Animal Model for Anxiety

Anxiety disorders are the most prevalent neuropsychological disorders around the world, especially in Pakistan, where about 34% population is suffering from the anxiety and depression which contribute to the incidents of crimes, and suicidal cases. In addition, it also effects the healthy work effic

Project Title

Animal Model for Anxiety

Project Area of Specialization

Biomedical Engineering

Project Summary

Anxiety disorders are the most prevalent neuropsychological disorders around the world, especially in Pakistan, where about 34% population is suffering from the anxiety and depression which contribute to the incidents of crimes, and suicidal cases. In addition, it also effects the healthy work efficiency, create damage in the work productivity of individuals. Several studies show the hereditary connections with the anxiety, similarly, genetic studies have highlighted the important tool to early diagnosis of trauma, stress, and anxiety. Therefore, there is much focus on neuroimaging and genetic research for the diagnosis of anxiety and other related disorders.  To understand the underlying mechanism that how some anxious genes responsible for the symptoms and pathogenesis of anxiety. Among those factors that can contribute to the symptoms of anxiety, there are some food and edible substances that can trigger the anxiety by various mechanisms. In previous studies we found that Juglans Regia (Walnut) which is rich in potentially neuroprotective compounds but unfortunately it can trigger the anxiety, our preliminary studies showed that walnuts produced significant anxiety in albino wistar male rats. Keeping this in mind, the present study is designed to identify the possible effect of Juglans Regia (Walnut) on the anxious genes in the animal brain.  Top candidate genes that are responsible for the anxiety are selected for this study including gamma-aminobutyric acid (GABA) B receptor-1 (GABBR1), FOS/FBJ osteosarcoma oncogene, interleukin 1 beta and Brain derived neurotropic factor (BDNF), it has been reported that there is significant higher expression level of these anxious genes in different brain areas.

Project Objectives

To observe the effect of Juglans Regia (Walnut) on anxiety level and to find out its possible effect on anxious genes expression level.

Project Implementation Method

1. Administration of Juglans Regia (Walnut) in rat model

All rats will be housed in a room that will maintained at 21-25°C with a controlled light-dark cycle. Daily Juglans Regia (Walnut) will be administered orally (at final concentration of 10 mg/ml) for 7 days.

2. Drugs

The Juglans Regia (Walnut) was crushed and suspended in distilled water. Diazepam (1 mg/ kg), anxiolytics drug is selected as a positive control for anxiolytic-like effects. While the normal control groups received only 0.9% normal saline orally.

3. Elevated plus maze

The elevated plus maze (EPM) was widely used as an animal model of anxiety. Entries in open arm and the time duration is open and closed arm will be recorded and interpreted as an index of potential anxiogenic or anxiolytic activity.

4. Open field test

Another test which is used to measure the anxiety level in animals is open field test. Behavioral pattern that reflects the anxiety index in animals will be observed including exploratory behaviors, total number of crossings, central area crossings, time spent in central zone, inner and outer zone rearing.

5. Determination of GABBR1, FOS/FBJ, interleukin 1 beta and BDNF mRNA

After 7 to 10 days treatment all animals will be sacrificed, and their brain samples will be collected and store it -20oC till further analysis. To evaluate the effect of walnut, diazepam on gene expression of different anxious genes, mRNA quantification will be performed by extraction of total using the RNA extraction commercial kit or by TRIZol method). The cDNA will be synthesized using commercial cDNA kit, according to the manufacturer's protocol. The values obtained for the target gene expression will normalized to Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and quantified relative to the expression in control samples.  The transcribed cDNA will be applied using commercial kit and using oligonucleotide primers corresponding to transcripts of anxious genes. The PCR mixture for reaction will be prepared as outlined in the protocol.

The PCR reaction product will be resolved on 1 % agarose gel containing ethidium bromide. The density of each band will be quantified using NIH online J image software.

6. Statistical analysis

Statistical analysis of the data will be carried out using analysis of variance (ANOVA).  The Bonferroni's post hoc test will used to determine which group mean differs. Any values below the level of 0.05 will considered as significant.

Benefits of the Project

After establishing the fact that walnut can cause an anxiety at genetic level or may induce neuroplastic changes that may lead in permanent damages, an awareness campaign will be started that the walnut consumption is very beneficial and nutritious, but it should be taken with other dry fruit such as raisin, so that the prevalence of anxiety and anxiety like other neurological disorder can be reduced significantly.

Technical Details of Final Deliverable

User Observation

Process modeling

Verify scope

Final Deliverable of the Project

HW/SW integrated system

Core Industry

Health

Other Industries

Core Technology

Others

Other Technologies

Sustainable Development Goals

Required Resources

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