Assessing the impact of distal femoral morphology using Citak's ratio: an independent risk factor for aseptic loosening in rotating hinge knee prosthesis.

BACKGROUND: Aseptic loosening (AL) is a frequent complication after rotating hinge knee (RHK) prosthesis. Citak's ratio has recently been developed to describe and classify distal femoral morphology into 3 groups (A, B, C). It consists in a ratio between the diameters of the femoral canal at 20 cm from the knee joint line and at 2 cm from the adductor tubercle. The objective of the study was to identify whether the femoral distal anatomical shape described with this ratio represents a risk factor for AL in RHK prosthesis. METHODS: Retrospective study of patients who had undergone primary or revision RHK prosthesis, with a follow-up of minimum 4 years. Citak's ratio was calculated, and patients were classified depending on its value. Univariate and bivariate statistical analysis was performed to identify AL risk factors. Receiver Operating Characteristics (ROC) analyses were conducted to examine diagnostic quality of the parameters of interest. RESULTS: Thirty-three patients were included. Most of them females (ratio 26:7), with a mean age of 78.2 (SD 6.9). Three patients presented AL (rate of 9%), all of them classified into group C (100%). Citak's ratio was significantly related to the AL rate (p < 0.001), and so was the femoral canal diameter at 20 cm from the knee joint (p 0.010). The ROC curve analysis yielded an Area Under the Curve (AUC) of 0.922 (CI 95% 0.819-1.000) for the Citak´s ratio. CONCLUSION: The inner femoral diameter at 20 cm proximal to the knee joint line and Citak's ratio help indentify patients at risk of AL after RHK prosthesis, and thus a better planning of the surgery.

Osmotic and metabolic responses to cold acclimation and acute cold challenge in a freeze avoidant lizard, Podarcis siculus.

Ectotherms survive exposure to subzero temperatures through freeze tolerance or freeze avoidance. Among vertebrate ectotherms, glucose is commonly used as a cryoprotectant in freeze tolerant strategies and as an osmolyte in freeze avoidant strategies, while also functioning as a metabolic substrate. Whereas some lizard species are capable of both freeze tolerance and freeze avoidance, Podarcis siculus is limited to freeze avoidance through supercooling. We hypothesized that, even in a freeze-avoidant species such as P. siculus, plasma glucose would accumulate with cold acclimation and would increase in response to acute exposure to subzero temperatures. To investigate this, we tested whether plasma glucose concentration and osmolality would increase in response to a subzero cold challenge before and after cold acclimation. In addition, we examined the relationship between metabolic rate, cold acclimation, and glucose by measuring metabolic rate during the cold challenge trials. We found that plasma glucose increased during the cold challenge trials, and that the increase was more pronounced after cold acclimation. However, baseline plasma glucose decreased throughout cold acclimation. Interestingly, total plasma osmolality did not change, and the increase in glucose only slightly altered freezing point depression. Metabolic rate during the cold challenge decreased after cold acclimation, and changes in respiratory exchange ratio suggest an increased relative use of carbohydrates. Overall, our findings demonstrate an important role for glucose in the response of P. siculus to an acute cold challenge, thus adding evidence for glucose as an important molecule for overwintering ectotherms that use freeze avoidant strategies.

Environmental Enrichment Differentially Activates Neural Circuits in FVB/N Mice, Inducing Social Interaction in Females but Agonistic Behavior in Males.

Environmental enrichment induces behavioral and structural modifications in rodents and influences the capability of mice to cope with stress. However, little is understood about hippocampal neurogenesis and the appearance of social/agonistic (aggressive) behavior upon activation of different neuronal circuits in FVB/N mice. Thus, in this study we hypothesized that environmental enrichment differentially regulates neurogenesis, neural circuit activation and social/agonistic behavior in male and female FVB/N mice. We explored the (1) neurogenic process as an indicative of neuroplasticity, (2) neuronal activation in the limbic system, and (3) social behavior using the resident-intruder test. On postnatal day 23 (PD23), mice were assigned to one of two groups: Standard Housing or Environmental Enrichment. At PD53, rodents underwent the resident-intruder test to evaluate social behaviors. Results revealed that environmental enrichment increased neurogenesis and social interaction in females. In males, environmental enrichment increased neurogenesis and agonistic behavior. Enriched male mice expressed higher levels of agonistic-related behavior than female mice housed under the same conditions. Neural circuit analysis showed lower activation in the amygdala of enriched males and higher activation in enriched females than their respective controls. Enriched females also showed higher activation in the frontal cortex without differences in male groups. Moreover, the insular cortex was less activated in females than in males. Thus, our results indicate that environmental enrichment has different effects on neuroplasticity and social/agonistic behavior in FVB/N mice, suggesting the relevance of sexual dimorphism in response to environmental stimuli.

Heat hardening in a pair of Anolis lizards: constraints, dynamics and ecological consequences.

Heat tolerance plasticity is predicted to be an important buffer against global warming. Nonetheless, basal heat tolerance often correlates negatively with tolerance plasticity ('trade-off hypothesis'), a constraint that could limit plasticity benefits. We tested the trade-off hypothesis at the individual level with respect to heat hardening in two lizard species, Anolis carolinensis and Anolis sagrei. Heat hardening is a rapid increase in heat tolerance after heat shock that is rarely measured in reptiles but is generally considered to be a first line of physiological defense against heat. We also employed a biophysical model of operative habitat temperatures to estimate the performance consequences of hardening under ecologically relevant conditions. Anolis carolinensis hardened by 2 h post-heat shock and maintained hardening for several hours. However, A. sagrei did not harden. Biophysical models showed that hardening in A. carolinensis reduces their overheating risk in the field. Therefore, while not all lizards heat harden, hardening has benefits for species that can. We initially found a negative relationship between basal tolerance and hardening within both species, consistent with the trade-off hypothesis. However, permutation analyses showed that the apparent trade-offs could not be differentiated from statistical artifact. We found the same result when we re-analyzed published data supporting the trade-off hypothesis in another lizard species. Our results show that false positives may be common when testing the trade-off hypothesis. Statistical approaches that account for this are critical to ensure that the hypothesis, which has broad implications for thermal adaptation and responses to warming, is assessed appropriately.

Heat hardening in a pair of Anolis lizards: constraints, dynamics and ecological consequences.

Heat tolerance plasticity is predicted to be an important buffer against global warming. Nonetheless, basal heat tolerance often correlates negatively with tolerance plasticity ('trade-off hypothesis'), a constraint that could limit plasticity benefits. We tested the trade-off hypothesis at the individual level with respect to heat hardening in two lizard species, Anolis carolinensis and Anolis sagrei Heat hardening is a rapid increase in heat tolerance after heat shock that is rarely measured in reptiles but is generally considered to be a first line of physiological defense against heat. We also employed a biophysical model of operative habitat temperatures to estimate the performance consequences of hardening under ecologically relevant conditions. Anolis carolinensis hardened by 2 h post-heat shock and maintained hardening for several hours. However, A. sagrei did not harden. Biophysical models showed that hardening in A. carolinensis reduces their overheating risk in the field. Therefore, while not all lizards heat harden, hardening has benefits for species that can. We initially found a negative relationship between basal tolerance and hardening within both species, consistent with the trade-off hypothesis. However, permutation analyses showed that the apparent trade-offs could not be differentiated from statistical artifact. We found the same result when we re-analyzed published data supporting the trade-off hypothesis in another lizard species. Our results show that false positives may be common when testing the trade-off hypothesis. Statistical approaches that account for this are critical to ensure that the hypothesis, which has broad implications for thermal adaptation and responses to warming, is assessed appropriately.

A Nonopioid, Nonbenzodiazepine Treatment Approach for Intractable Nausea and Vomiting in the Emergency Department.

GOAL: We sought to assess the feasibility and efficacy of a treatment protocol for nausea and vomiting using the combination of chlorpromazine, a dopamine antagonist antiemetic, and ketamine, a nonopioid analgesic. BACKGROUND: Increasing numbers of patients with cannabis use disorder are presenting to emergency departments with a poorly understood syndrome characterized by intractable nausea and vomiting. METHODS: This is a prospective, observational study involving a convenience sample of patients with unexplained nausea and vomiting. Subjects were given ketamine 15 mg slow intravenous push and chlorpromazine 12.5 mg intravenous over 15 minutes. Outcomes were number of episodes of emesis after study drug administration; change in nausea severity; change in pain severity; adverse events; and patient satisfaction. RESULTS: We enrolled 28 subjects on 30 emergency department visits. Twenty-three subjects (82%) reported at least weekly cannabis use with 19 reporting daily use. Initial symptoms were severe, with median pain and nausea scores both 10. After receiving study medication, the mean decrease in pain score over 120 minutes was 4.1 (95% confidence interval: 3.2, 5.0) and the mean decrease in nausea score was 4.9 (95% confidence interval: 4.0, 5.8). There were no adverse events. All 28 subjects who were asked reported they would want to receive these medications again. CONCLUSION: In this single-center study, the majority of patients presenting with intractable nausea and vomiting reported heavy cannabis use, and symptoms were severe. The combination of chlorpromazine plus ketamine resulted in rapid, definitive cessation of symptoms in most of these patients without the need for opioids or benzodiazepines.

Relationship between skin and urine colonization and surgical site infection in the proximal femur fracture: a prospective study.

PURPOSE: Antibiotic prophylaxis is routinely used in the surgical management of proximal femur fractures. The role of bacterial colonization of the skin and urine in the development of deep surgical site infections (SSI) is yet to be elucidated. This study aimed to evaluate the role of previous skin and urine colonization in the development of deep SSI after a proximal femoral fracture surgery. METHODS: We conducted a prospective observational study in 326 patients > 64 years old, who were scheduled to surgery. Cultures from skin samples of the surgical site and from urine were performed prior to the procedure, and cefazoline was administered as prophylaxis. RESULTS: Skin microbiota was isolated in 233 (71.5%) cases; 8 (2.5%) samples were positive for other bacteria, and 85 (26%) were negative. Of 236 urine samples, 168 were negative or contaminated (71.2%), and 68 (28.8%) were positive, being 58/236 for Enterobacterales (24.6%). Acute deep SSI were diagnosed in nine out of 326 patients (2.7%), and two (22%) were infected by Gram-negative bacilli. Of the 9 cases, normal skin microbiota was isolated in 7 (78%), and the remaining two were negative. Seven cases had negative or contaminated urine cultures, and the one with E. coli did not correlate with SSI bacteria. CONCLUSION: In our elderly hip fracture population, most patients harbored normal skin microbiota, and Enterobacterales urine cultures were positive in one-quarter of cases. There was no relationship between skin colonization, urine culture, and deep SSI. We therefore do not believe that our patients would benefit from modifying the current antibiotic prophylaxis.

Thermal tolerance varies with age and sex for the nonnative Italian Wall Lizard (Podarcis siculus) in Southern California.

Temperature has a substantial effect on both the physiology and behavior of ectothermic animals such as lizards. Physiology and behavior can also be influenced by ontogenetic and sex differences, but these effects are largely understudied in lizards. We examined ontogenetic and sex-based differences in thermal tolerances, preferred temperature, and temperature-dependent evaporative water loss rates in Italian Wall Lizards, Podarcis siculus, collected from an introduced population near Los Angeles, California, USA that were acclimated to laboratory conditions. Podarcis siculus has been introduced to multiple localities in the USA and the Mediterranean region and has demonstrated remarkable ability to adapt to novel climatic conditions. In the California population, adults of both sexes had a higher critical thermal maximum (CTmax) than juveniles, and adult females had a lower critical thermal minimum (CTmin) than juveniles and adult males. Thus, adult females had a significantly wider thermal breadth (CTmax - CTmin) compared to adult males and juveniles. Mass-specific evaporative water loss was higher in juveniles compared to adult males at intermediate temperatures. There was no significant difference among groups for preferred temperature. This implies that thermal tolerance, a physiological characteristic, varies with age and sex for this population, whereas thermal preference, a behavioral characteristic, does not. Interestingly, CTmin for all age and sex classes was above temperatures likely experienced by some nonnative populations in winter, suggesting individuals need to find urban thermal retreats. These results add to the growing literature demonstrating that thermal tolerances and breadths can vary between sexes and across age classes in squamate species.

GABAρ subunits confer a bicuculline-insensitive component to GFAP+ cells of cerebellum.

GABA-A receptors mediating synaptic or extrasynaptic transmission are molecularly and functionally distinct, and glial cells are known to express a plethora of GABA-A subunits. Here we demonstrate that GFAP(+) cells of the granular layer of cerebellum express GABAρ subunits during early postnatal development, thereby conferring peculiar pharmacologic characteristics to GABA responses. Electron microscopy revealed the presence of GABAρ in the plasma membrane of GFAP(+) cells. In contrast, expression in the adult was restricted to Purkinje neurons and a subset of ependymal cells. Electrophysiological studies in vitro revealed that astrocytes express functional receptors with an EC50 of 52.2 ± 11.8 µM for GABA. The evoked currents were inhibited by bicuculline (100 µM) and TPMPA (IC50, 5.9 ± 0.6 µM), indicating the presence of a GABAρ component. Coimmunoprecipitation demonstrated protein-protein interactions between GABAρ1 and GABAα1, and double immunofluorescence showed that these subunits colocalize in the plasma membrane. Three populations of GABA-A receptors in astrocytes were identified: classic GABA-A, bicuculline-insensitive GABAρ, and GABA-A-GABAρ hybrids. Clusters of GABA-A receptors were distributed in the perinuclear space and along the processes of GFAP(+) cells. Time-lapse microscopy showed GABAρ2-GFP accumulation in clusters located in the soma and along the processes. The clusters were relatively immobile, with mean displacement of 9.4 ± 0.9 µm and a net distance traveled of 1-2 µm, owing mainly to directional movement or simple diffusion. Modulation of GABAρ dynamics may be a novel mechanism of extrasynaptic transmission regulating GABAergic control of GFAP(+) cells during early postnatal development.

Anorexia Reduces GFAP+ Cell Density in the Rat Hippocampus.

Anorexia nervosa is an eating disorder observed primarily in young women. The neurobiology of the disorder is unknown but recently magnetic resonance imaging showed a volume reduction of the hippocampus in anorexic patients. Dehydration-induced anorexia (DIA) is a murine model that mimics core features of this disorder, including severe weight loss due to voluntary reduction in food intake. The energy supply to the brain is mediated by astrocytes, but whether their density is compromised by anorexia is unknown. Thus, the aim of this study was to estimate GFAP+ cell density in the main regions of the hippocampus (CA1, CA2, CA3, and dentate gyrus) in the DIA model. Our results showed that GFAP+ cell density was significantly reduced (~20%) in all regions of the hippocampus, except in CA1. Interestingly, DIA significantly reduced the GFAP+ cells/nuclei ratio in CA2 (-23%) and dentate gyrus (-48%). The reduction of GFAP+ cell density was in agreement with a lower expression of GFAP protein. Additionally, anorexia increased the expression of the intermediate filaments vimentin and nestin. Accordingly, anorexia increased the number of reactive astrocytes in CA2 and dentate gyrus more than twofold. We conclude that anorexia reduces the hippocampal GFAP+ cell density and increases vimentin and nestin expression.

Dehydration-Induced Anorexia Reduces Astrocyte Density in the Rat Corpus Callosum.

Anorexia nervosa is an eating disorder associated with severe weight loss as a consequence of voluntary food intake avoidance. Animal models such as dehydration-induced anorexia (DIA) mimic core features of the disorder, including voluntary reduction in food intake, which compromises the supply of energy to the brain. Glial cells, the major population of nerve cells in the central nervous system, play a crucial role in supplying energy to the neurons. The corpus callosum (CC) is the largest white matter tract in mammals, and more than 99% of the cell somata correspond to glial cells in rodents. Whether glial cell density is altered in anorexia is unknown. Thus, the aim of this study was to estimate glial cell density in the three main regions of the CC (genu, body, and splenium) in a murine model of DIA. The astrocyte density was significantly reduced (~34%) for the DIA group in the body of the CC, whereas in the genu and the splenium no significant changes were observed. DIA and forced food restriction (FFR) also reduced the ratio of astrocytes to glial cells by 57.5% and 22%, respectively, in the body of CC. Thus, we conclude that DIA reduces astrocyte density only in the body of the rat CC.

Modulation of GABA-A receptors of astrocytes and STC-1 cells by taurine structural analogs.

Taurine activates and modulates GABA receptors in vivo as well as those expressed in heterologous systems. This study aimed to determine whether the structural analogs of taurine: homotaurine and hypotaurine, have the ability to activate GABA-A receptors that include GABAρ subunits. The expression of GABA-A receptors containing GABAρ has been reported in the STC-1 cells and astrocytes. In both cell types, taurine, homo-, and hypotaurine gated with low efficiency a picrotoxin-sensitive GABA-A receptor. The known bimodal modulatory effect of taurine on GABAρ receptors was not observed; however, differences between the activation and deactivation rates were detected when they were perfused together with GABA. In silico docking simulations suggested that taurine, hypo-, and homotaurine do not form a cation-π interaction such as that generated by GABA in the agonist-binding site of GABAρ. This observation complements the electrophysiological data suggesting that taurine and its analogs act as partial agonists of GABA-A receptors. All the observations above suggest that the structural analogs of taurine are partial agonists of GABA-A receptors that occupy the agonist-binding site, but their structures do not allow the proper interaction with the receptor to fully gate its Cl(-) channel.

γ-Aminobutyric acid-ρ expression in ependymal glial cells of the mouse cerebellum.

The ependymal glial cells (EGCs) from the periventricular zone of the cerebellum were studied to determine their distribution and the functional properties of their γ-aminobutyric acid type A (GABA(A) ) receptors. EGCs were identified by the presence of ciliated structures on their ventricular surface and their expression of glial fibrillary acidic protein (GFAP). Interestingly, diverse cell types, including neurons, astrocytes, and other types of glia, were identified in the subventricular zone by their current profiles. Electron microscopy showed ciliated cells and myelinated axons in this zone, but we found no collateral connections to suggest the presence of functional synapses. GABA-mediated currents were recorded from EGCs in cerebellar slices from postnatal days 13 to 35 (PN13-PN35). These currents were blocked by TPMPA (a highly specific GABA(A) ρ subunit antagonist) and bicuculline (a selective antagonist for classic GABA(A) receptors). Pentobarbital failed to modulate GABA(A)-mediated currents despite the expression of GABAα1 and GABAγ2 subunits. In situ hybridization, RT-PCR, and immunofluorescence studies confirmed GABAρ1 expression in EGCs of the cerebellum. We conclude that cerebellar EGCs express GABAρ1, which is functionally involved in GABA(A) receptor-mediated responses that are unique among glial cells of the brain.

Regional density of glial cells in the rat corpus callosum.

Axons and glial cells are the main components of white matter. The corpus callosum (CC) is the largest white matter tract in mammals; in rodents, 99% of the cells correspond to glia after postnatal day 5 (P5). The area of the CC varies through life and regional differences related to the number of axons have been previously described. Whether glial cell density varies accordingly is unknown; thus the aim of this study was to estimate glial cell density for the genu, body and splenium -the three main regions of CC-, of P6 and P30 rats. Here we report that the density of CC glial cells reduced by ~10% from P6 to P30. Even so, the density of astrocytes showed a slight increase (+6%), probably due to differentiation of glioblasts. Interestingly, glial cell density decreased for the genu (-21%) and the body (-13%), while for the splenium a minor increase (+5%) was observed. The astrocyte/glia ratio increased (from P6 to P30) for the genu (+27%), body (+17%) and splenium (+4%). Together, our results showed regional differences in glial cell density of the CC. Whether this pattern is modified in some neuropathologies remains to be explored.

Rapid Physiological Plasticity in Response to Cold Acclimation for Nonnative Italian Wall Lizards (Podarcis siculus) from New York.

AbstractThermal physiology helps us understand how ectotherms respond to novel environments and how they persist when introduced to new locations. Researchers generally measure thermal physiology traits immediately after animal collection or after a short acclimation period. Because many of these traits are plastic, the conclusions drawn from such research can vary depending on the duration of the acclimation period. In this study, we measured the rate of change and extent to which cold tolerance (critical thermal minimum [CTmin]) of nonnative Italian wall lizards (Podarcis siculus) from Hempstead, New York, changed during a cold acclimation treatment. We also examined how cold acclimation affected heat tolerance (critical thermal maximum [CTmax]), thermal preference (Tpref), evaporative water loss (EWL), resting metabolic rate (RMR), and respiratory exchange ratio (RER). We predicted that CTmin, CTmax, and Tpref would decrease with cold acclimation but that EWL and RMR would increase with cold acclimation. We found that CTmin decreased within 2 wk and that it remained low during the cold acclimation treatment; we suspect that this cold tolerance plasticity reduces risk of exposure to lethal temperatures during winter for lizards that have not yet found suitable refugia. CTmax and Tpref also decreased after cold acclimation, while EWL, RMR, and RER increased after cold acclimation, suggesting trade-offs with cold acclimation in the form of decreased heat tolerance and increased energy demands. Taken together, our findings suggest that cold tolerance plasticity aids the persistence of an established population of invasive lizards. More generally, our findings highlight the importance of accounting for the plasticity of physiological traits when investigating how invasive species respond to novel environments.

Striatal synaptic changes and behavior in adult mouse upon prenatal exposure to valproic acid.

Autism spectrum disorders (ASD) are a group of neurodevelopmental disorders characterized by persistent deficits in social communication and social interaction. Altered synaptogenesis and aberrant connectivity responsible for social behavior and communication have been reported in autism pathogenesis. Autism has a strong genetic and heritable component; however, environmental factors including toxins, pesticides, infection and in utero exposure to drugs such as VPA have also been implicated in ASD. Administration of VPA during pregnancy has been used as a rodent model to study pathophysiological mechanisms involved in ASD, and in this study, we used the mouse model of prenatal exposure to VPA to assess the effects on striatal and dorsal hippocampus function in adult mice. Alterations in repetitive behaviors and shift habits were observed in mice prenatally exposed to VPA. In particular, such mice presented a better performance in learned motor skills and cognitive deficits in Y-maze learning frequently associated with striatal and hippocampal function. These behavioral changes were associated with a decreased level of proteins involved in the formation and maintenance of excitatory synapses, such as Nlgn-1 and PSD-95. In conclusion, motor skill abilities, repetitive behaviors, and impaired flexibility to shift habits are associated with reduced striatal excitatory synaptic function in the adult mouse prenatally exposed to VPA.

Expression of GABAρ receptors in the neostriatum: localization in aspiny, medium spiny neurons and GFAP-positive cells.

GABAergic transmission in the neostriatum plays a central role in motor coordination, in which a plethora of GABA-A receptor subunits combine to modulate neural inhibition. GABAρ receptors were originally described in the mammalian retina. These receptors possess special electrophysiological and pharmacological properties, forming a characteristic class of ionotropic receptors. In previous studies, we suggested that GABAρ receptors are expressed in the neostriatum, and in this report we show that they are indeed present in all the calretinin-positive interneurons of the neostriatum. In addition, they are located in calbindin-positive interneurons and projection neurons that express the dopamine D(2) receptor. GABAρ receptors were also located in 30% of the glial fibrillary acidic protein-positive cells, and may therefore also contribute to gliotransmission. Quantitative reverse transcription-PCR suggested that the mRNAs of this receptor do not express as much as in the retina, and that GABAρ2 is more abundant than GABAρ1. Electrophysiological recordings in brain slices provided evidence of neurons expressing a cis-4-aminocrotonic acid-activated, 1,2,5,6-tetrahydropyridine-4-yl methylphosphinic acid-sensitive ionotropic GABA receptor, indicating the presence of functional GABAρ receptors in the neostriatum. Finally, electron-microscopy and immunogold located the receptors mainly in perisynaptic as well as in extrasynaptic sites. All these observations reinforce the importance of GABAρ receptors in the neostriatum and contribute to the diversity of inhibitory regulation in this area.

Glial cells in anorexia.

Anorexia is a loss of appetite or an inability to eat and is often associated with eating disorders. However, animal anorexia is physiologically regulated as a part of the life cycle; for instance, during hibernation, migration or incubation. Anorexia nervosa (AN), on the other hand, is a common eating disorder among adolescent females that experience an intense fear of gaining weight due to body image distortion that results in voluntary avoidance of food intake and, thus, severe weight loss. It has been shown that the neurobiology of feeding extends beyond the hypothalamus. The prefrontal cortex (PFC) is involved in food choice and body image perception, both relevant in AN. However, little is known about the neurobiology of AN, and the lack of effective treatments justifies the use of animal models. Glial cells, the dominant population of nerve cells in the central nervous system, are key in maintaining brain homeostasis. Accordingly, recent studies suggest that glial function may be compromised by anorexia. In this review, we summarize recent findings about anorexia and glial cells.

Anorexia disrupts glutamate-glutamine homeostasis associated with astroglia in the prefrontal cortex of young female rats.

Anorexia nervosa (AN) is an eating disorder characterized by self-starvation and excessive weight loss with a notorious prevalence in young women. The neurobiology of AN is unknown but murine models, like dehydration induced anorexia (DIA), reproduce weight loss and avoidance of food despite its availability. Astrocytes are known to provide homeostatic support to neurons, but it is little explored if anorexia affects this function. In this study, we tested if DIA disrupts glutamate-glutamine homeostasis associated with astrocytes in the prefrontal cortex (PFC) of young female rats. Our results showed that anorexia reduced the redox state, as well as endogenous glutamate and glutamine. These effects correlated with a reduced expression of the glutamate transporters (GLT-1 and GLAST) and glutamine synthetase, all of them are preferentially expressed by astrocytes. Accordingly, the expression of GFAP was reduced. Anorexia reduced the astrocyte density, promoted a de-ramified morphology, and augmented the de-ramified/ramified astrocyte ratio in the medial prefrontal cortex (mPFC) and orbitofrontal cortex (OFC), but not in the motor cortex (M2). The increase of a de-ramified phenotype correlated with increased expression of vimentin and nestin. Based on these results, we conclude that anorexia disrupts glutamate-glutamine homeostasis and the redox state associated with astrocyte dysfunction.

Transcriptome-wide Cas13 guide RNA design for model organisms and viral RNA pathogens.

The recent characterization of RNA-targeting CRISPR nucleases has enabled diverse transcriptome engineering and screening applications that depend crucially on prediction and selection of optimized CRISPR guide RNAs (gRNAs). Previously, we developed a computational model to predict RfxCas13d gRNA activity for all human protein-coding genes. Here, we extend this framework to six model organisms (human, mouse, zebrafish, fly, nematode, and flowering plants) for protein-coding genes and noncoding RNAs (ncRNAs) and also to four RNA virus families (severe acute respiratory syndrome coronavirus 2 [SARS-CoV-2], HIV-1, H1N1 influenza, and Middle East respiratory syndrome [MERS]). We include experimental validation of predictions by testing knockdown of multiple ncRNAs (MALAT1, HOTAIRM1, Gas5, and Pvt1) in human and mouse cells. We developed a freely available web-based platform (cas13design) with pre-scored gRNAs for transcriptome-wide targeting in several organisms and an interactive design tool to predict optimal gRNAs for custom RNA targets entered by the user. This resource will facilitate CRISPR-Cas13 RNA targeting in model organisms, emerging viral threats to human health.