**1. Introduction**

The menopausal transition is often associated with a multiplicity of manifestations, the most standard being neuropsychiatric [1, 2]. The role of ovarian hormones in affect-related disorders is of great interest for women transitioning through menopause [2, 3]. Mood disorders during menopause could partly be explained due to a loss of estrogen is known to have neuroprotective effects on brain [3]. Numerous experimental and clinical studies have documented that estrogen deficiency during menopause increases the susceptibility to mood disturbances, including anxiety [4–6]. There has been a discussion that menopausal hormonal therapy (MHT) may improve the symptoms of affective-related disorders or decrease the risk of developing these, yet some uncertainty still exist around this topic because as research has

also found that MHT does not entirely stop the development of affective-related symptoms [7].

Females going through menopause are at higher risk of developing Vitamin D (VD) deficiency due to a VD poor diet, restricted outdoor activity resulting in less sun exposure as well as a decreased capacity to produce enough calcitriol as a result of an age related decline in hydroxylation by the kidneys [8]. Our previous experimental work has confirmed that hormonal profile in ovariectomized (OVX) female rodents is also characterized by VD deficiency or insufficiency [9, 10]. Traditional methods of affective-related disorders therapy, which also include antidepressants/ anxiolytics, are unfortunately of limited effectiveness [11]. Nutrient imbalance, especially VD3 deficiency, is considered as one of the critical causes, enabling the pathophysiological mechanisms for development of psychiatric disorders [12]. In the pathophysiological mechanisms of mood disorders, many trigger factors play a role, and it is argued that one of them could be a deficiency in VD3 [12].

VD3 deficiency has been proven to impact on the pathogenesis of various diseases, for example, autoimmune diseases, cardiovascular diseases, infections, osteoporosis, obesity, diabetes, and certain types of cancers [13–15]. A correlation between very low VD3 levels and numerous neuropsychiatric diseases and a correlation between an impact of VD3 levels and normal brain functioning have also been found in recent studies [14–16]. VD receptors (VDRs) have been found present in the central nervous system [17], in the brain structures involved in processes of mood regulation (cingulate cortex, hippocampus, thalamus, and hypothalamus) [18]. In this line, it can be assumed that VD3 likely has humoral or neurohumoral activities in these brain structures. VD3 involves in the neurogenesis, neuroplasticity, neuroprotection, and neuroimmunomodulation [19–21]. This fact creates a neurobiological basis to propose the involvement of VD in the mechanisms of neuropsychiatric disorders [22–26].

The neuroinflammation in the central nervous system is supposed to be one of the main trigger factors for the development of affective-related disorders [27, 28]. Taking this assumption into account, mood disturbances established in menopausal women might result from complex alterations in estradiol and VD3 levels, as well as neuroinflammation.

Nowadays, nuclear factor-kappa B (NF-κB) is postulated as the proinflammatory transcription factor that controls proinflammatory cytokines expression and is involved in the mechanisms of many inflammatory and neuroinflammatory diseases [29, 30]. NF-κB is triggered by stress and might mediate cellular responses to stressful life events, thereby critically involved in development of affective-related disorders [31–33]. The enhancement of NF-κB might induce the elevated production of proinflammatory cytokines and diminished neurohormonal stress feedback [34]. Furthermore, NF-κB pathway is involved in antidepressant action of different psychotropic drugs that used for treatment of mood disorders [35]. Clinical studies using patients with mood disorders have shown that NF-κB levels are increased in the serum of such patients [35–37]. Using genetic and environmental model of depression, it was shown that the antidepressant effect of such pharmacological treatments was dependent on NF-κB-p65 acetylation [36, 37].

The hippocampus is one of the key structures of the brain, which plays a role in affective-related disorders [38]. Both estrogen and VD3 have been associated with the successful functioning of the hippocampus [1, 21, 25]. Basic and clinical studies have suggested that alterations in NF-kB/p65 signaling and in 17β-E2/VD3 receptors expression in the hippocampus, as well changes of serum estradiol/VD contents are very often registered at affective-related disorders [1, 23, 39]. Animal studies have documented that the impaired behavioral profile in OVX rats is correlated with increased NF-kB/p65 levels in the brain [40, 41].

**155**

*Vitamin D3 Modulates NF-kB/p65, 17β-Estradiol, and Vitamin D Receptors Expression…*

sion in the hippocampus of long-term OVX adult rats with CUMS.

Recently, we found that VD3 (5.0 mg/kg, s.c.) reduced anhedonia and depression-like behavior of long-term adult ovariectomized (OVX) rats exposed to the chronic unpredictable mild stress (CUMS) in the sucrose preference (SPT) and forced swimming (FST), respectively [42]. However, the therapeutic effects of VD3 in a combination with low dose of 17β-E2 in female rats with long-lasting decline of estrogens exposed to CUMS remain unknown. Furthermore, it is still unclear whether the antidepressant-like action of VD3 plus 17β-E2 application implicates NF-kB/p65 signaling pathway and modifications of 17β-E2/VD3 receptors expres-

The current investigation was performed to clarify the antidepressant-like effect of a combination with VD3 plus low dose of 17β-E2 on a rat model of CUMS in the female rats with long-lasting decline of estrogens. Similar to previously published work [43], we used long-lasting estrogen deficiency caused by a post-ovariectomy period of 3 months. This animal model is widely utilized in preclinical behavioral research producing a menopausal-like state in women [44]. Such behavioral tests as sucrose preference (SPT), forced swimming (FST), and open-field (OFT) were carried out to examine the depression-like behavior. NF-kB/p65, 17β-E2/VD3 receptors levels in the hippocampus and serum estradiol and VD concentrations were determined to assess the possible mechanisms of the VD3 effects on the depression-like behavior in long-term OVX rats given with low dose of 17β-E2 subjected to CUMS.

A total of 49 Wistar rats of 3 months age, female sex (weighing 200–220 g) were purchased in this work. Animals were divided into experimental groups with access to rat standard food and water ad libitum. The female rats were placed under a 12 light-dark scheme (light was given between 07:00 and 19:00 h) and room temperature (23 ± 2°C). All behavioral procedures and CUMS model were performed in compliance with the National research council's guide for the care and use of laboratory animals and approved by the Ethical committee for experimental studies of I.P. Pavlov Institute of Physiology (statement No.: 1095/1/25.06.2012). Stress model

Three months before CUMS procedure, sham operation and long-term total ovariectomy with general anesthesia (ketamine 70 mg/kg and xylazine 10 mg/kg, i.p.) were performed. Long-term period (3 months) elimination of female gonadal hormones was chosen as experimental model of menopause in women [44, 45]. The removal of ovaries was carried out accordingly to our method as prescribed earlier [43]. After surgery or sham-operation (SHAM), the ovariectomized (OVX) females were placed in home cage with free access to food and water. During 12 weeks, sham-operated and OVX females had a recovery. Following 3 months of surgery, experimental rats were randomly distributed to the groups for the chronic stress

Chronic unpredictable mild stress (CUMS) paradigm is a valid and significant animal model of depression induced by stress procedure. This behavioral model

of depression was conducted with minimal pain for all groups of rats.

procedure, except for SHAM non-stressed control rats.

*DOI: http://dx.doi.org/10.5772/intechopen.89357*

**2. Materials and methods**

**2.1 Animals**

**2.2 Ovariectomy**

**2.3 CUMS model**

#### *Vitamin D3 Modulates NF-kB/p65, 17β-Estradiol, and Vitamin D Receptors Expression… DOI: http://dx.doi.org/10.5772/intechopen.89357*

Recently, we found that VD3 (5.0 mg/kg, s.c.) reduced anhedonia and depression-like behavior of long-term adult ovariectomized (OVX) rats exposed to the chronic unpredictable mild stress (CUMS) in the sucrose preference (SPT) and forced swimming (FST), respectively [42]. However, the therapeutic effects of VD3 in a combination with low dose of 17β-E2 in female rats with long-lasting decline of estrogens exposed to CUMS remain unknown. Furthermore, it is still unclear whether the antidepressant-like action of VD3 plus 17β-E2 application implicates NF-kB/p65 signaling pathway and modifications of 17β-E2/VD3 receptors expression in the hippocampus of long-term OVX adult rats with CUMS.

The current investigation was performed to clarify the antidepressant-like effect of a combination with VD3 plus low dose of 17β-E2 on a rat model of CUMS in the female rats with long-lasting decline of estrogens. Similar to previously published work [43], we used long-lasting estrogen deficiency caused by a post-ovariectomy period of 3 months. This animal model is widely utilized in preclinical behavioral research producing a menopausal-like state in women [44]. Such behavioral tests as sucrose preference (SPT), forced swimming (FST), and open-field (OFT) were carried out to examine the depression-like behavior. NF-kB/p65, 17β-E2/VD3 receptors levels in the hippocampus and serum estradiol and VD concentrations were determined to assess the possible mechanisms of the VD3 effects on the depression-like behavior in long-term OVX rats given with low dose of 17β-E2 subjected to CUMS.
