Dopamine D1-like receptors modulate synchronized oscillations in the hippocampal-prefrontal-amygdala circuit in contextual fear.

Contextual fear conditioning (CFC) is mediated by a neural circuit that includes the hippocampus, prefrontal cortex, and amygdala, but the neurophysiological mechanisms underlying the regulation of CFC by neuromodulators remain unclear. Dopamine D1-like receptors (D1Rs) in this circuit regulate CFC and local synaptic plasticity, which is facilitated by synchronized oscillations between these areas. In rats, we determined the effects of systemic D1R blockade on CFC and oscillatory synchrony between dorsal hippocampus (DH), prelimbic (PL) cortex, basolateral amygdala (BLA), and ventral hippocampus (VH), which sends hippocampal projections to PL and BLA. D1R blockade altered DH-VH and reduced VH-PL and VH-BLA synchrony during CFC, as inferred from theta and gamma coherence and theta-gamma coupling. D1R blockade also impaired CFC, as indicated by decreased freezing at retrieval, which was characterized by altered DH-VH and reduced VH-PL, VH-BLA, and PL-BLA synchrony. This reduction in VH-PL-BLA synchrony was not fully accounted for by non-specific locomotor effects, as revealed by comparing between epochs of movement and freezing in the controls. These results suggest that D1Rs regulate CFC by modulating synchronized oscillations within the hippocampus-prefrontal-amygdala circuit. They also add to growing evidence indicating that this circuit synchrony at retrieval reflects a neural signature of learned fear.

Attitudes to the use of animals in biomedical research: Effects of stigma and selected research project summaries.

Three groups of participants (largely recruited from the UK) completed a survey to examine attitudes to the use of animals in biomedical research, after reading the lay (N = 182) or technical (N = 201) summary of a research project, or no summary (N = 215). They then completed a survey comprising the animal attitude (AAS), animal purpose (APQ), belief in animal mind (BAM) and empathy quotient (EQ) scales. The APQ was adapted to assess attitudes towards the use of animals for research into disorders selected to be perceived as controllable and so 'blameworthy' and potentially stigmatised (addiction and obesity) and 'psychological' (schizophrenia and addiction) versus 'physical' (cardiovascular disease and obesity), across selected species (rats, mice, fish pigs and monkeys). Thus, the APQ was used to examine how the effects of perceived controllability and the nature of the disorder affected attitudes to animal use, in different species and in the three summary groups. As expected, attitudes to animal use as measured by the AAS and the APQ (total) correlated positively with BAM and EQ scores, consistent with the assumption that the scales all measured pro-welfare attitudes. Participants in the two research summary groups did not differentiate the use of rats, mice and fish (or fish and pigs in the technical summary group), whereas all species were differentiated in the no summary group. Participants given the lay summary were as concerned about the use of animals for schizophrenia as for addiction research. APQ ratings otherwise indicated more concern for animals used for addiction research (and for obesity compared to cardiovascular disease in all summary groups). Therefore, the information provided by a research project summary influenced attitudes to use of animals in biomedical research. However, there was no overall increase in agreement with animal use in either of the summary groups.

Pharmacological modulation of Kv3 voltage-gated potassium channels regulates fear discrimination and expression in a response-dependent manner.

Various psychiatric diseases are characterized by aberrant cognition and emotional regulation. This includes inappropriately attributing affective salience to innocuous cues, which can be investigated using translationally relevant preclinical models of fear discrimination. Activity in the underpinning corticolimbic circuitry is governed by parvalbumin-expressing GABAergic interneurons, which also regulate fear discrimination. Kv3 voltage-gated potassium channels are highly expressed in these neurons and are important for controlling their activity, suggesting that pharmacological Kv3 modulation may regulate fear discrimination. We determined the effect of the positive Kv3 modulator AUT00206 given systemically to female rats undergoing limited or extended auditory fear discrimination training, which we have previously shown results in more discrimination or generalization, respectively, based on freezing at retrieval. We also characterized darting and other active fear-related responses. We found that limited training resulted in more discrimination based on freezing, which was unaffected by AUT00206. In contrast, extended training resulted in more generalization based on freezing and the emergence of discrimination based on darting during training and, to a lesser extent, at retrieval. Importantly, AUT00206 given before extended training had dissociable effects on fear discrimination and expression at retrieval depending on the response examined. While AUT00206 mitigated generalization without affecting expression based on freezing, it reduced expression without affecting discrimination based on darting, although darting levels were low overall. These results indicate that pharmacological Kv3 modulation regulates fear discrimination and expression in a response-dependent manner. They also raise the possibility that targeting Kv3 channels may ameliorate perturbed cognition and emotional regulation in psychiatric disease.

Camellia sinensis solvent extract, epigallocatechin gallate and caffeine confer trophocidal and cysticidal effects against Acanthamoeba castellanii.

We examined the anti-acanthamoebic efficacy of green tea Camellia sinensis solvent extract (SE) or its chemical constituents against Acanthamoeba castellanii by using anti-trophozoite, anti-encystation, and anti-excystation assays. C. sinensis SE (625-5000 µg/mL) inhibited trophozoite replication within 24-72 h. C. sinensis SE exhibited a dose-dependent inhibition of encystation, with a marked cysticidal activity at 2500-5000 µg/mL. Two constituents of C. sinensis, namely epigallocatechin-3-gallate and caffeine, at 100 µM and 200 µM respectively, significantly inhibited both trophozoite replication and encystation. Cytotoxicity analysis showed that 156.25-2500 µg/mL of SE was not toxic to human corneal epithelial cells, while up to 625 µg/mL was not toxic to Madin-Darby canine kidney cells. This study shows the anti-acanthamoebic potential of C. sinensis SE against A. castellanii trophozoites and cysts. Pre-clinical studies are required to elucidate the in vivo efficacy and safety of C. sinensis SE.

URB597 induces subtle changes to aggression in adult Lister Hooded rats.

The endocannabinoid system has been implicated in both social and cognitive processing. The endocannabinoid metabolism inhibitor, URB597, dose-dependently improves non-social memory in adult Wistar and Sprague Dawley rats, whereas its effect on social interaction (SI) is affected by both rat strain and drug dose. Lister Hooded rats consistently respond differently to drug treatment in general compared with albino strains. This study sought to investigate the effects of different doses of URB597 on social and non-social memory in Lister Hooded rats, as well as analyzing the behavioral composition of the SI. Males were tested for novel object recognition (NOR), social preference (between an object and an unfamiliar rat), social novelty recognition (for a familiar vs. unfamiliar rat) and SI with an unfamiliar rat. URB597 (0.1 or 0.3 mg/kg) or vehicle was given 30 min before testing. During SI testing, total interaction time was assessed along with time spent on aggressive and explorative behaviors. Lister Hooded rats displayed expected non-social and social memory and social preference, which was not affected by URB597. During SI, URB597 did not affect total interaction time. However, the high dose increased aggression, compared to vehicle, and decreased anogenital sniffing, compared to the low dose of URB597. In summary, URB597 did not affect NOR, social preference or social recognition memory but did have subtle behavioral effects during SI in Lister hooded rats. Based on our findings we argue for the importance of considering strain as well as the detailed composition of behavior when investigating drug effects on social behavior.

Cannabidiol Prevents Spontaneous Fear Recovery after Extinction and Ameliorates Stress-Induced Extinction Resistance.

Cannabidiol, the main non-psychotropic constituent of cannabis, has potential as a treatment for anxiety-related disorders since it reduces learned fear expression and enhances fear extinction. The return of fear over time after successful extinction and stress-induced extinction resistance are potential barriers to the treatment of these disorders with extinction-based psychological therapy. In two experiments using rats subjected to auditory fear conditioning, we determined the effects of systemic cannabidiol treatment on (1) delayed extinction and later spontaneous fear recovery, and (2) extinction resistance caused by immediate extinction (the immediate extinction deficit (IED)). In Experiment 1, cannabidiol was given before delayed extinction occurring 24 h after conditioning, with extinction recall and spontaneous fear recovery tested drug-free 1 and 21 days after extinction, respectively. We found that cannabidiol had no effect on extinction recall but it prevented spontaneous fear recovery. In Experiment 2, the IED procedure was first validated, with immediate extinction occurring 30 min after conditioning. We confirmed that immediate extinction impaired extinction recall, compared to delayed extinction. Next, cannabidiol was given before immediate or no extinction, with extinction recall tested drug-free the next day. We found that cannabidiol rescued the IED, which did not involve effects on fear memory consolidation. In summary, cannabidiol prevented spontaneous fear recovery after delayed extinction and ameliorated extinction resistance caused by immediate extinction. Although the pharmacological mechanisms underlying these effects remain to be determined, our results add to evidence indicating that cannabidiol might prove useful as an adjunct for potentiating the psychological treatment of anxiety-related disorders.

Illuminating Host-Parasite Interaction at the Cellular and Subcellular Levels with Infrared Microspectroscopy.

Toxoplasma gondii (T. gondii) is an opportunistic protozoan that can cause brain infection and other serious health consequences in immuno-compromised individuals. This parasite has a remarkable ability to cross biological barriers and exploit the host cell microenvironment to support its own survival and growth. Recent advances in label-free spectroscopic imaging techniques have made it possible to study biological systems at a high spatial resolution. In this study, we used conventional Fourier-transform infrared (FTIR) microspectroscopy and synchrotron-based FTIR microspectroscopy to analyze the chemical changes that are associated with infection of human brain microvascular endothelial cells (hBMECs) by T. gondii (RH) tachyzoites. Both FTIR microspectroscopic methods showed utility in revealing the chemical alterations in the infected hBMECs. Using a ZnS hemisphere device, to increase the numerical aperture, and the synchrotron source to increase the brightness, we obtained spatially resolved spectra from within a single cell. The spectra extracted from the nucleus and cytosol containing the tachyzoites were clearly distinguished. RNA sequencing analysis of T. gondii-infected and uninfected hBMECs revealed significant changes in the expression of host cell genes and pathways in response to T. gondii infection. These FTIR spectroscopic and transcriptomic findings provide significant insight into the molecular changes that occur in hBMECs during T. gondii infection.

Hippocampal Disinhibition Reduces Contextual and Elemental Fear Conditioning While Sparing the Acquisition of Latent Inhibition.

Hippocampal neural disinhibition, i.e., reduced GABAergic inhibition, is a key feature of schizophrenia pathophysiology. The hippocampus is an important part of the neural circuitry that controls fear conditioning and can also modulate prefrontal and striatal mechanisms, including dopamine signaling, which play a role in salience modulation. Consequently, hippocampal neural disinhibition may contribute to impairments in fear conditioning and salience modulation reported in schizophrenia. Therefore, we examined the effect of ventral hippocampus (VH) disinhibition in male rats on fear conditioning and salience modulation, as reflected by latent inhibition (LI), in a conditioned emotional response (CER) procedure. A flashing light was used as the conditioned stimulus (CS), and conditioned suppression was used to index conditioned fear. In experiment 1, VH disinhibition via infusion of the GABA-A receptor antagonist picrotoxin before CS pre-exposure and conditioning markedly reduced fear conditioning to both the CS and context; LI was evident in saline-infused controls but could not be detected in picrotoxin-infused rats because of the low level of fear conditioning to the CS. In experiment 2, VH picrotoxin infusions only before CS pre-exposure did not affect the acquisition of fear conditioning or LI. Together, these findings indicate that VH neural disinhibition disrupts contextual and elemental fear conditioning, without affecting the acquisition of LI. The disruption of fear conditioning resembles aversive conditioning deficits reported in schizophrenia and may reflect a disruption of neural processing both within the hippocampus and in projection sites of the hippocampus.

In it together? The case for endocannabinoid-noradrenergic interactions in fear extinction.

Anxiety and trauma-related disorders, such as post-traumatic stress disorder (PTSD), are debilitating mental illnesses with great personal and socioeconomic costs. Examining memory formation and relevant behavioural responding associated with aversive stimuli may improve our understanding of the neurobiology underlying fear memory processing and PTSD treatment. The neurocircuitry underpinning learned fear and its inhibition through extinction is complex, involving synergistic interactions between different neurotransmitter systems in inter-connected brain areas. Endocannabinoid and noradrenergic transmission have both been implicated separately in fear memory processing and PTSD, but potential interactions between these systems in relation to fear extinction have received little attention to date. Their receptors are expressed together in brain areas crucial for fear extinction, which is enhanced by both cannabinoid and noradrenergic receptor activation in these areas. Moreover, cannabinoid signalling modulates the activity of locus coeruleus noradrenaline (NA) neurons and the release of NA in the medial prefrontal cortex, a brain area that is crucial for fear extinction. Interestingly, endocannabinoid-noradrenergic system interactions have been shown to regulate the encoding and retrieval of fear memory. Thus, noradrenergic regulation of fear extinction may also be driven indirectly in part via cannabinoid receptor signalling. In this perspective paper, we collate the available relevant literature and propose a synergistic role for the endocannabinoid and noradrenergic systems in regulating fear extinction, the study of which may further our understanding of the neurobiological substrates of PTSD and its treatment.

Endocannabinoid metabolism inhibition has no effect on spontaneous fear recovery or extinction resistance in Lister hooded rats.

Endocannabinoid transmission is emerging as a target for treating anxiety-related disorders, given its regulation of fear extinction. Boosting anandamide levels via inhibition of its metabolism by fatty acid amide hydrolase (FAAH) can enhance extinction, whereas inhibiting monoacylglycerol lipase (MAGL) to elevate 2-arachidonoylglycerol levels can impair extinction. However, whether endocannabinoids regulate fear relapse over time or extinction resistance remains unclear. In two experiments using auditory fear conditioned rats, we examined the effects of the FAAH inhibitor URB597 and the MAGL inhibitor JZL184 administered systemically on 1) spontaneous fear recovery after delayed extinction, and 2) extinction resistance resulting from immediate extinction [the immediate extinction deficit (IED)]. In Experiment 1, URB597 or JZL184 was given immediately after delayed extinction occurring 24 h after conditioning. Extinction recall and spontaneous fear recovery were tested drug-free 1 and 21 days later, respectively. We found no effects of either drug on extinction recall or spontaneous fear recovery. In Experiment 2, URB597 or JZL184 was given before immediate extinction occurring 30 min after conditioning and extinction recall was tested drug-free the next day. We also examined the effects of propranolol, a beta-adrenoceptor antagonist that can rescue the IED, as a positive control. JZL184 enhanced fear expression and impaired extinction learning but we found no lasting effects of URB597 or JZL184 on cued extinction recall. Propranolol reduced fear expression but, unexpectedly, had no enduring effect on extinction recall. The results are discussed in relation to various methodological differences between previous studies examining endocannabinoid and adrenergic regulation of fear extinction.

Could Cannabidiol Be a Treatment for Coronavirus Disease-19-Related Anxiety Disorders?

Coronavirus disease-19 (COVID-19)-related anxiety and post-traumatic stress symptoms (PTSS) or post-traumatic stress disorder (PTSD) are likely to be a significant long-term issue emerging from the current pandemic. We hypothesize that cannabidiol (CBD), a chemical isolated from Cannabis sativa with reported anxiolytic properties, could be a therapeutic option for the treatment of COVID-19-related anxiety disorders. In the global over-the-counter CBD market, anxiety, stress, depression, and sleep disorders are consistently the top reasons people use CBD. In small randomized controlled clinical trials, CBD (300-800 mg) reduces anxiety in healthy volunteers, patients with social anxiety disorder, those at clinical high risk of psychosis, in patients with Parkinson's disease, and in individuals with heroin use disorder. Observational studies and case reports support these findings, extending to patients with anxiety and sleep disorders, Crohn's disease, depression, and in PTSD. Larger ongoing trials in this area continue to add to this evidence base with relevant patient cohorts, sample sizes, and clinical end-points. Pre-clinical studies reveal the molecular targets of CBD in these indications as the cannabinoid receptor type 1 and cannabinoid receptor type 2 (mainly in fear memory processing), serotonin 1A receptor (mainly in anxiolysis) and peroxisome proliferator-activated receptor gamma (mainly in the underpinning anti-inflammatory/antioxidant effects). Observational and pre-clinical data also support CBD's therapeutic value in improving sleep (increased sleep duration/quality and reduction in nightmares) and depression, which are often comorbid with anxiety. Together these features of CBD make it an attractive novel therapeutic option in COVID-related PTSS that merits investigation and testing through appropriately designed randomized controlled trials.

Dopamine regulation of contextual fear and associated neural circuit function.

Learning to associate certain contexts with threat and adapting to changing environmental contingencies by learning that such contexts are no longer associated with threat are both crucial for survival. Research over the last few decades has made considerable progress in determining the brain areas involved in the encoding, retrieval and extinction of contextual fear. These studies have identified the hippocampus and amygdala, along with the prefrontal cortex and other inter-connected brain areas, as key players in contextual fear processing. In contrast to the neural circuit basis of contextual fear, the neurochemical mechanisms involved in its regulation remain poorly understood. Dopamine is well known for its role in appetitive learning but this neurotransmitter is also important for other types of learning, including spatial and aversive memory processing. Dopamine is ideally positioned to regulate contextual fear given that the areas involved receive dopamine input and express dopamine receptors. Moreover, neuronal activity, functional connectivity and synaptic plasticity in this neural circuitry are modulated by dopamine receptor signalling. Here, we review the evidence indicating that dopamine regulates various contextual fear processes, along with the more recent studies that have begun to elucidate the brain areas and neurophysiological mechanisms involved. From a fundamental research perspective, understanding how dopamine regulates contextual fear will lead to novel insights on the neurochemical modulation of neural circuit function underlying memory processing. This research may also have translational relevance given that contextual fear conditioning and extinction also provide useful preclinical models of certain aspects of anxiety-related disorders and their treatment.

Structural, Functional, and Metabolic Alterations in Human Cerebrovascular Endothelial Cells during Toxoplasma gondii Infection and Amelioration by Verapamil In Vitro.

Toxoplasma gondii (T. gondii), the causative agent of toxoplasmosis, is a frequent cause of brain infection. Despite its known ability to invade the brain, there is still a dire need to better understand the mechanisms by which this parasite interacts with and crosses the blood-brain barrier (BBB). The present study revealed structural and functional changes associated with infection and replication of T. gondii within human brain microvascular endothelial cells (BMECs) in vitro. T. gondii proliferated within the BMECs and disrupted the integrity of the cerebrovascular barrier through diminishing the cellular viability, disruption of the intercellular junctions and increasing permeability of the BMEC monolayer, as well as altering lipid homeostasis. Proton nuclear magnetic resonance (1H NMR)-based metabolomics combined with multivariate data analysis revealed profiles that can be attributed to infection and variations in the amounts of certain metabolites (e.g., amino acids, fatty acids) in the extracts of infected compared to control cells. Notably, treatment with the Ca2+ channel blocker verapamil rescued BMEC barrier integrity and restricted intracellular replication of the tachyzoites regardless of the time of treatment application (i.e., prior to infection, early- and late-infection). This study provides new insights into the structural and functional changes that accompany T. gondii infection of the BMECs, and sheds light upon the ability of verapamil to inhibit the parasite proliferation and to ameliorate the adverse effects caused by T. gondii infection.

In vitro activity of Camellia sinensis (green tea) against trophozoites and cysts of Acanthamoeba castellanii.

The effect of Camellia sinensis (green tea) on the growth of Acanthamoeba castellanii trophozoites was examined using a microplate based-Sulforhodamine B (SRB) assay. C. sinensis hot and cold brews at 75% and 100% concentrations significantly inhibited the growth of trophozoites. We also examined the structural alterations in C. sinensis-treated trophozoites using transmission electron microscopy (TEM) and scanning electron microscopy (SEM). This analysis showed that C. sinensis compromised the cell membrane integrity and caused progressive destruction of trophozoites. C. sinensis also significantly inhibited the parasite's ability to form cysts in a dose-dependent manner and reduced the rate of excystation from cysts to trophozoites. C. sinensis exhibited low cytotoxic effects on primary corneal stromal cells. However, cytotoxicity was more pronounced in SV40-immortalized corneal epithelial cells. Chromatographic analysis showed that both hot and cold C. sinensis brews contained the same number and type of chemical compounds. This work demonstrated that C. sinensis has anti-acanthamoebic activity against trophozoite and cystic forms of A. castellanii. Further studies are warranted to identify the exact substances in C. sinensis that have the most potent anti-acanthamoebic effect.

Sex differences in auditory fear discrimination are associated with altered medial prefrontal cortex function.

The increased prevalence of post-traumatic stress disorder (PTSD) that is observed in women may involve sex differences in learned fear inhibition and medial prefrontal cortex (mPFC) function. PTSD is characterized by fear overgeneralization involving impaired fear regulation by safety signals. We recently found that males show fear discrimination and females show fear generalization involving reduced safety signalling after extended fear discrimination training. Here we determined if these sex differences involve altered mPFC function. Male and female rats underwent three days of auditory fear discrimination training, where one tone (CS+) was paired with footshock and another tone (CS-) was presented alone. Local field potentials were recorded from prelimbic (PL) and infralimbic (IL) mPFC during retrieval. We found that males discriminated and females generalized based on cue-induced freezing at retrieval. This was accompanied by sex differences in basal theta and gamma oscillations in PL and IL. Importantly, males also showed PL/IL theta activation during safety signalling by the CS- and IL gamma activation in response to the threat-related CS+, both of which were absent in females. These results add to growing evidence indicating that sex differences in learned fear inhibition are associated with altered mPFC function.

Improved functional connectivity network estimation for brain networks using multivariate partial coherence.

OBJECTIVE: Graphical networks and network metrics are widely used to understand and characterise brain networks and brain function. These methods can be applied to a range of electrophysiological data including electroencephalography, local field potential and single unit recordings. Functional networks are often constructed using pair-wise correlation between variables. The objective of this study is to demonstrate that functional networks can be more accurately estimated using partial correlation than with pair-wise correlation. APPROACH: We compared network metrics derived from unconditional and conditional graphical networks, obtained using coherence and multivariate partial coherence (MVPC), respectively. Graphical networks were constructed using coherence and MVPC estimates, and binary and weighted network metrics derived from these: node degree, path length, clustering coefficients and small-world index. MAIN RESULTS: Network metrics were applied to simulated and experimental single unit spike train data. Simulated data used a 10x10 grid of simulated cortical neurons with centre-surround connectivity. Conditional network metrics gave a more accurate representation of the known connectivity: Numbers of excitatory connections had range 3-11, unconditional binary node degree had range 6-80, conditional node degree had range 2-13. Experimental data used multi-electrode array recording with 19 single-units from left and right hippocampal brain areas in a rat model for epilepsy. Conditional network analysis showed similar trends to simulated data, with lower binary node degree and longer binary path lengths compared to unconditional networks. SIGNIFICANCE: We conclude that conditional networks, where common dependencies are removed through partial coherence analysis, give a more accurate representation of the interactions in a graphical network model. These results have important implications for graphical network analyses of brain networks and suggest that functional networks should be derived using partial correlation, based on MVPC estimates, as opposed to the common approach of pair-wise correlation.

The neurobiological basis of sex differences in learned fear and its inhibition.

Learning that certain cues or environments predict threat enhances survival by promoting appropriate fear and the resulting defensive responses. Adapting to changing stimulus contingencies by learning that such cues no longer predict threat, or distinguishing between these threat-related and other innocuous stimuli, also enhances survival by limiting fear responding in an appropriate manner to conserve resources. Importantly, a failure to inhibit fear in response to harmless stimuli is a feature of certain anxiety and trauma-related disorders, which are also associated with dysfunction of the neural circuitry underlying learned fear and its inhibition. Interestingly, these disorders are up to twice as common in women, compared to men. Despite this striking sex difference in disease prevalence, the neurobiological factors involved remain poorly understood. This is due in part to the majority of relevant preclinical studies having neglected to include female subjects alongside males, which has greatly hindered progress in this field. However, more recent studies have begun to redress this imbalance and emerging evidence indicates that there are significant sex differences in the inhibition of learned fear and associated neural circuit function. This paper provides a narrative review on sex differences in learned fear and its inhibition through extinction and discrimination, along with the key gonadal hormone and brain mechanisms involved. Understanding the endocrine and neural basis of sex differences in learned fear inhibition may lead to novel insights on the neurobiological mechanisms underlying the enhanced vulnerability to develop anxiety-related disorders that are observed in women.

Cannabinoid Regulation of Fear and Anxiety: an Update.

PURPOSE OF REVIEW: Anxiety- and trauma-related disorders are prevalent and debilitating mental illnesses associated with a significant socioeconomic burden. Current treatment approaches often have inadequate therapeutic responses, leading to symptom relapse. Here we review recent preclinical and clinical findings on the potential of cannabinoids as novel therapeutics for regulating fear and anxiety. RECENT FINDINGS: Evidence from preclinical studies has shown that the non-psychotropic phytocannabinoid cannabidiol and the endocannabinoid anandamide have acute anxiolytic effects and also regulate learned fear by dampening its expression, enhancing its extinction and disrupting its reconsolidation. The findings from the relevant clinical literature are still very preliminary but are nonetheless encouraging. Based on this preclinical evidence, larger-scale placebo-controlled clinical studies are warranted to investigate the effects of cannabidiol in particular as an adjunct to psychological therapy or medication to determine its potential utility for treating anxiety-related disorders in the future.

Dopamine D1-like receptors in the dorsomedial prefrontal cortex regulate contextual fear conditioning.

RATIONALE: Dopamine D1 receptor (D1R) signalling is involved in contextual fear conditioning. The D1R antagonist SCH23390 impairs the acquisition of contextual fear when administered systemically or infused locally into the dorsal hippocampus or basolateral amygdala. OBJECTIVES: We determined if state dependency may account for the impairment in contextual fear conditioning caused by systemic SCH23390 administration. We also examined if the dorsomedial prefrontal cortex (dmPFC), nucleus accumbens (NAc), and ventral hippocampus (VH) are involved in mediating the effect of systemic SCH23390 treatment on contextual fear conditioning. METHODS: In experiment 1, SCH23390 (0.1 mg/kg) or vehicle was given before contextual fear conditioning and/or retrieval. In experiment 2, SCH23390 (2.5 µg/0.5 uL) or vehicle was infused locally into dmPFC, NAc, or VH before contextual fear conditioning, and retrieval was tested drug-free. Freezing was quantified as a measure of contextual fear. RESULTS: In experiment 1, SCH23390 given before conditioning or before both conditioning and retrieval decreased freezing at retrieval, whereas SCH23390 given only before retrieval had no effect. In experiment 2, SCH23390 infused into dmPFC before conditioning decreased freezing at retrieval, while infusion of SCH23390 into NAc or VH had no effect. CONCLUSIONS: The results of experiment 1 confirm those of previous studies indicating that D1Rs are required for the acquisition but not retrieval of contextual fear and rule out state dependency as an explanation for these findings. Moreover, the results of experiment 2 provide evidence that dmPFC is also part of the neural circuitry through which D1R signalling regulates contextual fear conditioning.

Metallome of cerebrovascular endothelial cells infected with Toxoplasma gondii using µ-XRF imaging and inductively coupled plasma mass spectrometry.

In this study, we measured the levels of elements in human brain microvascular endothelial cells (ECs) infected with T. gondii. ECs were infected with tachyzoites of the RH strain, and at 6, 24, and 48 hours post infection (hpi), the intracellular concentrations of elements were determined using a synchrotron-microfocus X-ray fluorescence microscopy (µ-XRF) system. This method enabled the quantification of the concentrations of Zn and Ca in infected and uninfected (control) ECs at sub-micron spatial resolution. T. gondii-hosting ECs contained less Zn than uninfected cells only at 48 hpi (p < 0.01). The level of Ca was not significantly different between infected and control cells (p > 0.05). Inductively Coupled Plasma Mass Spectrometry (ICP-MS) analysis revealed infection-specific metallome profiles characterized by significant increases in the intracellular levels of Zn, Fe, Mn and Cu at 48 hpi (p < 0.01), and significant reductions in the extracellular concentrations of Co, Cu, Mo, V, and Ag at 24 hpi (p < 0.05) compared with control cells. Zn constituted the largest part (74%) of the total metal composition (metallome) of the parasite. Gene expression analysis showed infection-specific upregulation in the expression of five genes, MT1JP, MT1M, MT1E, MT1F, and MT1X, belonging to the metallothionein gene family. These results point to a possible correlation between T. gondii infection and increased expression of MT1 isoforms and altered intracellular levels of elements, especially Zn and Fe. Taken together, a combined µ-XRF and ICP-MS approach is promising for studies of the role of elements in mediating host-parasite interaction.