Comparative analysis of rats’ behavior in the «elevated plus maze» after the intranasal administration of single-walled carbon nanotubes in a small dose in subacute experiments of different duration
Authors: Loginova N.A. , Loseva E.V. , Kryuchkova A.V. , Russu L.I. , Mezentseva M.V. 
Published: November 2021

The unique properties of carbon nanotubes (CNT) (increased strength, flexibility, electrical and thermal conductivity) make them very promising for use in various fields of human activity. But the question of their toxic effects on the body is actively discussed in the literature. There is a lot of body toxicity data of large doses of CNT, but the question of the potential danger of small doses of CNT is a practically unexplored issue. Earlier, we showed that a single intranasal injection of single-walled CNT (SWCNT) in a small dose (4 μg / kg) to rats inhibited the mRNA production of most cytokines in both spleen and brain. At the same time, the expression of the early c-fos gene in the olfactory bulb and the pyriform cortex was increased [Loseva E.V. et al., 2016]. It is known very little about the effect of low doses of SWCNT on animal behavior. The aim of this study was to investigate the effect of a small doses of SWCNT with subacute intranasal administration of different duration on the behavior of rats in the elevated plus-maze (EPM) test, which is used to estimate the anxiety. Male Wistar rats were daily intranasal administrated 100 μl of SWCNT suspensions in physiological saline (PS) in small (5.2 μg / kg, n = 9) dose for 4 (n = 9) or 13 (n = 9) days. Control rats were administrated analogically 100 μl of PS (n = 19). A statistical analysis of indicators of rats' behavior in EPM (5 minutes each) was performed using Kruskal-Wallis one-way analysis of variance, and then using the comparison on the non-parametric criterion of Mann-Whitney for independent features by the program STATISTICA 7.0. It was shown that in the EPM test in rats after 4-fold injection of a small dose of SWCNT, as compared with the control group, there was observed the behavior activation in the center and open arm, what was expressed in frequent and lengthy visits, but with a reduction of the average time of such visits. These animals also had increased the vertical exploratory activity (rears), horizontal exploratory activity (head dips) and motor activity, but the grooming indicators were decreased. After chronic administration of SWCNT in a small dose for 13 days, the direction of some of these changes was maintained, but was expressed to a much lesser degree (only at the level of the tendency in comparison to the control group). So, these rats, compared with the control, more often and longer visited the central site, more often visited open arms and less time were in closed arms. At the same time, the average time of the dark visits was smaller, and the average time of grooming, on the contrary, was longer than in the control. Thus, the intranasal administration of SWCNT in a small dose for 4 days in rats had provoked atypical symptoms of anxiety, which were expressed in an agitated behavior in the EPM, as evidenced by the strong activation of most behavioral reactions. The intranasal administration of SWCNT in a small dose for 13 days also has led to some activation of behavior or atypical anxiety, but it was expressed to a much lesser extent than with the acute injection of SWCNT. There wasn't observed an increasing of agitated behavior or a cumulative negative effect of SWCNT during chronic intranasal injection. We can suggest that nanotubes when administered in a small dose for 4 days can penetrate cells and their organelles, causing damage and death of tissue structures, in particular, when they penetrate into the brain, which leads to agitated behavior. At the same time, with subacute administration of SWCNT in a small dose during 13 days, protective reactions can gradually develop, leading to accelerated utilization of nanotubes, for example, by macrophages and astrocytes, which can absorb them, that leads to a diminution of the activation of behavior. To test this assumption, special morpho-functional studies are required.