Twenty first-day pregnant Albino-Wistar rats purchased Pasteur Institute of Iran, were employed in the present investigation (weight: 180-220 g). The advent of the vaginal plug after mating was computed as the first day of gestational day. The pregnant rats were kept in two separate temperature-controlled rooms (23±2°C): i) a standard unshielded room (for pregnant rats in the EMF group); ii) a shielded room, which was insulated against EMF using aluminum foil (for pregnant rats in control (CON) group).²⁸ The rats were independently housed in standard Plexiglas cages. They had access to unlimited food and water sources. Each group was exposed to a cycle of 12 hours of light and 12 hours of dark.²⁹ After delivery, 12 male neonates from EMF and CON maternal groups were randomly selected for the experiment using a simple randomization approach. Helsinki ethics were applied in this research.

At 56 post-natal days, the subjects were deeply anesthetized using Medetomidine hydrochloride, 1mg/ml (Dorbene Vet brand). Then the brain tissues of the rats were removed and stored in liquid nitrogen at -80°C. Hippocampal samples were centrifuged using a German Hectic centrifuge for 5 min at around 16,000 rpm. The protein content of the hippocampus in the brain tissue was then analyzed by a Rat BDNF ELISA kit (Zellbio, Germany) using the manufacturer’s instructions to an accuracy of 0.2. The results were evaluated using an ELISA reader (Stat Fax, US), and the absorbance of the samples was read at 450 nm.

The descriptive statistics that were evaluated in this study included the mean and standard deviation. Firstly, the normal data distribution was tested using a Shapiro-Wilk, and Leven statistic to test the homogeneity of variances. Anthropometric measurements, and body measurements in each group were assessed using ANOVAs with repeated measures at three stages: after delivery, 21 post-natal days, and 56 post-natal days. One-way ANOVA was used in Bonferroni post-hoc manner. Assigning differences between groups, the independence T-test was applied. Total outcomes were depicted as Mean±SD in all statistical differentiation. The scale for statistical significance was P≤0.05.

In Figure 1A-C, the comparison of the weight, body length, and tail length changes among the CON and EMF groups are displayed.

Statistical analysis revealed a significant rise in body weight, body length, and tail length during the three measurement periods. In the EMF group at A) delivery: F (1.01, 10) =345.85, P=0.0001; B) 21 PND: F (2, 10) =319.011, P=0.0001; C) 56 PND: F (2, 10) =358.93, P=0.0001; and in the CON group at A) delivery: F (1.46, 10) =4554.76, P=0.0001; B) 21 PND: F (2, 10) =1971.29, P=0.0001; C) 56 PND: F (2, 10) =2111.99, P=0.0001).

The Bonferroni post-hoc tests in the EMF group discovered consistent changes in body weight, body length, and tail length from delivery day to 56 PND (body weight: 6.21±0.11 gr, 35.55±6.33 gr, 206.55±27.14 gr; body length: 51.02±0.64 mm, 92.21±9.25 mm, 171.38±9.40 mm; tail length: 19.95±0.03 mm, 46.33±7.02 mm, 136.45±9.89 mm), and also in the CON group (body weight: 6.32±0.19 gr, 36.16±5.37 gr, body length: 229.73±4.83 gr; 45.58±0.32mm, 104.59±5.20mm, 181.10±3.87mm; tail length: 17.17±0.05 mm, 67.31±4.75 mm, 156.23±6.44 mm).

As illustrated in Figure 2, this study found that body weight in delivery, 21 postnatal days, and 56 postnatal days in neonates who were exposed to EMF (6.21±0.11gr; 35.55±6.33gr; 206.55±27.14gr, respectively) lower in comparison to that found in the control group (6.32±0.19 gr; 36.16±5.37gr; 229.73±4.83gr) however, these rates were not significant (t (10) = 1.20, P=0.257; t (10) = 0.182, P=0.859; t (10) = 2.06, P=0.066, respectively).

In contrast, the body length of the EMF group at delivery time was significantly higher than the control group: 51.02±0.64mm vs. 45.58±0.32mm, t (10) = -18.43, P=0.0001). However, the control group’s body length at 21 PND: 104.59±5.20 mm vs. 92.21±9.25 mm, correspondingly was significantly higher than that of the EMF group and at 56 PND: 181.10±3.87mm vs. 171.38±9.20 mm), t (10) = 2.85, P=0.017; t (10) = 2.34, P=0.041 (Figure 3).

Moreover, according to the Statistical analysis outcomes, the tail length of EMF group (19.95±0.03mm) in delivery time was significantly higher than the control group (17.17±0.05mm), t (10) = -96.69, P=0.0001. Conversely, at 21 PND, the control group’s tail length was significantly longer than that of the EMF group 67.31±4.75mm vs. 46.33±7.02mm; and at 56 PND 156.23±6.44mm vs. 136.45±9.89 mm, t (10) = 6.06, P=0.001, and t (10) = 4.100, P=0.002, respectively (Figure 4).

The BDNF factor concentration comparisons between groups were assessed by the independent t-test, and the outcomes are illustrated in radar Figure 5.

This study found that neonates who had been affected by EMF exposure had significantly lower, statistically BDNF concentrations (6.8±0.2 ng/ml) at 56 post-natal days compared to the control group (7.08±0.16 ng/ml), t (10) = 2.59, P=0.027.

Examining the effects of 2.4 GHz EMF Exposure on anthropometric measurements, BDNF concentrations in the fetuses of exposed pregnant rats were the primary study’s purpose. The results were: i) 2.4 GHz EMF exposure in the pregnancy period leads to the reduced physical growth of the fetuses after birth; ii) 2.4 GHz EMF exposure during pregnancy results in lower BDNF concentrations in male rats.

As mentioned previously, the fetus’s body weight in the EMF exposure group was significantly lower compared to the CON group. Moreover, body length and tail length were significantly lower in the EMF group than in the CON group.

In zoological texts, the index of rat growth is assessed by body weight,²⁶ and the BDNF levels are manipulated by body weight.³⁴ According to the animal resource center report, the expected average weight for male Albino-Wistar rats is 305 gr.³⁵ As our results illustrate, both groups are lower than the normal range. On the other hand, the CON group at 56 PND is heavier than the EMF group. Therefore, the control group had better physical development. Moreover, prior studies have indicated that both obesity and anorexia have a direct and inverse relationship with BDNF.^22,23,34 These findings are not in line with previous investigations that suggest that EMF exposure causes obesity.³⁶ Many investigations have shown that BDNF levels can affect appetite.^34,37,38 BDNF is a component of the neurotrophins of concealing signaling molecules that involve nerve growth factor, neurotrophin-3, and neurotrophin- 4/5.³⁹ The major molecule involved in controlling body weight is leptin, which is the protein product of the obese (ob) gene and is formed in and secreted by adipose tissue and acts as an indicator of fat mass.⁴⁰ Adjusting the formation, preservation, and action of neuronal connections is the way BDNF manipulates energy homeostasis.⁴¹ Various investigations have revealed that BDNF affects other anorexigenic factors, such as leptin and cholecystokinin, to adjust food intake and body weight.⁴¹ Introducing peripheral leptin was found to improve the BDNF mRNA level in the ventromedial hypothalamus,⁴² and the BDNF factor rate in the dorsal vagal complex.⁴³ Improving BDNF mRNA in dendrites results from these effects, while leptin grew dendritic BDNF mRNA translation in hypothalamic neurons.⁴⁴ Initiating BDNFexpressing neurons through a polysynaptic neural circuit is the way that leptin obliquely begins synthesis of BDNF mRNA and BDNF Factor.⁴¹ BDNF stimulates the growth and preservation of neural circuits.⁴⁵ So, obesity syndrome associated with BDNF inadequacy could result from structural impairment in neural circuits, controlling energy homeostasis, or reduced anorexigenic actions of BDNF.¹⁸