Influence of environmental enrichment during adolescence on behavioral and neuroinflammatory changes induced by maternal deprivation in male and female rats

Abstract

The neonatal period is extremely important for programming physiological and behavioral responses to stress in later stages of development, and a large part of this process depends on the quality of the mother-infant relationship. In rodents, the rupture of this relationship, through maternal deprivation (MD) for 24 h, during the period of hyporesponsiveness to stress, alters the trajectory of the neurodevelopment of the offspring. In previous work, our group demonstrated that MD on postnatal day 3 (MD3 group) produces increased anxiety-like behavior in young adult males and females, and reduced motivation to explore a conspecific in adolescent males. Males and females from groups MD3 and MD11 also showed an increase in the frequency of immobility in the forced swimming test, indicating that this neonatal manipulation results in manifestations of behaviors relevant to anxiety and depression in an age- and gender-dependent manner. Furthermore, neuroinflammation, highly correlated with chronic stress, causes hyperactivation of the kynurenine pathway, with increased production of pro-inflammatory cytokines and neurotoxic factors by glial cells, especially microglia and astrocytes, in addition to having an impact on the decrease concentrations of Brain-Derived Neurotrophic Factor (BDNF) and neurogenesis. Environmental enrichment (EE), in turn, represents a non-pharmacological intervention of physical, motor, sensory, cognitive and social stimulation that has shown positive results in the prevention and reversal of deleterious effects induced by stress protocols. In the present project, our aim is to evaluate the potential of EE to reverse the behavioral and neuroinflammatory effects induced by MD. For this, the litters will be standardized in 4 males and 4 females and distributed between control groups (CTL), MD3 and MD11 and, after weaning, 2 brothers of the same sex will be housed in standard house (SH) or in cages with EE for 4 weeks, followed by 1 week of SH accommodation. In each litter, one animal representative of each sex/group/housing will be subjected to the behavioral tests, while the other representative will not be tested. After the end of the EE, the animals will be submitted to the Sucrose Splash Test, to evaluate the depressive-like behavior, through the latency and frequency of the self-cleaning behavior; to Open Field, to assess anxiety-like behavior; and the Social Investigation and Interaction Tests to assess social preference and motivation. After 1 week housed in SH, the animals will be submitted again to the same tests and, on the following day, to the Elevated Plus Maze (EPM) to evaluate the anxious behavior. Thirty min after the EPM, the animals will be euthanized and blood, adrenal glands, spleen and brain will be collected. Measurement of plasma corticosterone concentrations (CORT) will be performed by radioimmunoassay. Brain concentrations (hippocampus and frontal cortex) of BDNF and of the main enzyme of the kynurenine pathway, idolamine 2,3-dioxygenase (IDO), will be determined by Western Blot. Pro-inflammatory cytokines will be measured in the hippocampus and frontal cortex by Multiplex Immunoassays. Immunohistochemistry will also be performed for the histological analysis of microglia (M1 and M2 profile) and astrocytes and for the evaluation of hippocampal neurogenesis through DCX and Neu-N proteins. The results of this project may provide evidence of the effectiveness of EA in reversing the harmful effects of PM on emotional behavior and neuroinflammation in males and females adolescent rats.