Ameliorative effects of ketamine administration in experimental model of Chronic Unpredictable Mild Stress.

Introduction

Depression, also known as major depressive disorders, is one of the commonest psychiatric disorders and causes of disability across the world. It presents a serious socio-medical burden that will become more serious in the coming years according to WHO expectation^[1]. Symptoms of depression vary according to its severity, spread to many domains of life affecting person’s emotional, social, occupa-tional functioning. These symptoms show changes in mood, anxiety, memory loss, anhedonia, optimism, sleep change rhythm, energy loss, appetite change,  loss of libido, and psychomotor activity^[2].

The treatment plans for depression consist of pharmacological and non-pharmacological strategies containing psychotherapeutic appro-ach as cognitive behavioral, relaxation and imagery techniques, acupressure with massage and nutritional supplements. Forty percent of patients do not respond adequately to currently available anti-depressant medications^[3], so they open the way for new medications to develop and assist in reducing the progression of depression and its possible complications ^[4].

The aim of the current study is to induce depression in adult male albino rats and to study in them some pathophysiological changes. Cumulatively, the present study showed the effect of treatment by a single sub-anesthetic ketamine dose on pathophysiological changes associated with depression.

Materials and methods

• Animals;

Thirty adult male albino rats of national strains with average weight 200 ± 20 gm were gained from the National Research Center, Cairo, Egypt. They were housed in well ventilated cages made of stainless steel cages at normal room temperature with normal dark and light sequences. They were left for two weeks after arrival for acclimatization. Rats were fed a commercial rat's diet (Nile Company, Egypt) and tap water ad libitum through the time of the study except the timing of food and water deprivation in stressor’s groups. All experiments were performed according under the appropriate animal licenses No.323:4/2022 permitted by the animal care committee of Faculty of Medicine-Minia University, according to the international guidelines. 

• Experimental design;

Rats were categorized into the following three equal groups of ten rats each: 

• control group (C): in which rats were allowed freely wandering in their cages with free access to food and water.
• Chronic unpredictable mild stress group (CUMS): in which rats faced different physical stressors for six weeks. Stressors like water deprivation, food deprivation, noise. These stressors were administered daily and to have the unpredictable procedure, two stressors were applied daily in random manner ^[5].
• Ketamine treated CUMS group (K. CUMS): in which rats were exposed to different physical stressors for six weeks^[5] then every rat received a single sub-anesthetic intraperitoneal injection of ketamine at a dose of 10 mg/kg, 24 hours prior to the end of the experiment^[6].

• Behavioral test;

After the end of the CUMS period, before and after ketamine administration, sucrose prefe-rence test was executed as follow: rats were singly housed in a cage after a 12 hours’ duration of food and water withdrawal. After that, all rats were given 2 bottles containing water or 2% sucrose solution. Sucrose and water bottles were put  and assigned randomly in sides of every cage, Sucrose preference test includes two sections: an initial for 24 hours before drug administration (the baseline sucrose test ) then for 2 hours after drug administration (the sucrose preference test). The weights of water and sucrose consumed were measured according the following equation: Sucrose solution (ml)/ (Sucrose solution [ml] + water [ml]) ×100% ^[7].

• Sample collection;

At the end of the whole experimental design, and after the end of second section of behavioral test, the surviving rats were sacrificed by cervical dislocation. The heads of sacrificed rats were rapidly dissected carefully and the brains were removed in gentle way. Brain was used to expose the hippocampus which was identified as a C shaped structure in coronal section. Then the hippocampal brain tissue was dissected, weighed, homogenized to assay serotonin that was determined spectroflurophotometrically as Ciarlone (1978)^[8] by using spectroflurophotometer (Shimaduz RF- 5000), Japan).

Statistical analysis of data:

The data were expressed as the means ± Standard deviation(SD), comparisons between different groups were performed by one-way analysis of variance (one-way ANOVA), then the least significant difference (LSD) test. Results with p < 0.001 were considered statistically significant. Statistics were analyzed using IBM SPSS 28.0 statistical package software (IBM; Armonk, New York, USA).

Results

• Effect of CUMS with or without ketamine on sucrose preference tests:

Table (1) show that CUMS reduced the sucrose tests as compared with control group in both the baseline (24 h) and the preference tests (2 h). No significant difference in baseline sucrose test before drug administration was seen between all stressed groups as compared with the stressed group (CUMS). Concerning sucrose preference test, the ketamine treated CUMS group showed a significant higher sucrose preference test as compared to CUMS group but still significantly lower than the control group.

• Effect of CUMS with or without ketamine on hippocampal Serotonin level:

The results of the current study exhibited that the hippocampal serotonin level in CUMS group showed significant lowest levels among all experimental groups. On the other hand, the hippocampal serotonin levels were significant higher in ketamine treated CUMS as compared to CUMS group but still lower than control group as shown in table (2).

Discussion

Depression is one of the most prevalent psychiatric illness in the world. Nearly 300 million people around the world suffer from depressive episode in their lives which equal to 4.4% of the world’s population ^[9].

Depression in animals can be induced by numbers of behaviorally tests, based mainly on social and environmental reinforcement with a good credibility and validity ^[10]. Despite the wide variety of depressive-inducing protocols, the chronic unpredictable mild stress (CUMS) animal model was chosen in the present study as it is one of the most accepted behavioral models. In CUMS, the laboratory animals faced external, variable, different and unpredictable tests simulating different unpredictable life stressors which elicit emotional and behavioral responses contributing to neuropsychiatric disorders, such as major depressive disorders in humans^{[11] , [12]}.

The mesolimbic system is also known as the reward system that is responsible for emotional expression, learning, behavioral response and other motivational cognition. Chronic stress induces reactive oxygen species (ROS) that cross BBB to change mesolimbic system activity. Depression is very common with its anhedonia behavior. CUMS model is obviously known as anhedonia-inducing potential ^[13]. In the present study, the investigation of the depressive and anhedonia like behaviors was done by sucrose preference test (SPT). The CUMS group significantly decreased SPT as chronic stress causes dendritic remodeling in prefrontal cortex as well as  functional changes in behavioral response^[14].

The present results demonstrated that the sucrose preference test was significant higher in ketamine treated CUMS groups as compared with CUMS group. Kingir and his colleagues 2023 explained the anti-depressant action of ketamine as they modulate the neurotrans-mission in mesolimbic system affecting behavioral response in animal ^[15].

Regarding to the monoamine theory, Monoa-mine oxidase (MAO) density is elevated during episodes of depression. This results in greater metabolism of catecholamines, such as serotonin (5-HT) as shown in the present study. Catecholamines usually participate in mental activities and emotional control in body and it is confirmed to related directly to the onset, pathophysiology and symptoms of depression^[16]. Additionally, the serotonin deficiency hypothesis confirmed multiple abnormalities of serotonin receptors with a decrease of its availability in multiple brain areas of patients with MDD, which regulates serotonin function in brain ^[17].

On the other hand, ketamine administration significantly increases hippocampal serotonin level by acting through AMPA receptors, which is localized to serotonergic neurons eventually arouses serotonin secretion by stimulating 5-HT1A receptors and 5 hydroxy tryptaminergic transmission in the hippocampal^18].