented for every image that was processed.For the quantification of TrkB clusters, the number of TrkB-positive puncta were manually counted by the experimenter. TrkB cluster numbers were then normalized to the investigated area. Allocation to the D1- or D2-SPNs was conducted via colocalization of the TrkB clusters with either td-Tomato or Met-Enkephalin, respectively. t-test was performed, and for more than two groups a one-way ANOVA with Tukey’ multiple comparison test was applied. For data where a control group has been set to 100%, a one sample t-test was performed. For all tests, a confidence level of p ≤ 0.05 was set. For the statistical analysis, GraphPad Prism software version 6.01  (RRID: SCR_002798) was used. The type of each statistical test used has been indicated in the respective figure legends.For all data used for quantification purposes, a normal distribution was assumed based on experience with similar kinds of experiments. For two groups, a two-sample To investigate DRD2-mediated effects on the translocation of TrkB in indirect pathway iSPNs, we developed and tested a protocol for the culture of flow cytometry-enriched DRD2 SPNs . Our sorDue to their important functional role in the control and modulation of SPNs in the striatum and the presence of DRD2 on their cell surface , we werei-coupled receptors would induce the inhibition of adenylyl cyclase activity and a progressive downregulation in cAMP levels. This decrease in cAMP could prevent TrkB from being translocated to the cell surface. In order to study such effects, we performed surface biotinylation and cell surface immunostainings of TrkB in the presence or absence of sumanirole. Biotinylation of cell surface proteins of DRD2 neurons treated under different time courses showed that TrkB surface levels decreased after 10 min of treatment with sumanirole and decreased further over time for up to 24 h (t-test was performed. Control: mean 100% (n = 7); 10 min. Sum: mean 77.6%, p = 0.05 (n = 7);; 60 min. Sum: mean 75.8%, p = 0.19 (n = 4); 6 h. Sum: mean 31.5%, p = 0.08 (n = 2); Sum 24 h: mean 28.0%, p = 0.06 (n = 2)). Confocal imaging also showed that DRD2’s activation with sumanirole led to a marked decrease in TrkB on the cell surface in comparison to their DA-depleted controls (t-test was performed. Control: mean 100% (n = 7); Sum + BDNF 10 min: mean 46.64, p = 0.006 (n = 7); Sum + BDNF 60 min: mean 95.37%, p = 0.807 (n = 3); Sum + BDNF 24 h: mean 114.2%, p = 0.728 (n = 3)). We also assessed the ratio of pTrkB to cell surface TrkB, as seen with the surface biotinylation experiments at 10 min, 60 min, and 24 h treatment of sumanirole. A gradual increase in pTrkB/surface TrkB was observed . Bafilomycin treatment for 10 min  or 4 h  did not enhance the colocalization of TrkB with SORCS2, indicating that a reduced fusion of endosomes with lysosomes has no positive effect on the recycling of TrkB to the cell surface.TrkB is usually present on the dendritic spines of iSPNs that receive inputs from BDNF-positive corticostriatal afferences. We therefore investigated how the activation of DRD2 with sumanirole affects dendritic TrkB in cultures of enriched DRD2-expressing striatal neurons. Activation of D2 receptors by sumanirole rapidly reduced cell surface TrkB expression in the soma and dendrites of iSPNs A. Blockip = 0.0011) and 4 h (mean: 61.2% (n = 5); one-way ANOVA with Tukey’s multiple comparisons test, CTR vs. 4 h Baf: p < 0.0001) of bafilomycin treatment (p = 0.0186), but the simultaneous treatment with bafilomycin and sumanirole could not further increase the bafilomycin-induced TrkB–lysosome association (mean: 63.8% (n = 2); one-way ANOVA with Tukey’s multiple comparisons test, CTR vs. 4 h Baf, 10 min. Sum: p = 0.0015). This indicates that TrkB is degraded by lysosomes in the cell soma. Taken together, these data demonstrate that TrkB is rapidly retracted from the cell surface both in the dendrites and cell soma after the activation of DRD2 with sumanirole, and that dendritic TrkB is retrogradely transported to the soma, where these TrkB-positive vesicles then fuse with lysosomes. When we investigated the effect of sumanirole treatment on TrkB distribution in the cell body of DRD2-expressing neurons in enriched cultures of striatal iSPNs, we found a rapid uptake of TrkB A and thereatment B,C. FurtPitx3−/− mouse model, we investigated TrkB distribution in striatal tissue slices from these mice at an age range from 6 months to 11 months. We found that TrkB clusters arose in the striatum of Pitx3−/− animals : 14.4; mean Pitx3−/− (n = 6): 28.9; unpaired t-test: p = 0.0376) higher number of these clusters than sections from wild-type control animals . Also, there was no significant change in the number of TrkB clusters after prolonged DA depletion .We previously showed that the subcellular distribution of TrkB in SPNs is altered upon dopamine depletion in a rat model in which 6-OHDA is unilaterally injected into the medial forebrain bundle . In this animals A and tha animals B. We fur/− model C,D. Thise lesion E,F, and p = 0.0003; 2 wk p.l. intact vs. 4 m p.l. intact: p = 0.0007; 2 wk p.l. lesioned vs. 1 m p.l. lesioned: p = 0.0574; 2 wk p.l. lesioned vs. 4 m p.l. lesioned: p = 0.0047; 2 wk p.l. intact vs. 2 wk p.l. lesioned: p = 0.9374; 1 m p.l. intact vs. 1 m p.l. lesioned: p = 0.1132; 4 m p.l. intact vs. 4 m p.l. lesioned: p = 0.9921) as well as in layer V  of the motor cortex, both of which are areas that send projections to the striatum. Interestingly, there was a trend towards a reduced number of BDNF-positive cells in the motor cortex of the lesioned hemisphere when compared to the intact hemisphere from 1 month after the lesion onwards  release from nigrostriatal terminals onto SPNs in the DLS modulates circuit activity and long-term plastic changes, including long-term potentiation (LTP) and long-term depression (LTD) at the glutamatergic synapses between corticostriatal afferences and striatal SPNs ,13,14,15in iSPNs . This prHere, we also showed that TrkB’s localization in DRD2 neurons can be further modulated. Chronic activation of D2 receptors in vitro was achieved by stimulating enriched DRD2 neurons in culture over a period of 24 h or longer. This led to a further reduction in TrkB receptors at the cell surface of DRD2 striatal neurons. In parallel, intracellular levels of TrkB increased. This suggests that not only was TrkB’s translocation to the cell surface inhibited, but that the existent surface receptors might have been internalized, consequently reducing sensitivity to BDNF after acute stimulation of D2 receptors. Altered TrkB phosphorylation was already observed at 5 min after treatment with sumanirole. Interestingly, at 60 min and 24 h of treatment with this DRD2 agonist, no further reduction in pTrkB was observed. At that time point, pTrkB levels increased again when related to the levels of cell surface-exposed TrkB. This could reflect a cell autonomous mechanism that is due to a sensitization of the TrkB receptors left at the cell surface after undergoing a long period of stimulation. Long-term D2 receptor stimulation with sumanirole could lead to a compensatory or adaptive mechanism specific for DRD2 striatal neurons against chronic D2 agonist exposure. This effect of enhanced TrkB phosphorylation, despite low levels of TrkB at the cell surface after long-term stimulation with a DRD2 agonist, might as well be explained by an additional non-cell-autonomous effect due to the presence of DRD2-expressing astrocytes in the sorted D2 receptor-expressing cells . AstrocyWe have also shown that 7.9% of the DRD2-expressing sorted striatal neurons are cholinergic interneurons. These interneurons regulate the synaptic plasticity and excitability of striatal SPNs and are thought to induce inhibition in these cells. Additionally, cholinergic interneurons are characterized by a multiphasic change in activity, showing a pause in firing which is triggered by aversive and salient stimuli. These pauses have been associated with changes in DA neuron activity. Furthermore, a blockage of D2 receptors in the cholinergic interneurons inhibited the generation of the pause, and the upregulation of D2 receptors prolonged the pause, which is associated with delayed inhibitory learning . The chrPitx3−/− mice, acute depletion of dopaminergic afferents in 6-OHDA-treated rodents, as well as chronic levodopa therapy in PD models that induce dyskinesia have shown to induce changes in the morphology and synaptic activity of SPNs from both direct and indirect pathways [Overall, many changes take place in SPNs during the course of PD pathophysiology. Long-term dopamine depletion from early developmental stages in pathways ,73,92,93pathways . On the pathways . These sPitx3−/− mice. Pitx3−/− or aphakia mice represent a genetic model of PD, in which most of the dopaminergic neurons from the substantia nigra do not develop properly. Consequently, dopaminergic innervation of the DLS is massively reduced from early developmental stages onwards. When we examined TrkB expression in striatal brain slices of adult Pitx3−/− mice, we found that they showed an increase in perinuclear TrkB cluster formation compared to their wild-type littermates. These clusters were apparent in both dSPNs and iSPNs in the DLS. Interestingly, the aggregation of TrkB was not restricted to the dSPNs, as opposed to what we have previously shown for 6-OHDA-treated mice and rats [Furthermore, we studied the effect of DA depletion in DRD2 striatal neurons and the subsequent alterations of TrkB expression under pathological conditions of chronic DA depletion in and rats . We systPitx3−/− mouse model, the animals already lack striatal dopaminergic inputs during early development. This intrinsic lack of DA in the DLS could lead to developmental compensatory mechanisms counteracting potentially harmful changes in the motor network. This is also reflected by the absence of an apparent motor phenotype in these animals [Pitx3−/− mice.A possible explanation for this finding is the differences between the two models. In the 6-OHDA model, dopaminergic denervation occurs rapidly within a few days after the 6-OHDA injection, while in the  animals . Thus, T2A receptor signaling. This would render iSPNs less prone to TrkB cluster formation, as intracellular TrkB levels are decreased compared to dSPNs in a DA-depleted state. In the Pitx3−/− mice, there might be a compensatory downregulation of A2A receptors to protect the iSPNs from excitotoxic signaling, thus reducing this protective effect on TrkB cluster formation. It is apparent that TrkB clusters form upon DA depletion in both models, but iSPNs are spared from TrkB cluster accumulation in the 6-OHDA model. Potentially, dSPNs are more vulnerable to DA depletion than iSPNs, which could explain that TrkB cluster formation appears exclusively in the dSPNs in this model. This could be due to the distinct signal transduction processes of the different dopamine receptors. Under physiological conditions, D1 receptor activation leads to TrkB surface expression by shuttling TrkB from the intracellular stores to the cell surface. When this action is missing, TrkB can no longer be transported to the cell surface and is retained in the cell soma, favoring TrkB cluster formation. Conversely, D2 receptor activation normally leads to decreased TrkB surface expression. When this mechanism is impaired through DA depletion, TrkB surface expression is favored, which is likely facilitated via still intact APitx3−/− mice. This might be due to the low abundance of these DRD1/DRD2-coexpressing cells. Also, we cannot exclude the possibility that these cells might exhibit mixed responses to DA depletion, leading to a more complex effect on TrkB translocation. We did not investigate this in the 6-OHDA rat model, as D1-SPNs could not be directly identified by suitable markers, making it impossible to identify DRD1-expressing neurons that also express DRD2 or Met-Enkephalin.Furthermore, in the striatum, there is also a SPN population which expresses both DRD1 and DRD2 simultaneously ,95,96. HInterestingly, BDNF expression in the cortex increased gradually after 6-OHDA injection both on the ipsilateral lesioned side and the contralateral non-lesioned side. As BDNF itself is crucial for motor learning and execution , and sin−/− model are not directly comparable due to their different natures and degrees of DA depletion within the striatum. The conclusion from one model cannot be directly transferred to the other. However, we see these two models rather as complementary to each other to investigate how DA depletion affects striatal pathology either in an acute or in a chronic manner. Furthermore, the number of animals used for the quantification of the 6-OHDA experiments was low (n = 3). This was accounted for using non-parametric statistical testing when the data were assumed to be not normally distributed. Western blot analyses showed strong tendencies; however, due to low n numbers, no statistical significance could be reported in some cases.There are several limitations in our study. We identified the presence of DRD2-expressing cholinergic interneurons in our cell culture model, but how the activation of DRD2 with sumanirole specifically in these cells contributes to TrkB internalization in this and other striatal cell populations has not been studied. For this, further investigations with striatal organoid or organotypic slice cultures are needed for being able to investigate the responses of these interneurons in a synaptic context with D1 and D2 dopamine receptor-expressing SPNs. Also, the 6-OHDA rat model and the Pitx3Pitx3−/− model of early onset dopaminergic depletion in the dorsolateral striatum, we found TrkB clusters within intracellular structures in the iSPNs and dSPNs. Thus, dendritic and spine atrophy correlate with altered TrkB cell surface expression, indicating that dysregulated BDNF/TrkB signaling contributes to the pathophysiology of direct and indirect pathway striatal projection neurons in PD, and that long-term stimulation of DRD2 might re-establish TrkB phosphorylation to basal levels, rescuing spine morphology. This regulatory mechanism might be crucial for positive or negative modulation of corticostriatal synaptic plasticity. Altogether, these findings provide new insights about the disease pathomechanisms and possible therapeutic targets.In summary, we found that the activation of D2 receptors in iSPNs reduces TrkB cell surface levels. Upon retraction from the cell surface, TrkB accumulates in intracellular clusters and associates with Lamp-1-positive lysosomal structures. Biotinylation experiments indicated that this reduction in TrkB increases over time, suggesting a role for iSPNs in the complications induced by chronic treatment with levodopa. Interestingly, TrkB cell surface reduction did not affect TrkB phosphorylation levels after long-term treatment with a DRD2 agonist, which might explain why previous studies have reported that high doses of levodopa that induced dyskinesia rescue spine morphology in iSPNs. Additionally, using a


Graphene presents practical applications in energy storage devices, especially supercapacitors. However, mainstream synthesis of graphene includes toxic chemical usage, which threatens the environment. With the recent attention shift to synthesizing nanomaterials from agro-waste due to their easy availability, cost-effectiveness, and, most importantly, their environmental friendliness, we present, in this work for the first time, a novel and green synthesis of few-layered graphene sheets using pomegranate peels as a precursor at a low temperature of 80 °C. The surface morphology and microstructural properties are determined by Transmission Electron Microscopy (TEM), Energy Dispersive X-Ray spectroscopy (EDX), X-Ray Diffraction (XRD), Fourier Transform Infrared Spectroscopy (FTIR), UV–visible spectroscopy (UV–vis), and the electrical properties determined by Hall Effect Measurement. The application as a supercapacitor is also examined using Cyclic Voltammetry (CV), Charge–Discharge Cycling (GCD), and Electrochemical Impedance Spectroscopy (EIS). The resulting supercapacitor delivers an areal capacitance of  Graphene is a unique nanomaterial with zero band gap4 and the only carbon allotrope where carbon atoms are tightly bonded to their neighbors by a unique electronic cloud deviating slightly from quantum mechanical principles5. These electrons move freely between the 2pz orbitals1, explaining why graphene exhibits unparalleled electrical conductivity5.Graphene is a two-dimensional planar structure of s6. Graphene’s unique large surface area finds practical applications in energy storage, specifically batteries and supercapacitors7. Despite all these excellent properties and uses, the synthesis of high-quality, large-area graphene in a cost-effective way remains a significant challenge. Graphene was first isolated through mechanical exfoliation8, which has remained the typical conventional method of graphene synthesis8. Also, synthesis has been carried out via routes such as chemical exfoliation, chemical synthesis, thermal chemical vapor deposition, and unzipping carbon nanotubes9.Graphene exhibits excellent electronic, mechanical, and thermal properties10. The process’s cost of highly pure graphite remains another challenge11. Consequently, there is an attention shift to the synthesis of nanomaterials and graphene from natural sources12, most especially agro-waste14. Apart from easy availability and access to agro-waste, synthesis from agro-waste is straightforward, safe, and typically produces more stable materials15. They also offer an effective way of waste management16. Consequently, there has been the synthesis of nanomaterials and graphene from waste in general and agro-waste as well17,18 and used in electrochemical storage devices, wastewater treatment, and metal ion sensing, among others10. For example, graphene nanosheets were obtained by oxidizing graphite into graphene oxide using the Hummer method and highly concentrated acids, then using pomegranate juice as a reducing and capping agent to reduce graphene oxide19. Also, rice husk was combusted at 1123 K and chemically treated to obtain graphene sheets20. Furthermore, green tea-synthesized graphene sheets using a high-temperature pyrolysis technique have also been reported21. In all these works, graphene was obtained at elevated temperatures, and the synthesis process includes several steps, including chemical treatment.A significant drawback to the conventional graphene synthesis method involves using chemicals that are usually highly corrosive, explosive, and toxic22 and are most readily available as waste, globally estimated to be 1.62 million tons22. The pomegranate peel contains a variety of bioactive substances, including hydrolyzable tannins, flavonoids, complex polysaccharides, and minerals, with gallic acid and punicalagin being the major phenolic compounds which makes them useful in many fields23. Although the peels have been utilized in synthesizing carbon dots for several applications28, the low-temperature synthesis of graphene from the pomegranate peel is yet to be reported.Pomegranate peels constitute about 43% w/w of the entire fruitHerein, we report on a one-pot synthesis of few-layered graphene sheets from a green and abundant precursor of pomegranate peel which was used solely to produce graphene, where no industrial chemicals were used in the synthesis process. The latter makes our method entirely green and energy efficient as the synthesis temperature does not exceed 80 °C. The morphology, microstructure, and electrical properties are determined, and the potential application as a supercapacitor is also examined.Pomegranate fruit was sourced from a local market. The peel was removed and washed thoroughly with distilled water to remove the dust and impurities. The clean peels were subsequently dried in an oven at 40 °C overnight and blended into powder form using a blender.The few-layered graphene sheets were synthesized via the hydrothermal method as follows:A mass of 0.3 g of fine pomegranate peel powder was dissolved in 45 ml of distilled water and sonicated for 60 min. The solution was then transferred into a 750 ml Teflon-Line stainless steel autoclave and heated at 80 °C for 10 h. The solution was allowed to cool naturally, and filtration using a syringe and a 0.22 μm filter membrane was done for two turns. The solution was further filtered using a dialysis membrane against deionized water for 48 h. The resultant supernatant was stored at 4 °C for further use.Quartz and FTO substrates were used in the studies for electrical measurements. In cleaning the substrates, firstly, the substrates were dipped in a solution of 2 ml of Hellmanex III and 200 ml of hot distilled water and sonicated for 10 min. The substrates were then rinsed thoroughly with distilled water. The substrates were further cleaned using an ozone cleaner for 10 min at a temperature of 24 °C. Thin film deposition of the few-layered graphene sheets was achieved by drop casting on the substrate and allowing it to air dry.29. One significant benefit of LIG is high conductivity30, a required condition for substrate selection in making electrodes for electrochemical measurements. Therefore, as part of this work, we also aim to examine the practicality of LIG as conducting substrates.Electrochemical measurements were performed on specially designed electrodes fabricated by the Laser-Induced Graphene (LIG) technique. The LIG is a graphene synthesis technique that involves exposing a polymeric precursor to laser light to cause photochemical and thermal conversion into grapheneLIG electrode patterning was achieved in one step on a Kapton paper by using a high-powered 200 μl of the synthesized graphene was then drop-casted on LIG, which functioned as the conducting substrate for our synthesized graphene. After drying at room temperature for 48 h, a gel electrolyte layer was deposited, and the sample was dried for 6 h. Gel electrolyte was made by dissolving 5 g of PVA powder in 5 ml of The microstructure of the samples was studied using a high-resolution transmission electron microscope (TEM), JEM-2100F from JEOL Company, fortified with a 200 kV field emission gun. The elemental analysis was achieved by EDX, using an Oxford instrument 80 Experimental research and field studies on plants (either cultivated or wild), including the collection of plant material, must comply with relevant institutional, national, and international guidelines and legislation.Figure The elemental composition estimated by Energy dispersive X-Ray spectroscopy (EDX) is tabulated below. From Table 31. For the 2D band, three peaks are observed at ~ 2834.42, 3087.70 21. Also, the observed 2D band at 2834.42 cm–1 is blue-shifted due to the strain and vibrational phonons related to impurities32. On the other hand, the combinations of deconvoluted peaks at 3087.7 cm-1 and 3199.02 cm-1 can be ascribed to the D + G band33. The G to the 2D band ratio (i.e. IG/I2D) was also calculated to be 0.98, indicating layer numbers n < 534, which is consistent with that obtained from the HRTEM images.Figure 35. The broad spectrum observed is due to the amorphous nature of the synthesized few-layered graphene sheets36, specifically, the disorder of graphite layers caused by empty rooms between graphite layers37. The broad peak is also observed due to the broadening of the d spacing21.The XRD pattern of the synthesized few-layered graphene sheets is presented in Fig. 38, and the peak located at 364 nm is attributed to the n-π* transitions of the fewer C=O bonds present.UV–vis spectra of the synthesized few-layered graphene were studied using the U-3900 spectrophotometer in the 200–800 nm wavelength range. As shown in Fig. –OH groups and the absorbed water molecules. The peaks observed at 2925.10 COOH group stretching vibration and C=C skeletal vibrations from graphitic domains in the ketone or quinone groups. The weak peak observed at 1380.91 C–O, and lastly, the observed peaks between 721.996 and 570.56 C-H41.Figure Oxygen functional groups on the surface of the synthesized few-layered graphene sheets make it hydrophilic, explaining its solubility in water.To examine the electrical properties of the synthesized few-layered graphene sheets, Hall Effect measurements were conducted. Figure The pomegranate peels consist mainly of hydrolyzable punicalagin, a bioactive compound with a chemical structure including gallagic acid and ellagic acid connected through a glucose molecule. This compound has a common feature, the presence of labile oxygen linkages, responsible for the cyclization and metamorphosis of the punicalagin molecules. Most likely, the heating temperature at 80 °C and the pressure inside the autoclave are capable of hydrolyzing punicalagin and breaking these labile oxygen linkages resulting in the release of ellagic acid. Therefore the reaction proceeds by the breaking of the labile bonds, followed by the release of 42. Rectangular-shaped CV curve also indicates low contact resistance and ideal charge propagation of the electrodes43.Cyclic voltammetry was performed for the LIG Fig. 44. A plot of the Capacitance versus the Scan rates is shown in Fig. 43. Also, at low scan rates, a more homogeneous potential with minimal variations is created along the material45. The specific areal capacitance at 20 m 46.Comparatively, LIG + PG has a larger CV area, indicating higher capacitance and better rate capability. Also, from Fig. The GCD behavior of the samples was studied at various current densities within the range of 10–50 μA 45. Voltage drop for LIG was found to be 0.11 V, and that of LIG + PG was found to be 0.082 V, with the difference equating to 0.028 V. This indicates that our synthesized few-layered graphene sheets contributed a relatively small amount, almost negligible, to the voltage drop.A sharp voltage drop is observed at the beginning of discharge for both samples, which is due to the diffusion-limited mobility of the electrolyte ions in the electrode pores and could result from the interface resistance between the current collector and the active layer or the conductivity of highly porous activated carbon47 and thus indicating good capacitor behavior of our synthesized graphene. A summary of the electrochemical performance of the sample can be found in Table A plot of the areal specific capacitance versus current density is shown in Fig. entclass1pt{minima42, corroborating the results obtained in the CV measurements.EIS was further used to characterize the performance of the graphene samples. The experiment was performed in the frequency range of 100 MHz to 0.1 Hz. Figure 46. The charge transfer resistance (46. Our synthesized graphene's relatively lower resistant values make it an excellent choice for energy storage since lower resistant values are necessary for minimizing supercapacitor losses and increasing overall efficiency.EIS plot fitting was done using the EIS Spectrum Analyzer to get the equivalent circuit model of the EIS plot, shown in Fig. In summary, we have successfully developed for the first time a green, facile, and energy-efficient method of synthesizing few-layered graphene sheet from pomegranate peels via the hydrothermal method at a low temperature of 80 °C without the aid of any passivating agent. The microstructure, morphology, and electrical properties studies show that our synthesized graphene is of high quality. Excellent electrochemical performance has also been achieved with the areal capacitance of 3.39 mF