Valid Animal Model Research Articles

Artificial Neural Network to Predict Varicocele Impact on Male Fertility through Testicular Endocannabinoid Gene Expression Profiles.

The relationship between varicocele and fertility has always been a matter of debate because of the absence of predictive clinical indicators or molecular markers able to define the severity of this disease. Even though accumulated evidence demonstrated that the endocannabinoid system (ECS) plays a central role in male reproductive biology, particularly in the testicular compartment, to date no data point to a role for ECS in the etiopathogenesis of varicocele. Therefore, the present research has been designed to investigate the relationship between testicular ECS gene expression and fertility, using a validated animal model of experimental varicocele (VAR), taking advantage of traditional statistical approaches and artificial neural network (ANN). Experimental induction of VAR led to a clear reduction of spermatogenesis in left testes ranging from a mild (Johnsen score 7: 21%) to a severe (Johnsen score 4: 58%) damage of the germinal epithelium. However, the mean number of new-borns recorded after two sequential matings was quite variable and independent of the Johnsen score. While the gene expression of biosynthetic and degrading enzymes of AEA (NAPE-PLD and FAAH, respectively) and of 2-AG (DAGLα and MAGL, respectively), as well as their binding cannabinoid receptors (CB1 and CB2), did not change between testes and among groups, a significant downregulation of vanilloid (TRPV1) expression was recorded in left testes of VAR rats and positively correlated with animal fertility. Interestingly, an ANN trained by inserting the left and right testicular ECS gene expression profiles (inputs) was able to predict varicocele impact on male fertility in terms of mean number of new-borns delivered (outputs), with a very high accuracy (average prediction error of 1%). The present study provides unprecedented information on testicular ECS gene expression patterns during varicocele, by developing a freely available predictive ANN model that may open new perspectives in the diagnosis of varicocele-associated infertility.

Liver Injury, Endotoxemia, and Their Relationship to Intestinal Microbiota Composition in Alcohol-Preferring Rats.

There is strong evidence that alcoholism leads to dysbiosis in both humans and animals. However, it is unclear how changes in the intestinal microbiota (IM) relate to ethanol (EtOH)-induced disruption of gut-liver homeostasis. We investigated this issue using selectively bred Sardinian alcohol-preferring (sP) rats, a validated animal model of excessive EtOH consumption. Independent groups of male adult sP rats were exposed to the standard, home-cage 2-bottle "EtOH (10% v/v) versus water" choice regimen with unlimited access for 24h/d (Group Et) for 3 (T1), 6 (T2), and 12 (T3) consecutive months. Control groups (Group Ct) were composed of matched-age EtOH-naïve sP rats. We obtained samples from each rat at the end of each experimental time, and we used blood and colon tissues for intestinal barrier integrity and/or liver pathology assessments and used stool samples for IM analysis with 16S ribosomal RNA gene sequencing. Rats in Group Et developed hepatic steatosis and elevated serum transaminases and endotoxin/lipopolysaccharide (LPS) levels but no other liver pathological changes (i.e., necrosis/inflammation) or systemic inflammation. While we did not find any apparent alteration of the intestinal colonic mucosa, we found that rats in Group Et exhibited significant changes in IM composition compared to the rats in Group Ct. These changes were sustained throughout T1, T2, and T3. In particular, Ruminococcus, Coprococcus, and Streptococcus were the differentially abundant microbial genera at T3. The KEGG Ortholog profile revealed that IM functional modules, such as biosynthesis, transport, and export of LPS, were also enriched in Group Et rats at T3. We showed that chronic, voluntary EtOH consumption induced liver injury and endotoxemia together with dysbiotic changes in sP rats. This work sets the stage for improving our knowledge of the prevention and treatment of EtOH-related diseases.

Effect of Exercise Intervention on Cardiac Function in Type 2 Diabetes Mellitus: A Systematic Review.

The effect of exercise on cardiac function/structure in type 2 diabetes mellitus (T2DM) with or without diabetic cardiomyopathy (DCM) is not yet completely understood. To date, results of studies have been controversial with variable outcomes due to the variety of exercise modalities. The aim of the present review was to examine the impact of exercise intervention, and different types of exercise, on cardiac function and structure in T2DM through a systematic literature review, combining both pre-clinical and clinical studies. A systematic literature search was performed on PubMed, Web of Science, and PEDro to identify studies up to 2 April 2018. Articles were included when well-defined exercise protocols were provided, and cardiac function in T2DM patients or validated animal models was examined. In diabetic animals, improvements in both diastolic and systolic function through exercise therapy were mainly attributed to reduced collagen deposition. In T2DM patients, improvements were observed in diastolic function, but not consistently in systolic function, after endurance (and combined resistance) exercise training. Different exercise intervention modalities and exercise types seemed equally effective in improving cardiac structure and function. Exercise training elicits significant improvements in diastolic function and beneficial remodeling in T2DM and DCM animal models, but not necessarily improvements in systolic function and left ventricular structure, regardless of exercise type. Therefore, exercise intervention should be a cornerstone in the treatment of T2DM patients not only to improve glycemic control but also to specifically enhance cardiac function.

Stress-Enhanced Fear Learning, a Robust Rodent Model of Post-Traumatic Stress Disorder

Fear behaviors are important for survival, but disproportionately high levels of fear can increase the vulnerability for developing psychiatric disorders such as post-traumatic stress disorder (PTSD). To understand the biological mechanisms of fear dysregulation in PTSD, it is important to start with a valid animal model of the disorder. This protocol describes the methodology required to conduct stress-enhanced fear learning (SEFL) experiments, a preclinical model of PTSD, in both rats and mice. SEFL was developed to recapitulate critical aspects of PTSD, including long-term sensitization of fear learning caused by an acute stressor. SEFL uses aspects of Pavlovian fear conditioning but produces a distinct and robust sensitized fear response far greater than normal conditional fear responses. The trauma procedure involves placing a rodent in a conditioning chamber and administering 15 unsignaled shocks randomly distributed over 90 minutes (for rat experiments; for mouse experiments, 10 unsignaled shocks randomly distributed over 60 minutes are used). On day 2, rodents are placed in a novel conditioning context where they receive a single shock; then, on day 3 they are placed back in the same context as on day 2 and tested for changes in freezing levels. Rodents that previously received the trauma display enhanced levels of freezing on the test day compared to those that received no shocks on the first day. Thus, with this model, a single highly stressful experience (the trauma) produces extreme fear of the stimuli associated with the traumatic event.

The Impact of Chronic Pelvic Ischemia on LUTS and Urinary Levels of Neuroinflammatory, Inflammatory, and Oxidative Stress Markers in Elderly Men: A Case-control Study

ObjectiveTo investigate lower urinary tract symptoms (LUTS) and urinary levels of neuroinflammatory, inflammatory, and oxidative stress markers in elderly men with chronic pelvic ischemia (CPI) caused by significant aortoiliac disease. Materials and MethodsThirteen men aged over 60 years, with aorta, unilateral or bilateral common/internal iliac artery occlusion documented by computed tomography angiography or angiography, were enrolled from the vascular surgery department. Twelve sex- and age-matched controls without significant aortoiliac disease were used for comparison. Exclusion criteria included neurogenic bladder dysfunction, bladder or prostate cancer, prostatic surgery, pelvic radiotherapy, or chronic treatment for LUTS. Participants underwent urological examination, including assessment of International Prostate Symptom Score (IPSS), uroflowmetry, postvoid residual (PVR), and prostate volume. Urine samples were collected, and levels of neuroinflammatory (nerve growth factor, NGF), inflammatory (cytokines), and oxidative stress markers (8-hydroxy-2′-deoxyguanosine) were determined by enzyme-linked immunosorbent assay. ResultsGroups were similar for age, PVR, prostate volume, and most cardiovascular risk factors. IPSS was higher in patients with CPI (11 ± 3 vs 8 ± 2, P = .02), with a significant mean difference between groups of three points. Urinary NGF was significantly higher in men with CPI (3.7 ± 0.8 vs 2.9 ± 0.7, P = .02), but no differences were found in inflammatory and oxidative biomarkers among groups. ConclusionSevere CPI in elderly men is associated with a significant increase in LUTS and bladder neurogenic inflammation, as suggested by the increase of NGF release in urine, sensitizing bladder afferents. These findings confirm the relevance of ischemia in bladder function and appear to validate animal models of bilateral iliac artery occlusion.

ITGB4 deficiency in bronchial epithelial cells directs airway inflammation and bipolar disorder-related behavior

BackgroundChronic persistent airway inflammation has been associated with the comorbidity of asthma and bipolar disorder (BD). However, the direct relevance between airway inflammation and BD-like psychiatric comorbidity is almost unknown. Integrin β4 (ITGB4) is downregulated on the airway epithelial of asthma patients, which might play a critical role in the parthenogenesis of airway inflammation. So this study aimed to examine the role of ITGB4 deficiency in mediating airway inflammation and further leading to the BD-like behaviors.MethodsITGB4−/− mice were generated by mating ITGB4fl/fl mice with CCSP–rtTAtg/−/TetO-Cretg/tg mice. Mania-like behavior tests were performed, including hyperlocomotion, d-amphetamine-induced hyperactivity, open-field test, and elevated plus-maze test. Depressive-like behavior tests were carried out, including sucrose preference, forced swimming, and learned helplessness. Inflammatory cells (Th17, Th1, Th2) in the lung were examined by flow cytometry. Futhermore, inflammatory cytokines (IL-4, IL-13) in bronchoalveolar lavage fluid and sera were detected by ELISA. Protein expression of the IL-4Rα on choroid plexus, microglial marker (IBA1), and synapse-associated proteins (synaptophysin, SYP) in the hippocampus and prefrontal cortex were examined by western blotting. Additionally, proinflammatory cytokines (IL-1β, IL-6, and TNF-α) in the hippocampus and prefrontal cortex were detected by immunohistochemistry. Inflammatory disorder in the lung, hippocampus, and prefrontal cortex was tested by hematoxylin and eosin (H&E) staining. And cell apoptosis in the hippocampus and prefrontal cortex was measured by TUNEL test.ResultsITGB4−/− mice exhibited mania-like behavior, including hyperlocomotion, d-amphetamine-induced hyperactivity, and reduced anxiety-like behavior. While under stressful conditions, ITGB4−/− mice manifested depressive-like behavior, including anhedonia, behavioral despair, and enhanced learned helplessness. At the same time, ITGB4−/− mice mainly exerted Th2-type inflammation in periphery, like the number and major cytokines IL-4 and IL-13 of Th2-type inflammation. ITGB4−/− mice also showed a significant increase of microglia and pro-inflammatory cytokines such as IL-1β, IL-6, and TNF-α in the hippocampus and prefrontal cortex. Additionally, neuron damage, increased neuron apoptosis, and the decrease of SYP were found in ITGB4−/− mice.ConclusionsThese findings confirmed that airway inflammatory induced by ITGB4 deficiency is the important incentive for the BD-like behavior during asthma pathogenesis. The ITGB4-deficient mice provide a validated animal model for us to study the possible mechanism of BD-like psychiatric comorbidity of asthma patients.

Histological and molecular techniques utilized to investigate animal models of depression. An updated review.

This review aimed to summarize the different histopathological techniques and procedures utilized during investigating the different animal models of depression in order to explore the pathophysiological aspect of depression and testing the efficacy of the antidepressant drugs or new treatments. This will be helpful while designing researches aiming to achieve these objectives. It was found that the major obstacle during investigating the animal models of depression was the restricted availability of validated animal models. The chronic stress models have face, construct, and predictive validity. It was found that the histological techniques used in investigating the animal models of depression that was described in the literatures fall under three categories; the light microscopic, the electron microscopic and the molecular biological studies. The light microscope studies were performed using the routine histological staining and immunohistochemical technique that aimed to describe the hippocampal histopathological changes induced by depression. Establishment of a preclinical behavioral science laboratory is highly recommended. It will encourage and support the conduction of high quality, multidisciplinary researches targeting anxiety and other psychiatric disorders and will indirectly improve the health care provided to the psychiatric patients. RESEARCH HIGHLIGHTS: Chronic stress models are valid ones. Light microscope was utilized to examine the routinely or immunohistochemically stained sections in hippocampus of animal models of depression while electron microscope was utilized to examine its ultrastructure.

Cardiovascular consequences of KATP overactivity in Cantu syndrome.

Cantu syndrome (CS) is characterized by multiple vascular and cardiac abnormalities including vascular dilation and tortuosity, systemic hypotension, and cardiomegaly. The disorder is caused by gain-of-function (GOF) mutations in genes encoding pore-forming (Kir6.1, KCNJ8) and accessory (SUR2, ABCC9) ATP-sensitive potassium (KATP) channel subunits. However, there is little understanding of the link between molecular dysfunction and the complex pathophysiology observed, and there is no known treatment, in large part due to the lack of appropriate preclinical disease models in which to test therapies. Notably, expression of Kir6.1 and SUR2 does not fully overlap, and the relative contribution of KATP GOF in various cardiovascular tissues remains to be elucidated. To investigate pathophysiologic mechanisms in CS we have used CRISPR/Cas9 engineering to introduce CS-associated SUR2[A478V] and Kir6.1[V65M] mutations to the equivalent endogenous loci in mice. Mirroring human CS, both of these animals exhibit low systemic blood pressure and dilated, compliant blood vessels, as well dramatic cardiac enlargement, the effects being more severe in V65M animals than in A478V animals. In both animals, whole-cell patch-clamp recordings reveal enhanced basal KATP conductance in vascular smooth muscle, explaining vasodilation and lower blood pressure, and demonstrating a cardinal role for smooth muscle KATP dysfunction in CS etiology. Echocardiography confirms in situ cardiac enlargement and increased cardiac output in both animals. Patch-clamp recordings reveal reduced ATP sensitivity of ventricular myocyte KATP channels in A478V, but normal ATP sensitivity in V65M, suggesting that cardiac remodeling occurs secondary to KATP overactivity outside of the heart. These SUR2[A478V] and Kir6.1[V65M] animals thus reiterate the key cardiovascular features seen in human CS. They establish the molecular basis of the pathophysiological consequences of reduced smooth muscle excitability resulting from SUR2/Kir6.1-dependent KATP GOF, and provide a validated animal model in which to examine potential therapeutic approaches to treating CS.

Chronic Social Stress Leads to Reduced Gustatory Reward Salience and Effort Valuation in Mice.

Pathology of reward processing is a major clinical feature of stress-related neuropsychiatric disorders including depression. Several dimensions of reward processing can be impacted, including reward valuation/salience, learning, expectancy and effort valuation. To establish the causal relationships between stress, brain changes, and reward processing pathologies, valid animal models are essential. Here, we present mouse experiments investigating behavioral effects of chronic social stress (CSS) in association learning tests of gustatory reward salience and effort valuation. The reward salience test (RST) comprised Pavlovian pairing of a tone with gustatory reward. The effort valuation test (EVT) comprised operant responding for gustatory reinforcement on a progressive ratio schedule (PRS). All testing was conducted with mice at 100% baseline body weight (BBW). In one experiment, mice underwent 15-day CSS or control handling (CON) and testing was conducted using sucrose pellets. In the RST on days 16–17, CSS mice made fewer feeder responses and had a longer tone response latency, than CON mice. In a shallow EVT on days 19–20, CSS mice attained a lower final ratio than CON mice. In a second CSS experiment, mice underwent CSS or CON and testing was conducted with chocolate pellets and in the presence of standard diet (low effort/low reward). In the RST on days 16–18, CSS mice made fewer feeder responses and had a longer tone response latency, than CON mice. In a steep EVT on days 19–20, CSS and CON mice attained less pellets than in the RST, and CSS mice attained a lower final ratio than CON mice. At day 21, blood levels of glucose and the satiety adipokine leptin were similar in CSS and CON mice. Therefore, CSS leads to consistent reductions in reward salience and effort valuation in tests based on association learning. These reward pathology models are being applied to identify the underlying neurobiology and putative molecular targets for therapeutic pharmacology.

Biological and Behavioral Patterns of Post-Stroke Depression in Rats.

Post-stroke depression (PSD) is the most frequent psychiatric complication following ischemic stroke. It affects up to 60% of all patients and is associated with increased morbidity and mortality following ischemic stroke. The pathophysiology of PSD remains elusive and appears to be multifactorial, rather than "purely" biological or psychosocial in origin. Thus, valid animal models of PSD would contribute to the study of the etiology (and treatment) of this disorder. The present study depicts a rat model for PSD, using middle cerebral artery occlusion (MCAO). The two-way shuttle avoidance task, Porsolt forced-swim test, and sucrose preference test were employed to assess any depression-like behavior. Localized brain expressions of brain-derived neurotrophic factor (BDNF) protein levels were evaluated to examine the possible involvement of the brain neuronal plasticity in the observed behavioral syndrome. The raw data were subjected to unsupervised fuzzy clustering (UFC) algorithms to assess the sensitivity of bio-behavioral measures indicative of depressive symptoms post MCAO. About 56% of the rats developed significant depressive-like behavioral disruptions as a result of MCAO compared with 4% in the sham-operated control rats. A pattern of a depressive-like behavioral response was common to all affected MCAO animals, characterized by significantly more escape failures and reduced number of total avoidance shuttles, a significant elevation in immobility duration, and reduced sucrose preference. Significant downregulations of BDNF protein levels in the hippocampal sub-regions, frontal cortex, and hypothalamus were observed in all affected MCAO animals. The UFC analysis supports the behavioral analysis and thus, lends validity to our results.

Development of a Porcine Full-Thickness Burn Hypertrophic Scar Model and Investigation of the Effects of Shikonin on Hypertrophic Scar Remediation.

Hypertrophic scars formed after burns remain a challenge in clinical practice. Development of effective scar therapies relies on validated animal models that mimic human hypertrophic scars. A consistent porcine full-thickness burn hypertrophic scar model has yet to be developed. We have previously reported that Shikonin induces apoptosis and reduces collagen production in hypertrophic scar fibroblasts in vitro and may therefore hold potential as a novel scar remediation therapy. In this study, we aimed to validate the potential of Shikonin on scar remediation in vivo. A novel porcine hypertrophic scar model was created after full-thickness burn wounds, and the effect of Shikonin on scar remediation was investigated. Clinical scar assessments, histology, and immunohistochemistry were used to evaluate scar appearance, morphology, and protein expression. Eight weeks after scar formation, clinical scar assessment indicated that the score of hypertrophic scars treated with Shikonin was significantly lower than that of the control group. Hypertrophic scars treated with Shikonin appeared flat, pink, and pliable. In addition, histological analysis indicated that hypertrophic scars treated with Shikonin exhibited reduced thickness of the epidermis and dermis, thin and even epithelial layers, reduced numbers of keratinocytes, uniform distribution of fibroblasts, and a parallel and loose arrangement of collagen fibers in the dermis. Moreover, immunohistochemical analysis indicated that Shikonin inhibited the expression of p63, cytokeratin 10, alpha-smooth muscle actin, transforming growth factor-beta 1, and collagen I, which play important roles in hypertrophic scar formation. Based on these results, we conclude that Shikonin has potential as a novel scar therapy.

Reverse Translation: Using Computational Modeling to Enhance Translational Research.

Animal models provide powerful tools for examining human disease; however, translation of findings from these models to human patients is often challenging. To this end, we discuss modern tools to support the process of selecting and validating animal models with relevance to humans. We draw from data mining and computational modeling approaches to examine how large data sets may be leveraged to identify suitable models with the greatest translational potential. Rodent models for human disease are indispensable tools for modern biomedical research, particularly because of the high degree of control they afford in examination of complex pathophysiological phenomena. There is no doubt within the scientific community that animal models are both useful and necessary; however, the value of any animal model depends on its ability to replicate the human condition. Curing disease in a mouse is fascinating but short on merit without application to the understanding of human disease. Thus, there is great need for a robust process to translate a human disease into an appropriate rodent model, termed reverse translation, to distinguish it from the more familiar act of moving findings from animal models into human patients (translation). Studies on the quality of rodent models have reported disparate results depending on the specific physiological system of study. For example, studies on the acute inflammatory response to blunt trauma, burn, and endotoxin have found a poor correlation between the response in mouse and that in human, leading to the conclusion that murine models have limited translational potential.1 This weak association may be due, in part, to a highly variable response in mice that is not observed in humans, underscoring the low success rate in translating mouse findings on acute inflammatory response to human. However, rodents fare better as a model in chronic inflammation, where there is a much better …

Animal Models of Chronic Migraine.

Chronic migraine (CM) is a recalcitrant subtype of migraine which causes high degrees of disability, poor treatment responses, and frequent recurrences in sufferers. However, the pathophysiological mechanisms underlying the development and chronification of migraine attacks remain incompletely understood. A validated animal model could help to decipher the pathogenic mechanism of the disease, facilitating the development of possible therapeutic strategies for CM. In this review, we aimed to summarize current animal models of CM and discuss the validity of these models. Several methods have been available to induce recurrent headache-like behaviors or biochemical changes in rodents, including repeated dural application of inflammatory soup, chronic systemic infusion of nitroglycerin, repeated administration of acute migraine abortive treatment to simulate medication overuse headache, or genetic modification. These models exhibit some features that are believed to be associated with migraine; however, none of the model can recapitulate all the clinical phenotypes found in humans and each has its own weakness. The complex features of CM increase the difficulty of constructing a proper animal model. Nonetheless, currently available models are valid to certain degrees. Future directions might consider simulating the spontaneity and chronicity of migraine by combining known genetic substrates and allostatic loads into the same model.

Mnemonic and behavioral effects of biperiden, an M1-selective antagonist, in the rat.

There is a persistent pressing need for valid animal models of cognitive and mnemonic disruptions (such as seen in Alzheimer's disease and other dementias) usable for preclinical research. We have set out to test the validity of administration of biperiden, an M1-acetylcholine receptor antagonist with central selectivity, as a potential tool for generating a fast screening model of cognitive impairment, in outbred Wistar rats. We used several variants of the Morris water maze task: (1) reversal learning, to assess cognitive flexibility, with probe trials testing memory retention; (2) delayed matching to position (DMP), to evaluate working memory; and (3) "counter-balanced acquisition," to test for possible anomalies in acquisition learning. We also included a visible platform paradigm to reveal possible sensorimotor and motivational deficits. A significant effect of biperiden on memory acquisition and retention was found in the counter-balanced acquisition and probe trials of the counter-balanced acquisition and reversal tasks. Strikingly, a less pronounced deficit was observed in the DMP. No effects were revealed in the reversal learning task. Based on our results, we do not recommend biperiden as a reliable tool for modeling cognitive impairment.

Assessment of Reamer Irrigator Aspirator System (RIA) filtrate for its osteoinductive potential in a validated animal model

PurposePrevious studies indicate that Reamer Irrigator Aspirator (RIA) filtrate contains proteins that have the potential to stimulate bone healing. This study aimed to determine the osteoinductive capabilities of RIA filtrate in a validated in vivo model. MethodsWith Institutional Review Board approval, RIA filtrates from 9 patients were collected. The filtrate was processed to remove cells and inorganic particles. A portion of each sample was set aside for protein analysis while the remainder was lyophilized and prepared for implantation. With Animal Care and Use Committee approval, athymic mice (n = 16; 32 hind limbs) were randomly assigned to 1 of 4 groups (n = 8 limbs per group) for percutaneous gastrocnemius muscle injection of demineralized bone matrix (DBM) (10 mg), lyophilized RIA powder (10 mg), RIA liquid (10 mg of lyophilized RIA powder in 100ul phosphate buffered saline (PBS)), or DBM (10 mg) + RIA liquid (10 mg in 100ul PBS). Radiographs were obtained 2, 4, and 8 weeks after injection. At 8 weeks, mice were sacrificed and the entire gastrocnemius muscle from each hind limb was collected and processed for histologic examination. Histological sections and radiographs were assessed for ossification/calcification. Data were compared for statistically significant (p < 0.05) differences among groups and strong (R > 0.7) correlations between outcome measures. ResultsThe protein composition of RIA filtrates was consistent among patients and matched previous data. For all groups, radiographic scores were significantly (p < 0.014) higher (more calcification/ossification) at 8 weeks compared to 2 weeks. Radiographic scores for the DBM and DBM + RIA liquid groups were significantly higher than RIA liquid and RIA powder at 4 weeks and 8 weeks (p < 0.019 and p < 0.049, respectively). Histologic scores were significantly (p = 0.004) higher in the DBM + RIA liquid group compared to the RIA liquid group at 8 weeks. Histologic scores showed strong correlations (r > 0.77) to radiographic scores for all groups. ConclusionRIA filtrate liquid and powder were osteoinductive in vivo with new bone formation being most abundant using a combination of DBM and RIA filtrate in this validated animal model. RIA filtrate has potential for clinical use in augmenting bone healing treatments.

Voluntary Exercise During Adolescence Mitigated Negative the Effects of Maternal Separation Stress on the Depressive-Like Behaviors of Adult Male Rats: Role of NMDA Receptors.

Evidence indicates that experiencing early-life stress (ELS) is a risk factor for the development of mental disorders such as depression. Maternal separation stress (MS) is a valid animal model of ELS that caused to induce long-lasting effects on the brain and behaviors of animals. It hypothesized that adolescence is a critical stage in which the brain is still developing, and applying (non)pharmacological therapies in this period may attenuate the effects of ELS on the brain and behavior. Male rats were subjected to MS from postnatal day (PND) 2-14, and the stressed animals were then treated with (1) chronic fluoxetine (FLX) (5mg/kg) and (2) voluntary running wheel exercise (RW) from PND 30, for 30 days. Then, we subjected the animals to behavioral and molecular assessments at PND 60. Our data showed that MS provoked depressive-like behaviors in rats, tested by the forced swimming test, splash test, and sucrose preference test. Additionally, we found that MS increased the gene expression of the NR2A (and not NR2B) subunit of N-methyl-D-aspartate (NMDA) receptors in the hippocampus of adult rats. Both FLX and RW treatments during adolescence were able to mitigate the negative effects of ELS on stressed animals. These results highlighted the importance of adolescence in treating stressed animals with FLX/voluntary RW exercise to alleviate the depressive effects of ELS. In addition, we found that ELS altered the transcriptional level of Grin2a (and not Grin2b) in the hippocampus. Finally, our results showed that FLX/voluntary RW exercise during adolescence could normalize altered expression of Grin2a in the hippocampus of adult rats.

Triglycerides: A Potential Hematological Marker in Animal Models of Cognitive Impairment

Mild Cognitive Impairment (MCI) is a clinical disorder characterized as an intermediate stage of cognitive impairment between normal cognitive aging and more advanced cognitive impairment associated with dementia. MCI causes measurable decline in cognitive abilities including memory and reasoning. Occurrence of MCI has been linked to altered physiological functions, co‐morbid conditions, and adverse events related to therapeutic drugs or toxins. MCI is commonly diagnosed by a physician's clinical judgement based on the following parameters: complete medical history, assessment of independent function and daily activities, input from family members, assessment of mental status (i.e. Mini Mental Status Exam), neurological examinations, and laboratory tests including blood tests and imaging of the brain's structure. The testing required is extensive and often subjective in nature. Currently, there are few biochemical markers thought to correlate with the onset of early‐stage MCI and those that do exist require expensive or invasive techniques to measure accurately. Therefore, using valid animal models representative of cognitive impairment, this study aims to determine the validity of using hematological markers as predictors of early‐stage MCI development.In this study, we measured the hematological markers in animal models of cognitive impairment compared to their respective control group. Cognitive impairment was observed in a chemotherapeutic (CT) rodent group dosed with doxorubicin (2 mg/kg, IP) and cyclophosphamide (50 mg/kg, IP) once weekly for 4 weeks. Cognitive impairment was also observed in a hyperglycemia rodent group dosed with Streptozotocin (STZ) 55 mg/kg, once, IP. Blood samples were withdrawn, and hematological markers analyzed. There was a significant increase in creatine kinase and lactate dehydrogenase (LDH) in the CT model versus control. Glucose levels were significantly increased in the STZ model versus control. With regard to BUN, there was a significant increase in the STZ model compared to their control group, however there was a decrease in the CT model compared to their control group. Interestingly, triglycerides were significantly elevated in both the STZ and CT treated animal models. Previous studies with human subjects have shown a potential link between the increase in triglyceride levels influencing cognitive impairment. Likewise, our data indicates a possible correlation with an increase in triglycerides to cognitive impairment in animal models. This suggests elevated levels of triglycerides may prove to be a potential, minimally invasive, hematological marker for the increased risk of cognitive deficit in these animal models. However, further studies are necessary to determine the causal relationship between elevated triglyceride levels and early‐stage MCI.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Translation-relevant EEG phenotypes in a mouse model of Fragile X Syndrome

Identification of comparable biomarkers in humans and validated animal models will facilitate pre-clinical to clinical therapeutic pipelines to treat neurodevelopmental disorders. Fragile X Syndrome (FXS) is a leading known genetic cause of intellectual disability with symptoms that include increased anxiety, social and sensory processing deficits. Recent EEG studies in humans with FXS have identified neural oscillation deficits that include enhanced resting state gamma power and reduced inter-trial coherence of sound evoked gamma oscillations. To determine if analogous phenotypes are present in an animal model of FXS, we recorded EEGs in awake, freely moving Fmr1 knock out (KO) mice using similar stimuli as in the human studies. We report remarkably similar neural oscillation phenotypes in the Fmr1 KO mouse including enhanced resting state gamma power and reduced evoked gamma synchronization. The gamma band inter-trial coherence of neural response was reduced in both auditory and frontal cortex of Fmr1 KO mice stimulated with a sound whose envelope was modulated from 1 to 100 Hz, similar to that seen in humans with FXS. These deficits suggest a form of enhanced ‘resting state noise’ that interferes with the ability of the circuit to mount a synchronized response to sensory input, predicting specific sensory and cognitive deficits in FXS. The abnormal gamma oscillations are consistent with parvalbumin neuron and perineuronal net deficits seen in the Fmr1 KO mouse auditory cortex indicating that the EEG biomarkers are not only clinically relevant, but could also be used to probe cellular and circuit mechanisms of sensory hypersensitivity in FXS.

Development of Harmaline-induced Tremor in a Swine Model.

BackgroundIn the field of translational neuroscience research, it is critical to utilize a large animal model to test the feasibility, safety, and functionality of novel therapies. Here, we describe a protocol for the development of a large animal model of tremor.MethodsIn a pig model, tremor was induced with harmaline and measured with wireless accelerometers attached to the limbs. Three different doses of harmaline were tested and three repetitive injections were made at 72-hour intervals. To fully characterize the drug-induced tremor, onset time, tremor amplitude, maintained duration, and peak tremor frequency were analyzed.ResultsHarmaline-induced tremor appeared immediately following intravenous injection of harmaline. Tremor was maintained over 2 hours. Its frequency was 10–16 Hz, which was independent of doses. Dose-dependent responses were observed in tremor amplitude, triggering time, and tremor-maintained duration. Repetitive injection of harmaline desensitized the harmaline effect.DiscussionWe provide a detailed protocol for training, drug injection, device selection, and tremor recording optimized to create a swine model of tremor with harmaline. Our protocol provides reliable tremor in pigs and suggests pig as a valid translational large animal model of tremor.

P.2.008 - Paradoxical kinesia induced by appetitive 50-kHz ultrasonic vocalisations in rats depends on glutamatergic mechanisms in the inferior colliculus

Immobile Parkinsonian patients [1] may be able to make quick movements, such as catching a ball or running, when excited by external stimuli. There are several reports of this phenomenon called paradoxical kinesia which refers to a sudden transient ability of akinetic Parkinsonian patients to perform motor tasks they are otherwise unable to perform [2]. The mechanisms underlying this phenomenon are unknown due to a paucity of valid animal models that faithfully reproduce paradoxical kinesia. However, in a previous study [3] we proposed a new animal model to investigate paradoxical kinesia in akinetic rats using species-relevant signals, namely rat ultrasonic vocalizations (USV) which are typical for social situations with positive valence like juvenile play or sexual encounters [4]. Our aim in the present study was to uncover underlying brain mechanisms. We focused on the inferior colliculus (IC) since it not only serves as an acoustic relay station, but can also modulate haloperidol-induced catalepsy [5]. To test the role of the IC in paradoxical kinesia induced by 50-kHz USV, male rats received intracollicular administration of NMDA (30 nmol) or diazepam (10 µg or 20 µg) or its respective controls (i.e. physiological saline or vehicle) 10 min before haloperidol (0.5 mg/kg; ip). Approximately 50 min later, they were exposed to playback of 50-kHz USV, time- and amplitude-matched white noise, background noise or silence, 10 min each with 5 min intervals. The catalepsy test was measured during the bar test, which consists of placing the rat with its forepaws on a horizontal bar. The time until it stepped down with both forepaws was measured (maximum 600s). In animals which had received saline microinjections into the IC, playback of 50-kHz USV significantly reduced haloperidol-induced catalepsy (main effect stimulus: F3,20 = 3.14, P = 0.048), and no such effects were observed in the case of other stimuli. However, the intracollicular administration of NMDA prevented this effect of 50-kHz playback presentation on haloperidol–induced catalepsy (main effect drug treatment: F1,12 = 32.78, P