Open Arms Of Elevated Plus-maze Research Articles

6569 An Increase, Rather Than Absolute Amount, Of Corticotropin-Releasing Hormone (Crh) Drives Mouse Anxiety-Like Behavior

Abstract Disclosure: H.R. Friedman: None. L. Gaston: None. L. Chan: None. J.A. Majzoub: None. Approximately 300 million people have an anxiety disorder. We previously found that anxiety-like behavior (AB) is decreased in mice which lack hypothalamic (HT) Crh (1), so we hypothesized that increased HT Crh, due to primary adrenal insufficiency (PAI), would induce increased AB. To evaluate this, we compared AB in wildtype (WT) mice to those with global deletion of either the Crh gene (CrhKO) or the Mrap gene (MrapKO, causing PAI (2)), measuring: 1) HT Crh mRNA by in situ RNAscope, 2) stress-induced (after 15 min of restraint) plasma corticosterone (CORT) and adrenocorticotropin hormone (ACTH) by ELISA, and 3) AB with elevated plus maze (EPM), and open field (OF) automatically-recorded behaviors. Comparisons were evaluated by ANOVA with post hoc Bonferroni corrections vs. WT. As expected, Crh mRNA was highest in MrapKO (n=16) compared to WT (n=11), and undetectable in CrhKO (n=10), and post-restraint CORT was highest in WT (11±2 mcg/dL), and undetectable in CrhKO and MrapKO. CrhKO mice displayed decreased AB (increased EPM open arm duration) vs. WT animals (140±11 vs. 68±8 s, respectively, P<0.0001). However, AB was comparable between MrapKO (97±10 s) vs. WT (68±8 s; P=0.0807) despite higher HT Crh mRNA in the former. To determine if chronically high vs. a dynamic rise in Crh mRNA is associated with increased AB, we treated MrapKO mice with dexamethasone (DEX) for 5-8 weeks to suppress HT Crh mRNA and assayed hypothalamic-pituitary-adrenal axis function and behavior both on DEX (MrapKO-D; N=4) and 1-week post-withdrawal (MrapKO-DW; N=5). As expected, ACTH was highest in the MrapKO-DW vs. WT (1689±535 vs. 378±35 pg/mL, respectively; P<0.0001) and undetectable in MrapKO-D. Notably, MrapKO-DW mice displayed increased AB (decreased OF center duration) vs. WT animals (56±3 s vs. 114±10 s; P=0.0439). In summary, AB was not present in MrapKO mice with chronically elevated HT Crh mRNA but only developed after they were DEX-treated followed by withdrawal from treatment. These data suggest that in mice, a dynamic rise in HT Crh, but not a chronically elevated level, drives AB, and suggest that Crh is a possible target for anxiety treatment.

The Comparative Effects Between Long-Term and Short-Term Treatment of Finasteride on Anxiety-Like and Depression-Like Behaviors in Early Senescent Male Rats.

This study aims to compare the efficacy of 5-alpha-reductase inhibitors (5ARIs) on anxiety and depression between long-term and short-term treatment followed by withdrawal in d-galactose (Dgal)-induced senescent male rats. Thirty-two, 8-week-old, male Wistar rats were divided into two groups: control rats and Dgal-treated rats (150 mg/kg/day; subcutaneously) for 18 weeks. At week 13, Dgal-treated rats were subdivided into three subgroups: (1) vehicle (DgV), (2) long-term treatment with 5ARIs, Finasteride 5 mg/kg/day, per oral for 6 weeks (DgF), (3) short-term treatment with 5ARIs, Finasteride 5 mg/kg/day, per oral for 2 weeks followed by a 4-week withdrawal period (DgW). Anxiety and depression were assessed using the elevated-plus maze (EPM) and splash test (ST). Blood was collected for biochemical analysis. After euthanasia, the brains were removed to examine brain inflammation, oxidative stress, neuroactive steroids, brain metabolites, and brain senescent markers. We found that DgV rats exhibited metabolic disturbance with a reduced preference index of the EPM, and grooming duration in ST. Increased brain neurotoxic metabolites, along with increased brain inflammation/oxidative stress, and reduced microglia complexity were observed in the DgV rats. Both therapeutic approaches improved metabolic parameters and preference index in the open arm of EPM in Dgal-treated rats, while grooming duration and microglia complexity were increased only in DgF rats. Our results indicate that Fin reduces depression-like and anxiety-like behaviors by reducing brain inflammation, oxidative stress, and brain senescent. In conclusion, long-term treatment with 5ARIs is more effective in alleviating depression than short-term treatment followed by withdrawal in Dgal-induced early senescent male rats.

Identification of a Locus Coeruleus-amygdala Angiotensinergic Circuit: Implications for Stress-related Cardiovascular Diseases

Background: The locus coeruleus (LC) is a significant noradrenergic nucleus in the brain sensitive to afferent interoceptive signals. It responds to behavioral challenges by increasing noradrenaline release through ascending projections to the hypothalamus, thalamus, cortex, and amygdala. The brain renin-angiotensin system (RAS) plays a role in stress-related cardiovascular diseases, and previous research has identified angiotensin II (Ang II) and its receptors in the LC. The current study aimed to gain a deeper understanding of the function of the Ang II type 1 receptor (AT1R) in the LC, particularly its involvement in transmitting interoceptive cardiovascular signals by regulating LC activity. Methods: Using AT1R-eGFP and AT1R-Cre mice combined with neuroanatomical tract-tracing, chemogenetic and behavioral approaches, we examined AT1R expressing neurons in the LC. Dual immunohistochemistry was used in AT1R-eGFP reporter mice to characterize LC-AT1R-eGFP+ cells by looking at the colocalization of noradrenergic neuron maker tyrosine hydroxylase (TH) and GFP. Cre-inducible tracing with AT1R-cre mice was applied for circuit anterograde analysis and in vivo chemogenetics were used for behavior testing and analysis. Results: The majority of AT1R-eGFP+ neurons (94%) in the LC were found to be co-localized with TH. By using anterograde labeling with cre-inducible GFP virus (AAV-DIO-GFP) in AT1R-Cre mice, it was revealed that the AT1R+ neurons in the LC predominantly project to the amygdala and extended amygdala regions. Only a few or no projections to other brain areas known to receive LC inputs were observed. Within the amygdala, the AT1R+ nerve terminals were specifically limited to the medial division of the central amygdala (CeM) and the basomedial amygdala (BMA). Subsequently, AT1R was deleted from LC by injecting Cre virus into AT1R-Flox mice. Although the general anxiety level remained unchanged, restraint stress-induced anxiety was attenuated following LC AT1R deletion as shown by increased open arm distance in the elevated plus maze (EPM) test (21.7±6.6mm AAV-GFP vs. 98.1±27.5mm AAV-Cre, p<0.05, n=10). Moreover, the study employed Cre-dependent inhibitory and excitatory designer receptors exclusively activated by designer drug (hM4Di and hM3Dq DREADD) with Clozapine-n-oxide (CNO) to selectively silence and activate the AT1R+ neurons in the LC. The inhibition of AT1R+ neurons decreased the baseline anxiety level in mice (36.6±8.6mm Saline vs. 153.4±38.9mm CNO, p<0.01, n=8, EPM open arm distance), while activation of AT1R+ neurons increased baseline anxiety level (963.1±223.1mm Saline vs. 408.5±92mm CNO, p<0.01, n=10, EPM open arm distance). Additionally, the acoustic startle response, which is associated with anxiety levels, was reduced after silencing the LC AT1R+ neurons (324±39.2mV Saline vs. 168.8±54.1mV CNO, p<0.05, n=8). Conclusion: These findings suggest that a specific group of angiotensinergic LC neurons sends projections to the amygdala, highlighting the potential significance of AT1R in mediating noradrenergic activation between the LC and amygdala. Future investigations will explore the response of LC AT1R+ neurons to stress-induced cardiovascular stimuli and afferent interoceptive LC inputs, which may offer novel insights, and new therapeutic direction for the link between stress disorders and cardiovascular disease risk. NIH 1R01HL137103-01A1. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Unmoving and uninflamed: Characterizing neuroinflammatory dysfunction in the Wistar-Kyoto rat model of depression.

Reductionistic research on depressive disorders has been hampered by the limitations of animal models. Recently, it has been hypothesized that neuroinflammation is a key player in depressive disorders. The Wistar-Kyoto (WKY) rat is an often-used animal model of depression, but no information so far exists on its neuroinflammatory profile. As such, we compared male young adult WKY rats to Wistar (WS) controls, with regard to both behavioral performance and brain levels of key neuroinflammatory markers. We first assessed anxiety- and depression-like behaviors in a battery consisting of the Elevated Plus Maze (EPM), the Novelty Suppressed Feeding (NSFT), Open Field (OFT), Social Interaction (SIT), Forced Swim (FST), Sucrose Preference (SPT), and Splash tests (ST). We found that WKY rats displayed increased NSFT feeding latency, decreased OFT center zone permanence, decreased EPM open arm permanence, decreased SIT interaction time, and increased immobility in the FST. However, WKY rats also evidenced marked hypolocomotion, which is likely to confound performance in such tests. Interestingly, WKY rats performed similarly, or even above, to WS levels in the SPT and ST, in which altered locomotion is not a significant confound. In a separate cohort, we assessed prefrontal cortex (PFC), hippocampus and amygdala levels of markers of astrocytic (GFAP, S100A10) and microglial (Iba1, CD86, Ym1) activation status, as well as of three key proinflammatory cytokines (IL-1β, IL-6, TNF-α). There were no significant differences between strains in any of these markers, in any of the regions assessed. Overall, results highlight that behavioral data obtained with WKY rats as a model of depression must be carefully interpreted, considering the marked locomotor activity deficits displayed. Furthermore, our data suggest that, despite WKY rats replicating many depression-associated neurobiological alterations, as shown by others, this is not the case for neuroinflammation-related alterations, thus representing a novel limitation of this model.

Anti-anxiety and Hypnotic Effects of Lawsonia inermis Hydroalcoholic Extract

Background: Sleep is a vital biological feature, and insomnia causes maladaptive physical and psychological functions. The main flaws of current insomnia medications are significant side effects. A suitable substitute can be herbal products. We aimed to evaluate the anti-anxiety, sleep-inducing, and - prolonging effects of Lawsonia inermis extract and fractions in mice. Methods: Male albino mice were pretreated intraperitoneally (i.p.) with either three doses (40, 80, and 160 mg/kg) of L. inermis extract or n-butanol fraction (NBF), ethyl acetate fraction (EAF), and water fraction (WF), 30 minutes before i.p. injection of 30 mg/kg pentobarbital. Sleep latency and duration of sleep were recorded. For anxiolytic activity, elevated plus-maze (EPM) tests were used. Moreover, the toxicity of the extract was determined in both in vivo and in vitro experiments. Results: L. inermis extract (160 mg/kg) significantly reduced sleep latency and increased sleep duration. EAF at 160 mg/kg decreased sleep latency and increased sleep duration. Flumazenil reversed the hypnotic effect of L. inermis extract (160 mg/kg). L. inermis extract (80, 160 mg/kg) increased the time spent and the number of entries in the open arms of EPM. The tested extracts and fractions administration found no adverse effects on PC12 cell viability. The LD50 was 2.4 g/kg. Conclusion: L. inermis extracts exhibit anxiolytic and hypnotic effects, probably modulating the GABAergic system.

Pharmacological Manipulation of Corticotropin-Releasing Factor Receptors in the Anterior and Posterior Subregions of the Insular Cortex Differently Affects Anxiety-Like Behaviors in the Elevated Plus Maze in Rats.

Neuroimaging data in humans and neurobiological studies in rodents have suggested an involvement of the insular cortex (IC) in anxiety manifestations. However, the local neurochemical mechanisms involved are still poorly understood. Corticotropin-releasing factor (CRF) neurotransmission has been described as a prominent neurochemical mechanism involved in the expression of anxiety-like behaviors, but the brain sites related are poorly understood. Additionally, several findings indicate that control of physiological and behavioral responses by the IC occurs in a site-specific manner along its rostrocaudal axis. Thus, this study is aimed at evaluating the effect of CRF receptor agonism and antagonism within the anterior and posterior subregions of the IC in controlling anxiety-related behaviors in the elevated plus maze (EPM). For this, independent groups (six groups) of animals received bilateral microinjections of vehicle, the selective CRF1 receptor antagonist CP376395, or CRF into either the anterior or posterior subregions of the IC. Ten minutes later, the behavior in the EPM was evaluated for five minutes. Treatment of the anterior IC with CP376395, but not with CRF, increased the time and number of entries into the open arms of the EPM. CRF, but not the CRF1 receptor antagonist, microinjected into the posterior IC also increased exploration of the EPM open arms. Taken together, these data indicate that CRFergic neurotransmission in the anterior IC is involved in the expression of anxiety-related behaviors in the EPM. This neurochemical mechanism does not seem to be activated within the posterior IC during exposure to the EPM, but the effects caused by CRF microinjection indicate that activation of CRF receptors in this IC subregion might evoke anxiolytic-like effects.

Post-natal antibiotic exposure in mother rat (F0) induces anxiety like behavior in adult rat offspring (F1) by activating HPA axis and down-regulating the Nr3c1 gene.

Early postnatal administration of antibiotics has been linked to lasting effects on brain development and behavior. Research conducted on animals that are free from germs has demonstrated that the impact of microbiome colonization on the regulation of the hypothalamic-pituitary-adrenal (HPA) axis and neuroendocrine pathways is substantial, which play a crucial role in stress management. Nevertheless, it is still uncertain if the exposure to antibiotics in rat dams (F0-generation) before weaning is associated with neurobehavioral changes in rat offspring (F1-generation) during adulthood. In order to investigate the effects, we perturbed the intestinal microbiota of rat dams (F0 generation) by administering cefixime (CEF), an antibiotic commonly used for obstetric purposes, at clinically relevant doses (1 mg/kg, 2.5 mg/kg or 5 mg/kg). Anxiety-like behaviors in adult offspring was evaluated through the utilization of elevated plus maze (EPM) and open field paradigm (OFP) following a six-week interval from birth (PND42). Subsequent to behavioral assessments, the rats were euthanized, and their brains and blood was collected for biochemical analysis. Plasma corticosterone concentration was used to assess HPA activity, whereas the quantitative real-time polymerase chain reaction (PCR) was employed to determine the transcription levels of the glucocorticoid receptor (GR) Nr3c1. The offspring of F1 that were administered antibiotics before being weaned spent less time in the EPM open arm. The alterations were accompanied by increased levels of corticosterone in the bloodstream. The gene expression study revealed a decrease in the levels of mRNA transcription of Nr3c1. This research emphasizes the possible long-term effects of antibiotic exposure before weaning on the development of anxiety in offspring upon adulthood.

Early life stress exacerbates the obesogenic and anxiogenic effects of a Western diet without worsening cardiac ischaemic tolerance in male mice.

Early life stress (ELS) and a Western diet (WD) promote mood and cardiovascular disorders, however, how these risks interact in disease pathogenesis is unclear. We assessed effects of ELS with or without a subsequent WD on behaviour, cardiometabolic risk factors, and cardiac function/ischaemic tolerance in male mice. Fifty-six new-born male C57BL/6J mice were randomly allocated to a control group (CON) undisturbed before weaning, or to maternal separation (3h/day) and early (postnatal day 17) weaning (MSEW). Mice consumed standard rodent chow (CON, n = 14; MSEW, n = 15) or WD chow (WD, n = 19; MSEW + WD, n = 19) from week 8 to 24. Fasted blood was sampled and open field test and elevated plus maze (EPM) tests undertaken at 7, 15, and 23 weeks of age, with hearts excised at 24 weeks for Langendorff perfusion (evaluating pre- and post-ischaemic function). MSEW alone transiently increased open field activity at 7 weeks; body weight and serum triglycerides at 4 and 7 weeks, respectively; and final blood glucose levels and insulin resistance at 23 weeks. WD increased insulin resistance and body weight gain, the latter potentiated by MSEW. MSEW + WD was anxiogenic, reducing EPM open arm activity vs. WD alone. Although MSEW had modest metabolic effects and did not influence cardiac function or ischaemic tolerance in lean mice, it exacerbated weight gain and anxiogenesis, and improved ischaemic tolerance in WD fed animals. MSEW-induced increases in body weight (obesity) in WD fed animals in the absence of changes in insulin resistance may have protected the hearts of these mice.

Treadmill Exercise Reverses the Adverse Effects of Intermittent Fasting on Behavior and Cortical Spreading Depression in Young Rats.

Intermittent fasting (IF) and physical exercise (PE) have beneficial psychological and physiological effects, improving memory and anxiety-like behavior. However, the impact of this combination on brain electrophysiological patterns is unknown. We aimed to evaluate the behavior and parameters of a brain excitability-related phenomenon named cortical spreading depression (CSD) in young rats (31-87 days of life) submitted to IF and treadmill PE for eight weeks. Sixty-four male and female Wistar rats aged 24 days were randomized into control, IF, PE, and IF+PE groups. Behavioral tests (open field (OF), object recognition, and elevated plus maze (EPM)) were performed, and the CSD propagation features were recorded. IF caused behavioral responses indicative of anxiety (lower number of entries and time spent in the OF center and EPM open arms). IF also reduced the discrimination index for object recognition memory tests and increased the propagation velocity of CSD. PE rats displayed more entries into the OF center and lowered CSD propagation speed. Data suggest that IF worsens anxiety-like behavior and memory and accelerates CSD in young rats. In contrast, PE reverted the unfavorable effects of IF. The brain effects of IF and PE at younger ages are recommended for study.

Chronic kidney disease is associated with cognitive dysfunction and cardiovascular remodeling in aged male and female Sprague‐Dawley rats

AbstractBackgroundCardiovascular diseases (CVD), including increased central arterial stiffness (CAS), accompany chronic kidney disease (CKD) development. An increase in pulse wave velocity (PWV), an index for CAS, is implicated in age‐associated changes in the brain and potentiates the development of CI in CVD and CKD in humans and animal models. The aim of this study was to determine whether inducing CKD accelerates the development of CI and CAS in aged male and female rats.MethodTen months‐old male and female Sprague‐Dawley rats were fed with 0.25% adenine diet to accelerate CKD (n = 10‐12; CKD‐male, CKD‐female) or regular diet (n = 8‐10; CNT‐male, CNT‐female) for 8weeks. Body weight (BW), blood pressure (BP), heart rate (HR), aortic‐PWV (aPWV), behavioral tests, blood urea nitrogen (BUN), creatinine and hematocrit were assessed at the endpoint. The anxiety‐like behavior was tested in open field test (OFT) and elevated plus maze (EPM). Morris water maze (MWM) and cross maze (CM) were used to test spatial memory. The data were analyzed using two‐way ANOVA.ResultCKD development in both sexes was accompanied by kidneys enlargement, increase in BUN and creatinine, and reduction in hematocrit. In CKD‐male the kidney function was more compromised with higher levels of BUN and creatinine vs. CKD‐female (Table 1). CKD‐male had lower BW and SBP vs. CNT‐male (Table 1). CKD induced a reduction in HR, increase in aPWV and aortic weight in both sexes vs. respective CNT‐groups (Figure 1). Both CKD‐male and CKD‐female demonstrated a tendency of increase in path length to find the hidden platform in MWM, e.g., both sexes had numerically impaired spatial memory. CKD‐male spent less time in the center of OFT, a measure of anxiety, and loss of spatial memory, with a reduction of spontaneous alternation task assessed by CM. CKD‐female exhibited higher level of anxiety spending less time in open arm of EPM vs. CNT‐female (Figure 2).ConclusionCKD development in old male and female Sprague‐Dawley rats was accompanied by an increase in CAS, cardiovascular remodeling, and cognitive dysfunction. In CKD, CI affected different cognitive domains in the old males than in the age‐matched females.Supported by NIA/NIH/IRP

Chronic Exposure of 5‐Alpha Reductase Inhibitor in Young Male Rats Induces Not Only Metabolic Disturbance, But Also Depressive‐Like Behaviors as Similar with Obese Condition

AbstractBackgroundFinasteride (FIN), a 5‐alpha reductase inhibitor (5‐ARIs), is one of the drugs approved for the treatment of androgenetic alopecia (AGA) and benign prostatic hyperplasia (BPH). AGA has been found to be associated with obesity. Obesity and AGA themselves often co‐occurs with anxiety and depression. Nevertheless, several studies reported adverse effects of FIN on anxiety and depression, especially in young men. A previous study also reported that FIN caused anxiety and depression in male rats; however, the effects of chronic FIN exposure in young obese condition has not been investigated.MethodTwenty male Wistar rats (8‐week‐old) were divided into two groups (N = 10/group) to feed with either normal diet (ND) or high‐fat diet (HFD) for 18 weeks. At week 13, rats in each dietary group were subdivided into two subgroups to treat with either vehicle as control or FIN (5 mg/kg/day) via oral gavage. At the end of the experimental protocol, the anxiety/depression‐like behaviors were measured using the Elevated‐Plus Maze (EPM) and the Splash Test (ST), while the Open‐Field Test (OFT) was examined for locomotor activity. In addition, blood in each rat was collected for determining the metabolic profiles.ResultHFD‐fed rats developed obesity and hypercholesterolemia, with an increase in systemic oxidative stress (Figure 1A‐C). Anxiety and depressive‐like behaviors were observed in vehicle‐treated HFD‐fed rats, as indicated by decreasing % preference index in the open arm of EPM and grooming time in ST (Figure 1D‐E). FIN treated ND‐fed rats also showed depression‐like behaviors, similar to those in vehicle‐treated HFD‐fed rats (Figure 1E). Although FIN increased plasma cholesterol levels in HFD‐fed rats, it did not aggravate anxiety and depression‐like behaviors in HFD‐fed rats. Regarding the locomotor activity, no significant difference was found among all groups. (Figure 1F).ConclusionAll of these findings suggest that FIN exposure induced metabolic disturbance, and depression‐like behaviors as similar to obesity without the aggravating effects of FIN and obesity on depressive‐like behaviors.

Chronic Exposure with 5‐Alpha Reductase Inhibitor Ameliorates Anxiety and Depression‐like Behaviors By Reducing Systemic Oxidative Stress in D‐galactose‐Induced Aging Male Rats

AbstractBackgroundThe most common geriatric psychiatric disorders are anxiety and depression. The 5‐alpha reductase inhibitor (5‐ARIs), finasteride (Fin), is a standard medication used to treat benign prostatic hyperplasia (BPH), that often occurs in the elderly. However, the effects of Fin on mental health in aged population are still unclear. D‐galactose (D‐gal)‐induced aging has been recognized worldwide, as a model to mimic natural aging in rodents. Therefore, we investigated the effect of Fin on anxiety and depression‐like behaviors in D‐gal‐induced aging male rats.MethodTwenty‐male Wistar rats (8‐week‐old) were randomly divided into two groups (n = 10/group) to receive either vehicle as control or D‐gal (150 mg/kg/day via subcutaneous injection) for 18 weeks. At week 13, rats in each group were divided into two subgroups (n = 5/subgroup) in order to receive either vehicle (drinking water) or Fin (5 mg/kg/day) via oral gavage. At the end of the treatment, elevated‐plus maze (EPM), and splash test (ST) were performed for quantifying anxiety and depression‐like behaviors, while open‐field test (OFT) to determine locomotor activity. In addition, blood in each rat was collected for determining the metabolic profiles.ResultD‐gal‐treated rat showed no effect on metabolic disturbance, but increased systemic oxidative stress (Figure 1A‐C). Anxiety and depression‐like behavior were observed in D‐gal‐treated rats, as indicated by a decrease % preference index in the open arm of EPM and grooming time in ST (Figure 1D‐E). Fin‐treated control rats demonstrated hypercholesterolemia (Figure 1A‐B), and depression‐like behavior was observed (Figure 1E). Anxiety and depression‐like behaviors were attenuated in Fin‐treated D‐gal rats (Figure 1D‐E). Regarding the locomotor activity, no significant difference was found among all groups. (Figure 1F).ConclusionFIN exposure induced metabolic disturbance, and depression‐like behaviors in male rats. In contrast, Fin ameliorated anxiety and depression‐like behaviors in D‐gal‐induced aging rat, which may due to a reducing systemic oxidative stress. The findings in this study provide information for future clinical applications such as the use of Fin for therapeutic approach for anxiety/depression‐like behaviors in the elderly. However, caution is advised when using Fin in young men since it may induce depressive‐like behaviors.

Experimental evaluation of anxiolytic and antidepressant activities of methanolic extract of Triticum aestivum (wheatgrass) in albino mice

Background: Anxiety and depression are common psychiatric conditions. The present study was carried out to find antianxiety and antidepressant activity of methanolic extract of Triticum aestivum (wheat grass) in mice. Materials and methods: The methanolic extract of Triticum aestivum (META) was screened for antianxiety activity by elevated plus maze (EPM) and light and dark box (LDB) and for antidepressant activity by forced swim test (FST) and tail suspension test (TST) in mice. Animals were divided into four groups having six animals in each group. Group I served as control and received gum acacia aqueous suspension 10 ml/kg, Groups II and III served as test groups and received META 200 and 400 mg/kg, respectively, Group IV served as standard group and received diazepam 1 mg/kg for antianxiety activity and fluoxetine (20 mg/kg) for antidepressant activity once daily for thirty days. Result: META 200 and 400 mg/kg showed significant (P < 0.01) dose-dependent increase in entries and stay in the open arms in EPM and entries and stay in the light compartment in LDB as compared to control. Antianxiety effect of META at dose of 400 mg/kg was comparable (P > 0.05) with diazepam 1mg/kg. META 200 and 400 mg/kg also produced dose-dependent significant (P < 0.01) antidepressant effect, indicated by reduction in the immobility time as compared to control in both FST and TST. The antidepressant activity of META at dose of 400 mg/kg was comparable (P > 0.05) with fluoxetine 20 mg/kg. Conclusion: Results of our study suggested that META possess dose-dependent significant antianxiety and antidepressant activities.

Investigation of effects of transferrin-conjugated gold nanoparticles on hippocampal neuronal activity and anxiety behavior in mice.

Gold nanoparticles (GNPs) have been widely used in medicine such as imaging, drug delivery and therapeutics due to their multifunctional properties. Alterations in neuronal function may contribute to various neurological diseases. Transferrin plays a primary role in iron transportation and delivery and has recently been utilized for drug delivery to the brain. We have investigated effects of transferrin-conjugated GNPs (Tf-GNPs) on anxiety and locomotor behavior in vivo and also hippocampal neuronal activity ex vivo. Electrophysiological effects of Tf-GNP on hippocampal neurons were determined by patch clamp method. Fifteen male young adult C57BL/6 mice were randomly divided into three groups as control (200 µL PBS), GNP (bare GNP; 2.2μg/g in PBS) and Tf-GNPs (2.2μg/g Tf-GNP). Animals intraperitoneally received the respective treatments for seven consecutive days and were subjected to elevated plus maze (EPM) and open field tests (OFT). Ex vivo, firing frequency of the neurons significantly increased by GNP treatment (p < 0.001). In vivo, animals in Tf-GNP group showed significantly longer distance in open arms but significantly lower number of entries to the open arms in EPM (p < 0.05). Mice received bare GNPs had significantly higher locomotor activity in OFT (p < 0.05), while Tf-GNP did not alter the locomotor activity significantly (p = 0.051). Animals in Tf-GNP group spent significantly longer time in the peripheral zone in OFT (p < 0.05). The present findings have shown that Tf-GNP induces anxiety-like behavior without altering spontaneous firing rate of hippocampal neurons. We suggest that neurobiological effects of Tf-GNP should be pre-determined before using in medical applications.

Neurotoxicity of Bisphenol A and the Impact of Melatonin Administration on Oxidative Stress, ERK/NF-kB Signaling Pathway, and Behavior in Rats.

Bisphenol A (BPA) exposure can be associated with neurodevelopmental disorders due to impairment of cell proliferation and synaptic development. Our study evaluated the effects of melatonin (MEL) on ambulatory activity, lipid peroxidation, cytokines, ERK/NF-kB signaling pathway in the hippocampus and frontal lobe, and histopathological changes in the hippocampus of the BPA-treated rats. The animals were divided into 4 groups: control, BPA, BPA + MEL I, and BPA + MEL II. MEL I (20mg/kg b.w.) and MEL II (40mg/kg b.w.) were orally administered for 28days. On the 29th day, BPA (1mg/kg b.w.) was intraperitoneally administered, and, after 24h, an open field test (OFT) and an elevated plus maze (EPM) were conducted. The results showed that the MEL II group made significantly more entries in the open arms of EPM, traveled significantly greater distance, and spent more time in the central part of OFT. Malondialdehyde levels were diminished by MEL II in the hippocampus and by MEL I in the frontal lobe. In the hippocampus, the MAPK level was significantly lowered by both doses of MEL (p < 0.05) while in the frontal lobe, only MEL II reduced the MAPK activation. MEL I and II significantly decreased the γH2AX and upregulated the NFkB and pNFkB expressions in the hippocampus while MEL II downregulated the MCP1 expression. Both doses of MEL attenuated the BPA-evoked histopathological alterations in the hippocampus. These data indicate that MEL can mediate the neuroprotection against BPA-induced neurotoxicity and improves behavioral changes suggesting a real potential as a protective agent in brain toxicity.

Evaluation of the posterior insular cortex involvement in anxiogenic response to emotional stress in male rats: Functional topography along the rostrocaudal axis

The insular cortex (IC) is engaged in behavioral and physiological responses to emotional stress. Control of physiological functions and behavioral responses has been reported to occur in a site-specific manner along the rostrocaudal axis of the IC. However, a functional topography of the IC regulation of anxiogenic responses caused by stress has never been evaluated. Therefore, we investigated the role of rostrocaudal subregions in the posterior IC in anxiogenic-like effect caused by exposure to acute restraint stress in male rats. For this, rats received bilateral microinjection of the non-selective synaptic inhibitor CoCl2 or vehicle into either the rostral, intermediate or caudal portions of the posterior IC before exposure to acute restraint stress. Then, behavior in the elevated plus maze (EPM) was evaluated immediately after restraint stress. The behavior of non-stressed animals in the EPM was also investigated. We observed that acute restraint stress decreased the exploration of the EPM open arms in animals treated with vehicle in all regions of the posterior IC, thus indicating an anxiogenic-like effect. The avoidance of the EPM open arms was completely inhibited in animals subjected to microinjection of CoCl2 into the intermediate posterior IC. Nevertheless, the same pharmacological treatment into either the rostral or caudal subregions of the posterior IC did not affect the restraint-evoked behavioral changes in the EPM. Taken together, these results suggest that regulation of anxiogenic-like effect to emotional stress along the rostrocaudal axis of the posterior IC might occur in a site-specific manner, indicating a role of the intermediate subregion.

Stress resilience-associated behaviors following predator scent stress are accompanied by upregulated nucleus accumbens mGlu5 transcription in female Sprague Dawley rats

Despite the higher prevalence of post-traumatic stress disorder (PTSD) in women, the majority of preclinical research has been conducted utilizing male subjects. We have found that male rats exposed to the predator scent 2,4,5-trimethyl-3-thiazoline (TMT) show heterogenous long-term anxiety-like behavior and conditioned fear to the TMT environment. Stress-Resilient males exhibit increased mGlu5 mRNA expression in the basolateral amygdala (BLA) and prefrontal cortex (PFC). Here we sought to determine whether the same behavioral and genetic responses would be observed in female rats exposed to TMT. Female Sprague-Dawley rats were exposed to TMT for ten minutes, while Controls were exposed to an unscented environment. Anxiety and anhedonia were assessed 7–14 days later with elevated plus maze (EPM), acoustic startle response, light-dark box, and sucrose preference test (SPT). TMT-exposed females spent less time in the EPM open arms, exhibited greater startle amplitude, and reduced sucrose intake compared to Controls. Median split analyses conducted on EPM and SPT data yielded stress-Susceptible and -Resilient phenotypes that displayed behavior in the light-dark box consistent with EPM and SPT behavior. Susceptible females displayed greater BLA mGlu5 mRNA expression than Resilient and Control rats and did not show conditioned fear, in contrast to previous results in males. Resilient females displayed greater mGlu5 mRNA expression in the nucleus accumbens. These data indicate that the predator scent stress model of PTSD produces distinct stress-Susceptible and Resilient phenotypes in female rats that are associated with changes in mGlu5 mRNA expression in several brain regions.

LOSARTAN, ANGIOTENSIN II TYPE 1 RECEPTOR BLOCKER, REDUCED AORTIC STIFFNESS AND IMPROVED COGNITIVE FUNCTION IN DAHL SALT SENSITIVE RAT MODEL OF AGE-ASSOCIATED VASCULAR DEMENTIA

Objective: Cardiovascular diseases are the major contributors to the pathogenesis of vascular dementia. The Dahl salt sensitive (Dahl-S) rat model is characterized by the development of central arterial stiffness (CAS), hypertension and cognitive decline with age on a normal salt diet. We hypothesized that treatment with losartan (LOS), angiotensin II type 1 receptor (AT1R) blocker, via reduction of blood pressure (BP) and CAS, will improve cognitive function in aged Dahl-S rats. Design and method: Male Dahl-S rats (n = 30) were kept on a normal salt diet (0.5% NaCl) for the duration of the study. Baseline measurements were taken at 3- and/or 6-mo of age following by the treatment with LOS in drinking water (30 mg/kg/day, n = 14) or control treatment (n = 16) for 6-mo. Measurements were taken every 3 months and included systolic and diastolic BP (SBP, DBP), pulse wave velocity (PWV), a measure of CAS, left ventricular posterior wall thickness in diastole (LVPWd), fractional shortening (FS) and LV mass (by echocardiography). At 12-mo of age the rats were tested in operant chambers using a simple reaction task to assess attention and impulsivity, and in elevated plus maze (EPM) to assess anxiety-like behavior. Statistical analyses were performed using 2-way ANOVA and t-test. Data are presented as mean ± SEM. Results: Prior to the treatment, LOS and control groups did not display differences in any parameter analyzed. Following the treatment, LOS rats had lower SBP, DBP, PWV, thinner LVPWd, higher FS and smaller LV mass at 9- and 12-mo of age vs. control group (Fig.1A-F). In the reaction task, LOS rats demonstrated more correct responses and fewer premature responses. The LOS rats exhibited lower anxiety level, because they spent more time in the open arm of EPM vs. control rats (Fig.1G,H). Conclusions: Beneficial effect of LOS on cognitive function in aged male Dahl-S rats was associated with the improvement of cardiovascular function and remodeling. The CAS stabilizing and BP reduction by LOS treatment was associated with improved attentional performance, lower impulsivity and lower anxiety. The mechanistic basis of these effects of LOS on cognition via cerebrovascular and brain changes will be further investigated.

STREStrogen: The Dichotomous Role of Estrous Cycle on Anxiety‐like Behavior is Dependent upon Stress History

Stress‐induced psychiatric disorders are some of the most debilitating conditions worldwide and occur more often in women than men. Increased risk coincides with the reproductive years when circulating estrogen (E) levels peak, alluding to a critical role of ovarian hormones in susceptibility. Human stress studies support this association as E is linked to negative mood and anxiety during social stressors. However, preclinical research often reports opposing findings of protective effects of E in non‐stressed subjects, indicating reduced anxiety‐like behavioral responses. Yet, rodent studies often neglect to acknowledge the latent effects of stress on E‐induced behavioral differences, like the negative impact of a traumatic experience that perpetuates future interactions. Thus, this divergence between clinical and preclinical findings limits the capability of identifying adequate pharmacological targets for use in naturally cycling females. Therefore, we investigated if prior exposure to stress in intact adult female rats sensitized behavioral outcomes during anxiogenic tasks and whether this was contingent upon the current phase of estrous. Rats were exposed for 15 mins/day over 5 consecutive days to either control handling (CON) or witness stress (WS), a modified social defeat paradigm whereby a female observes social defeat from within a protected compartment of the aggressive resident’s cage. Forty‐eight hours after the 5th and final CON/WS exposure, rats underwent elevated plus maze (EPM) testing, and then acoustic startle response (ASR) testing the next day. Vaginal lavage was obtained immediately following EPM and ASR to confirm estrous phase. Analyses were conducted in a 2 x 2 fashion: stress exposure x estrous phase, in which metestrus/diestrus (M/D) signified putatively low levels of E, and proestrus/estrus (P/E) signified high levels. Results revealed a prominent stress exposure x estrous phase interaction for percentage of time spent in the closed and open arms of EPM. First, CON rats demonstrated reduced avoidance of the open arms when tested during P/E vs M/D. This is in line with studies reporting an anxiolytic effect of E. However, our findings in WS rats supports the notion that E is a critical component of sensitizing anxiety‐like responses in females with a history of stress. Specifically, WS rats demonstrated greater avoidance of open arms in the EPM during P/E vs M/D. Importantly, we also found that not all tests are impacted by cycle stage. Females with a history of stress demonstrated a sensitized startle amplitude compared with CON rats, independent of estrous phase, when tested for magnitude of the innate startle response. Further investigations into E receptor expression and subsequent signaling mechanisms in stress‐sensitive brain regions are necessary to determine how a history of stress modifies behavioral outcomes in the presence of high circulating E. These data can then elucidate novel therapeutic targets important for decreasing the high incidence of psychological disorders in this susceptible population.

Swimming Exercise Modulates Gut Microbiota in CUMS-Induced Depressed Mice.

BackgroundGut microbiota is associated with anxiety and depression, while exercise has been proved to alleviate depressive symptoms. However, the interaction of exercise, depression, and gut microbiota remains unclear.MethodsMale C57/BL6J mice were exposed to chronic unpredictable mild stress (CUMS) for 6 weeks and then were subjected to a 5-week swimming program. Behavioral tests, including sucrose preference test (SPT), open field test (OFT), elevated plus-maze (EPM) test, and tail suspension test (TST), were conducted to assess the anxiety-like and depressive behaviors. Gut microbiota analysis was carried out after sample collection.ResultsThis study showed that CUMS induced depressive behaviors, but swimming exercise increased sucrose preference rate in the SPT, increased time in the center and number of rearing in the OFT, decreased time in the closed arm and increased time in the open arm in EPM, and decreased immobility time in the TST. Firmicutes were the predominant phylum in the gut microbiome, followed by the phyla Bacteroidetes and Proteobacteria. We further found that CUMS and swimming influenced the relative abundance of the genus Desulfovibrio, genus Streptococcus, genus p-75-a5. Among the metabolic pathways, aromatic biogenic amine degradation (PWY-7431), mono-trans and polycis decaprenyl phosphate biosynthesis (PWY-6383), chlorosalicylate degradation (PWY-6107), mycothiol biosynthesis (PWY1G-0), mycolyl-arabinogalactan-peptidoglycan complex biosynthesis (PWY-6397), toluene degradation I (aerobic) (via o-cresol) (PWY-5180), toluene degradation II (aerobic) (via 4-methylcatechol) (PWY-5182), and starch degradation III (PWY-6731) may be related to the mechanism of anti-depression effect.ConclusionSwimming exercise reverses CUMS-induced depressive behaviors, and the alteration of gut microbiota composition and regulation of microbiota metabolic pathways are involved.