Potentiation of the M 1 muscarinic acetylcholine receptor normalizes neuronal activation patterns and improves apnea severity in Mecp2+/- mice.

Rett syndrome (RTT) is a neurodevelopmental disorder that is caused by loss-of-function mutations in the methyl-CpG binding protein 2 ( MeCP2 ) gene. RTT patients experience a myriad of debilitating symptoms, which include respiratory phenotypes that are often associated with lethality. Our previous work established that expression of the M 1 muscarinic acetylcholine receptor (mAchR) is decreased in RTT autopsy samples, and that potentiation of the M 1 receptor improves apneas in a mouse model of RTT; however, the population of neurons driving this rescue is unclear. Loss of Mecp2 correlates with excessive neuronal activity in cardiorespiratory nuclei. Since M 1 is found on cholinergic interneurons, we hypothesized that M 1 -potentiating compounds decrease apnea frequency by tempering brainstem hyperactivity. To test this, Mecp2 +/- and Mecp2 +/+ mice were screened for apneas before and after administration of the M 1 positive allosteric modulator (PAM) VU0453595 (VU595). Brains from the same mice were then imaged for c-Fos, ChAT, and Syto16 using whole-brain light-sheet microscopy to establish genotype and drug-dependent activation patterns that could be correlated with VU595's efficacy on apneas. The vehicle-treated Mecp2 +/- brain exhibited broad hyperactivity when coupled with the phenotypic prescreen, which was significantly decreased by administration of VU595, particularly in regions known to modulate the activity of respiratory nuclei (i.e. hippocampus and striatum). Further, the extent of apnea rescue in each mouse showed a significant positive correlation with c-Fos expression in non-cholinergic neurons in the striatum, thalamus, dentate gyrus, and within the cholinergic neurons of the brainstem. These results indicate that Mecp2 +/- mice are prone to hyperactivity in brain regions that regulate respiration, which can be normalized through M 1 potentiation.

Differences in the learning curve of robotic transabdominal preperitoneal inguinal hernia repair according to surgeon's robotic experience.

PURPOSE: In this study, we aim to analyze the learning curve of each step of robotic transabdominal pre-peritoneal inguinal hernia repair (rTAPP) in two surgeons with varying degrees of expertise with the robotic platform but no experience with laparoscopic hernia repair. METHODS: Data on 124 rTAPP cases performed by two surgeons were retrospectively reviewed. Cumulative sum (CUSUM) analysis was applied to visualize the learning curve of rTAPP on operation time of each step of the procedure [the peritoneal flap creation (T1), the completion of the critical view of the myopectineal orifice (T2), the mesh application (T3) and the peritoneal flap closure (T4)]. Each intraoperative and postoperative outcome was compared according to surgeon's experience with the robotic platform and learning phase. The robotic surgeon mentored the surgeon-in-training and was present during all surgeries in his learning period. RESULTS: The surgeon in training with the robotic platform showed a learning phase till the 20th procedure followed by a gradual improvement in performances. The expert surgeon showed a learning phase till the 35th procedure after which a constant decrease of operative time was recorded till the last procedure included. The operative times of each step of the procedures of both surgeons were significantly improved after the learning phase. In the late phase, the surgeon in training could achieve operative times in T2 and T3, which are similar to those of an experienced robotic surgeon with no experience with TAPP before the completion of the learning phase. CONCLUSIONS: In conclusion, the learning phase of rTAPP surgery may vary between 20 and 35 cases, depending on the surgeon's experience in robotic surgery.

Growth fitness, heme uptake and genomic variants in mutants of oxygen-tolerant Lacticaseibacillus casei and Lactiplantibacillus plantarum strains.

Adaptive Laboratory Evolution (ALE) is a powerful tool to improve the fitness of industrially relevant microorganisms, because it circumvents some of the problems related to the use of genetically modified strains. In this study, we used an ALE strategy involving serial batch cultivations in aerobic and respiratory conditions to generate spontaneous mutants from the respiration-competent strain Lacticaseibacillus casei N87. Genotypic changes in selected mutants were investigated using whole genome sequencing (WGS). The O2-tolerant Lactiplantibacillus plantarum C17 and its mutant C17-m58 (obtained from a previous ALE study) were included in heme uptake experiments and in WGS and variant calling analyses. Several Lcb. casei N87 mutants cultivated under aerobic and respiratory conditions showed improved biomass production, O2 uptake and oxidative stress tolerance compared to the parental strain. Mutants of Lcb. casei and Lpb. plantarum differed from the parental strains in the ability to use heme and menaquinone. High heme concentrations (> 10 mg/L), however, were toxic for all strains. Single nucleotide modifications (SNPs) were detected in some genes encoding for proteins and transcriptional regulators involved in carbon metabolism, oxidative stress, redox balance, and cell wall properties, but their role in the evolved phenotypes needs further investigations. We conclude that prolonged adaptation to aerobic and respiratory life-style may be used as natural strategy to generate strains with improved O2-consuming ability and oxidative stress tolerance, two important features to develop robust cultures and to reduce oxidative processes in foods.

Clinical and Preclinical Evidence for M1 Muscarinic Acetylcholine Receptor Potentiation as a Therapeutic Approach for Rett Syndrome.

Rett syndrome (RTT) is a neurodevelopmental disorder that is characterized by developmental regression, loss of communicative ability, stereotyped hand wringing, cognitive impairment, and central apneas, among many other symptoms. RTT is caused by loss-of-function mutations in a methyl-reader known as methyl-CpG-binding protein 2 (MeCP2), a protein that links epigenetic changes on DNA to larger chromatin structure. Historically, target identification for RTT has relied heavily on Mecp2 knockout mice; however, we recently adopted the alternative approach of performing transcriptional profiling in autopsy samples from RTT patients. Through this mechanism, we identified muscarinic acetylcholine receptors (mAChRs) as potential therapeutic targets. Here, we characterized a cohort of 40 temporal cortex samples from individuals with RTT and quantified significantly decreased levels of the M1, M2, M3, and M5 mAChRs subtypes relative to neurotypical controls. Of these four subtypes, M1 expression demonstrated a linear relationship with MeCP2 expression, such that M1 levels were only diminished in contexts where MeCP2 was also significantly decreased. Further, we show that M1 potentiation with the positive allosteric modulator (PAM) VU0453595 (VU595) rescued social preference, spatial memory, and associative memory deficits, as well as decreased apneas in Mecp2+/- mice. VU595's efficacy on apneas in Mecp2+/- mice was mediated by the facilitation of the transition from inspiration to expiration. Molecular analysis correlated rescue with normalized global gene expression patterns in the brainstem and hippocampus, as well as increased Gsk3ß inhibition and NMDA receptor trafficking. Together, these data suggest that M1 PAMs could represent a new class of RTT therapeutics.

Optimized Administration of the M4 PAM VU0467154 Demonstrates Broad Efficacy, but Limited Effective Concentrations in Mecp2+/- Mice.

Hypofunction of cholinergic circuits and diminished cholinergic tone have been associated with the neurodevelopmental disorder Rett syndrome (RTT). Specifically, deletion of Mecp2 in cholinergic neurons evokes the same social and cognitive phenotypes in mice seen with global Mecp2 knockout, and decreased choline acetyltransferase activity and vesamicol binding have been reported in RTT autopsy samples. Further, we recently identified significant decreases in muscarinic acetylcholine receptor subtype 4 (M4) expression in both the motor cortex and cerebellum of RTT patient autopsies and established proof of concept that an acute dose of the positive allosteric modulator (PAM) VU0467154 (VU154) rescued phenotypes in Mecp2+/- mice. Here, we expand the assessment of M4 PAMs in RTT to address clinically relevant questions of tolerance, scope of benefit, dose response, chronic treatment, and mechanism. We show that VU154 has efficacy on anxiety, social preference, cognitive, and respiratory phenotypes in Mecp2+/- mice; however, the therapeutic range is narrow, with benefits seen at 3 mg/kg concentrations, but not 1 or 10 mg/kg. Further, sociability was diminished in VU154-treated Mecp2+/- mice, suggestive of a potential adverse effect. Compound efficacy on social, cognitive, and respiratory phenotypes was conserved with a 44-day treatment paradigm, with the caveat that breath rate was moderately decreased with chronic treatment in Mecp2+/+ and Mecp2+/- mice. VU154 effects on respiratory function correlated with an increase in Gsk3ß inhibition in the brainstem. These results identify the core symptom domains where efficacy and adverse effects may present with M4 administration in RTT model mice and advocate for the continued evaluation as potential RTT therapeutics.

Exploration of group II metabotropic glutamate receptor modulation in mouse models of Rett syndrome and MECP2 Duplication syndrome.

Rett syndrome (RTT) and MECP2 Duplication syndrome (MDS) have opposing molecular origins in relation to expression and function of the transcriptional regulator Methyl-CpG-binding protein 2 (MeCP2). Several clinical and preclinical phenotypes, however, are shared between these disorders. Modulation of MeCP2 levels has recently emerged as a potential treatment option for both of these diseases. However, toxicity concerns remain with these approaches. Here, we focus on pharmacologically modulating the group II metabotropic glutamate receptors (mGlu), mGlu2 and mGlu3, which are two downstream targets of MeCP2 that are bidirectionally affected in expression in RTT patients and mice (Mecp2Null/+) versus an MDS mouse model (MECP2Tg1/o). Mecp2Null/+ and MECP2Tg1/o animals also exhibit contrasting phenotypes in trace fear acquisition, a form of temporal associative learning and memory, with trace fear deficiency observed in Mecp2Null/+ mice and abnormally enhanced trace fear acquisition in MECP2Tg1/o animals. In Mecp2Null/+ mice, treatment with the mGlu2/3 agonist LY379268 reverses the deficit in trace fear acquisition, and mGlu2/3 antagonism with LY341495 normalizes the abnormal trace fear learning and memory phenotype in MECP2Tg1/o mice. Altogether, these data highlight the role of group II mGlu receptors in RTT and MDS and demonstrate that both mGlu2 and mGlu3 may be potential therapeutic targets for these disorders.

Profiling beneficial and potential adverse effects of MeCP2 overexpression in a hypomorphic Rett syndrome mouse model.

De novo loss-of-function mutations in methyl-CpG-binding protein 2 (MeCP2) lead to the neurodevelopmental disorder Rett syndrome (RTT). Despite promising results from strategies aimed at increasing MeCP2 levels, additional studies exploring how hypomorphic MeCP2 mutations impact the therapeutic window are needed. Here, we investigated the consequences of genetically introducing a wild-type MECP2 transgene in the Mecp2 R133C mouse model of RTT. The MECP2 transgene reversed the majority of RTT-like phenotypes exhibited by male and female Mecp2 R133C mice. However, three core symptom domains were adversely affected in female Mecp2R133C/+ animals; these phenotypes resemble those observed in disease contexts of excess MeCP2. Parallel control experiments in Mecp2Null/+ mice linked these adverse effects to the hypomorphic R133C mutation. Collectively, these data provide evidence regarding the safety and efficacy of genetically overexpressing functional MeCP2 in Mecp2 R133C mice and suggest that personalized approaches may warrant consideration for the clinical assessment of MeCP2-targeted therapies.

A Chiral fNIRS Spotlight on Cerebellar Activation in a Finger Tapping Task.

Functional Magnetic Resonance Imaging (fMRI) has been so far the golden standard to study the functional aspects of the cerebellum. In this paper, a low-cost alternative imaging, i.e. functional Near-Infrared Spectroscopy (fNIRS) is demonstrated to achieve successful measurements of the cerebellar hemodynamics towards the challenging observation of motor and cognitive processes at the cerebellar level. The excitation and reception optodes need to be properly placed to circumvent a major hindering from the shielding by the neck muscles. A simple experimental protocol, i.e. finger tapping task, was implemented to observe the subject's engagement and the presence of functional asymmetries. Marked differences among subjects with different levels of lateralization were clearly noticed in terms of activation and latencies, together with peaks in the hemodynamic response following neural activation. These preliminary results suggest also differences in the hemodynamic behavior between the brain and the cerebellum and encourage future and extended analysis in this direction.Clinical Relevance-This establishes the possibility to use a novel technique (fNIRS) to study cerebellar hemodynamics instead of fMRI.

Single-Trial Detection of Event-Related Potentials with Integral Shape Averaging: An Application to the Elusive N400.

The estimation of Event-Related Potentials (ERPs) from the ambient EEG is a difficult task, usually achieved through the synchronous averaging of an extensive series of trials. However, this technique has some caveats: the ERPs have to be strictly time-locked with similar shape, i.e. emitted with the same latency and the same profile, with minor fluctuations of their amplitudes. Also, the method requires a huge number of valid trials (~100) to efficiently raise the ERPs from the EEG trials. In the case of cognitive ERPs, as with the N400, the delivered stimulus has to be different for each trial, the latencies are varying, and the number of available trials is usually low. In this paper, an alternative method, coined Integral Shape Averaging (ISA) and its derivatives are detailed. ISA is robust to varying latencies and affine transforms of shape. Furthermore, a new method coined ISAD can be derived to extract ERPs even from a single trial experiment. The aim here is to illustrate the potential of ISAD for N400 component extraction on real EEG data, with emphasis on its general applicability for ERPs computation and its major assets like reduced experimental protocol. Some insights are also given on its potential use to study ERP variability, through shape and latency.Clinical Relevance- The proposed algorithm aims to be a helpful tool in clinical practice to analyze and interpret evoked responses in real experimental settings, especially for particularities in neurology.

Comparison across 12 countries on knowledge, attitude, and behavior scores about medication errors in Intensive Care Units: an international study.

OBJECTIVE: The aim of the study was to explore the degree of agreement of intensive care unit nurses working on a set of medication error preventive strategies and to examine possible predictors of nurses' knowledge, attitude and behavior. MATERIALS AND METHODS: Observational, international, and cross-sectional study. Iran, Malta, Spain, Pakistan, Nepal, Qatar, Ecuador, Australia, Finland, Italy, Egypt, and Jordan were the countries included in this survey. To collect data, the Knowledge, Attitude and Behavior in Medication Errors questionnaire was used. A descriptive statistical analysis was performed for the socio-demographic characteristics of the sample and three multiple logistic regressions were performed. RESULTS: The international sample consists of 1383 nurses, of whom 478 (34.6%) were men and 900 (65.1%) were women and their mean age was 35.61 years with a range of 19-61. Descriptive statistics conducted on the international sample show a medium to high degree of agreement among participants concerning some preventive strategies of medication error. In addition, the results of the present study show a strong relationship between positive nurses' attitudes and correct behaviors and/or adequate knowledge, as well as between adequate knowledge and correct behaviors (p< 0.01). CONCLUSIONS: Further studies are needed to explore the issue of medication error concerning nurses' cultural backgrounds, as well as to assess similarities and disparities among international nurses.

Activating mGlu3 Metabotropic Glutamate Receptors Rescues Schizophrenia-like Cognitive Deficits Through Metaplastic Adaptations Within the Hippocampus.

BACKGROUND: Polymorphisms in GRM3, the gene encoding the mGlu3 metabotropic glutamate receptor, are associated with impaired cognition and neuropsychiatric disorders such as schizophrenia. Limited availability of selective genetic and molecular tools has hindered progress in developing a clear understanding of the mechanisms through which mGlu3 receptors regulate synaptic plasticity and cognition. METHODS: We examined associative learning in mice with trace fear conditioning, a hippocampal-dependent learning task disrupted in patients with schizophrenia. Underlying cellular mechanisms were assessed using ex vivo hippocampal slice preparations with selective pharmacological tools and selective genetic deletion of mGlu3 receptor expression in specific neuronal subpopulations. RESULTS: mGlu3 receptor activation enhanced trace fear conditioning and reversed deficits induced by subchronic phencyclidine. Mechanistic studies revealed that mGlu3 receptor activation induced metaplastic changes, biasing afferent stimulation to induce long-term potentiation through an mGlu5 receptor-dependent, endocannabinoid-mediated, disinhibitory mechanism. Selective genetic deletion of either mGlu3 or mGlu5 from hippocampal pyramidal cells eliminated effects of mGlu3 activation, revealing a novel mechanism by which mGlu3 and mGlu5 interact to enhance cognitive function. CONCLUSIONS: These data demonstrate that activation of mGlu3 receptors in hippocampal pyramidal cells enhances hippocampal-dependent cognition in control and impaired mice by inducing a novel form of metaplasticity to regulate circuit function, providing a clear mechanism through which genetic variation in GRM3 can contribute to cognitive deficits. Developing approaches to positively modulate mGlu3 receptor function represents an encouraging new avenue for treating cognitive disruption in schizophrenia and other psychiatric diseases.

A GRM7 mutation associated with developmental delay reduces mGlu7 expression and produces neurological phenotypes.

The metabotropic glutamate receptor 7 (mGlu7) is a G protein-coupled receptor that has been recently linked to neurodevelopmental disorders. This association is supported by the identification of GRM7 variants in patients with autism spectrum disorder, attention deficit hyperactivity disorder, and severe developmental delay. One GRM7 mutation previously reported in 2 patients results in a single amino acid change, I154T, within the mGlu7 ligand-binding domain. Here, we report 2 new patients with this mutation who present with severe developmental delay and epilepsy. Functional studies of the mGlu7-I154T mutant reveal that this substitution resulted in significant loss of mGlu7 protein expression in HEK293A cells and in mice. We show that this occurred posttranscriptionally at the level of protein expression and trafficking. Similar to mGlu7-global KO mice, mGlu7-I154T animals exhibited reduced motor coordination, deficits in contextual fear learning, and seizures. This provides functional evidence that a disease-associated mutation affecting the mGlu7 receptor was sufficient to cause neurological dysfunction in mice and further validates GRM7 as a disease-causing gene in the human population.

Socio-demographic analysis of free-lance nurses in a non-institutional employment regimen.

INTRODUCTION: The free-lance nurse, not bound to rigid organizational systems, can offer personalized assistance always respecting the rights of the person and of the profession. More recent graduates have decided to undertake the nursing profession by moving towards the free-lance nursing, considering it both as a career opening and as a professional opportunity, although this option never got much attention from the researchers in the Italian nursing scene. Free-lance nursing is now considered a valuable opportunity to develop a nursing career. This market is destined to grow for different reasons, such as an increasing chronicity of health conditions of more and more ageing population and the deficits of the National Health Service (Servizio Sanitario Nazionale - SSN) in community and home care. AIM: The aim of the study was to evaluate the correlation between the development of the free-lance nursing and the Italian socio-economic context. METHODS: The design of the study was descriptive - observational. Data collection and observation was carried out from January 2018 until April 2108. For the analysis a linear regression model was adopted to quantify a cause-effect relationship between one or more independent variables and the dependent variable which interprets the phenomenon investigated. The regression carried out was descriptive to analytically express the observed reality and represent it in a plausible way. The specification model was represented as: Free-lance nurses per capita = per capita income + Out of Pocket expense per capita + waiting lists in days + number of beds per inhabitants + NHS nurses per inhabitants. RESULTS: The estimate carried out had an R of 0.813, R-square equal to 0.6612, adjusted R-square 0.540 and standard error of the estimate 1.277, highlighting a correlation between the variables adopted in the model and a p = 0.005. From the analysis of the variables used, the average per capita income (p = 0.045) and the nurses working in the National Health Service /1,000 beds (p = 0.017) were statistically significant. CONCLUSIONS: It can be stated that the free-lance nursing profession is costly for patients and therefore develops more revenue where the average per capita income grows, but the research also seems to show that, where the National Health Service has too few nurses, the private demand increases in order to satisfy healthcare needs.

Phenotypic profiling of mGlu7 knockout mice reveals new implications for neurodevelopmental disorders.

Neurodevelopmental disorders are characterized by deficits in communication, cognition, attention, social behavior and/or motor control. Previous studies have pointed to the involvement of genes that regulate synaptic structure and function in the pathogenesis of these disorders. One such gene, GRM7, encodes the metabotropic glutamate receptor 7 (mGlu7 ), a G protein-coupled receptor that regulates presynaptic neurotransmitter release. Mutations and polymorphisms in GRM7 have been associated with neurodevelopmental disorders in clinical populations; however, limited preclinical studies have evaluated mGlu7 in the context of this specific disease class. Here, we show that the absence of mGlu7 in mice is sufficient to alter phenotypes within the domains of social behavior, associative learning, motor function, epilepsy and sleep. Moreover, Grm7 knockout mice exhibit an attenuated response to amphetamine. These findings provide rationale for further investigation of mGlu7 as a potential therapeutic target for neurodevelopmental disorders such as idiopathic autism, attention deficit hyperactivity disorder and Rett syndrome.

mGlu5 Positive Allosteric Modulators Facilitate Long-Term Potentiation via Disinhibition Mediated by mGlu5-Endocannabinoid Signaling.

Metabotropic glutamate (mGlu) receptor type 5 (mGlu5) positive allosteric modulators (PAMs) enhance hippocampal long-term potentiation (LTP) and have cognition-enhancing effects in animal models. These effects were initially thought to be mediated by potentiation of mGlu5 modulation of N-methyl-d-aspartate receptor (NMDAR) currents. However, a biased mGlu5 PAM that potentiates Gαq-dependent mGlu5 signaling, but not mGlu5 modulation of NMDAR currents, retains cognition-enhancing effects in animal models, suggesting that potentiation of NMDAR currents is not required for these in vivo effects of mGlu5 PAMs. However, it is not clear whether the potentiation of NMDAR currents is critical for the ability of mGlu5 PAMs to enhance hippocampal LTP. We now report the characterization of effects of two structurally distinct mGlu5 PAMs, VU-29 and VU0092273, on NMDAR currents and hippocampal LTP. As with other mGlu5 PAMs that do not display observable bias for potentiation of NMDAR currents, VU0092273 enhanced both mGlu5 modulation of NMDAR currents and induction of LTP at the hippocampal Schaffer collateral (SC)-CA1 synapse. In contrast, VU-29 did not potentiate mGlu5 modulation of NMDAR currents but induced robust potentiation of hippocampal LTP. Interestingly, both VU-29 and VU0092273 suppressed evoked inhibitory postsynaptic currents (eIPSCs) in CA1 pyramidal cells, and this effect was blocked by the cannabinoid receptor type 1 (CB1) antagonist AM251. Furthermore, AM251 blocked the ability of both mGlu5 PAMs to enhance LTP. Finally, both PAMs failed to enhance LTP in mice with the restricted genetic deletion of mGlu5 in CA1 pyramidal cells. Taken together with previous findings, these results suggest that enhancement of LTP by mGlu5 PAMs does not depend on mGlu5 modulation of NMDAR currents but is mediated by a previously established mechanism in which mGlu5 in CA1 pyramidal cells induces endocannabinoid release and CB1-dependent disinhibition.

A Coordinated Attack: Rett Syndrome Therapeutic Development.

Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in the Methyl CpG binding protein 2 (MeCP2) gene. This Science & Society article focuses on pharmacological strategies that attack RTT treatment from multiple angles, including drug repurposing and de novo discovery efforts, and discusses the impacts of preclinical study design and translationally relevant outcome measures.

Shaping nursing profession regulation through history - a systematic review.

AIM: The aim of this systematic review was to provide a critical synthesis of the factors that historically shaped the advancements of nursing regulators worldwide. BACKGROUND: An in-depth examination of the different factors that moulded regulatory changes over time is pivotal to comprehend current issues in nursing. INTRODUCTION: In the light of global health scenarios, the researchers explored the factors that historically influenced the socio-contextual circumstances upon which governments made regulatory changes. METHODS: A systematic search was performed on the following databases: PubMed, CINAHL, Scopus, OpenGrey and ScienceDirect. The review included papers from January 2000 to October 2016 published in English. The authors used the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) and an inductive thematic approach for synthesis. RESULTS: Two main themes were identified: factors underpinning current challenges and historical and contextual triggers of regulation. The first theme was composed of three aspects: education, migration and internationalization, and policy and regulation; the second theme consisted of four attributes: demographics, economics, history of registration and wars, and historical changes in nursing practice. DISCUSSION: Factors that shaped nursing regulation were linked to changing demographics and economics, education, history of nursing registration, shifting patterns of migration and internationalization, nursing practice, policy and regulation and significant societal turns often prompted by wars. CONCLUSION: A deeper understanding of the developments of the nursing regulatory institutions provides the foundation for portable standards that can be applied across an array of jurisdictions to guarantee a better public safety. IMPLICATION FOR NURSING AND HEALTH POLICY: Understanding factors that socially, legislatively and politically have influenced the development of regulatory bodies over time helps to mould local, national and international policies that have a stronger impact on health worldwide. To achieve this, there must be effective cooperation among systems of nursing regulations globally.

Mechanisms underlying prelimbic prefrontal cortex mGlu3/mGlu5-dependent plasticity and reversal learning deficits following acute stress.

Stress can precipitate or worsen symptoms of many psychiatric illnesses. Dysregulation of the prefrontal cortex (PFC) glutamate system may underlie these disruptions and restoring PFC glutamate signaling has emerged as a promising avenue for the treatment of stress disorders. Recently, we demonstrated that activation of metabotropic glutamate receptor subtype 3 (mGlu3) induces a postsynaptic form of long-term depression (LTD) that is dependent on the activity of another subtype, mGlu5. Stress exposure disrupted this plasticity, but the underlying signaling mechanisms and involvement in higher-order cognition have not yet been investigated. Acute stress was applied by 20-min restraint and early reversal learning was evaluated in an operant-based food-seeking task. We employed whole-cell patch-clamp recordings of layer 5 prelimbic (PL)-PFC pyramidal cells to examine mGlu3-LTD and several mechanistically distinct mGlu5-dependent functions. Acute stress impaired both mGlu3-LTD and early reversal learning. Interestingly, potentiating mGlu5 signaling with the mGlu5 positive allosteric modulator (PAM) VU0409551 rescued stress-induced deficits in both mGlu3-LTD and reversal learning. Other aspects of PL-PFC mGlu5 function were not disrupted following stress; however, signaling downstream of mGlu5-Homer interactions, phosphoinositide-3-kinase (PI3K), Akt, and glycogen synthase kinase 3ß was implicated in these phenomena. These findings demonstrate that acute stress disrupts early reversal learning and PL-PFC-dependent synaptic plasticity and that potentiating mGlu5 function can restore these impairments. These findings provide a framework through which modulating coordinated mGlu3/mGlu5 signaling may confer benefits for the treatment of stress-related psychiatric disorders.

In Vitro to in Vivo Translation of Allosteric Modulator Concentration-Effect Relationships: Implications for Drug Discovery.

Allosteric modulation of GPCRs represents an increasingly explored approach in drug development. Due to complex pharmacology, however, the relationship(s) between modulator properties determined in vitro with in vivo concentration-effect phenomena is frequently unclear. We investigated key pharmacological properties of a set of metabotropic glutamate receptor 5 (mGlu5) positive allosteric modulators (PAMs) and their relevance to in vivo concentration-response relationships. These studies identified a significant relationship between in vitro PAM cooperativity (αß), as well as the maximal response obtained from a simple in vitro PAM concentration-response experiment, with in vivo efficacy for reversal of amphetamine-induced hyperlocomotion. This correlation did not exist with PAM potency or affinity. Data across PAMs were then converged to calculate an in vivo concentration of glutamate putatively relevant to the mGlu5 PAM mechanism of action. This work demonstrates the ability to merge in vitro pharmacology profiles with relevant behavioral outcomes and also provides a novel method to estimate neurotransmitter concentrations in vivo.