The influence of sex and neonatal stress on medullary microglia in rat pups.

NEW FINDINGS: What is the central question of the study? Does neonatal stress, in the form of neonatal maternal separation, influence the maturation of microglial density, morphology and neuronal signalling in medullary regions regulating cardiorespiratory function in rat pups? What is the main finding and its importance? Using Iba-1 immunohistochemistry, we show that neonatal maternal separation augments microglial density and the proportion of cells with an amoeboid morphology in the medulla. Although the current understanding of the effect of early life stress on medullary development is relatively limited, these data show that within this area, microglia are affected by neonatal stress. Microglia could therefore be important effectors in cardiorespiratory disorders resulting from maternal separation. ABSTRACT: Neonatal stress has wide-ranging consequences for the developing brain, including the medullary cardiorespiratory network. In rat pups, the reflexive cardiorespiratory inhibition triggered by the presence of liquids near the larynx is augmented by neonatal maternal separation (NMS), especially in males. Sex-specific enhancement of synaptic connectivity by NMS might explain this cardiorespiratory dysfunction. Microglia influence the formation, maturation, activity and elimination of developing synapses, but their role in the wiring of medullary networks is unknown. Owing to their sensitivity to sex hormones and stress hormones, microglial dysfunction could contribute to the abnormal cardiorespiratory phenotype observed in NMS pups. Here, we first used ionized calcium-binding adapter molecule-1 (Iba-1) immunolabelling to compare the density and morphology of microglia in the medulla of male versus female rat pups (14-15 days old) that were either undisturbed or subjected to NMS (3 h day-1 ; postnatal days 3-12). Neonatal maternal separation augmented the density of Iba-1+ cells (caudal region of the NTS), increased the size of the soma and reduced the arborization area (especially in the dorsal motor nucleus of the vagus). Sex-based differences were not observed. Given that the actions of microglia are regulated by neuronal fractalkine (CX3 CL1 ), we then used western blot analysis to compare the expression of CX3 CL1 and its microglial receptor (CX3 CR1 ) in medullary homogenates from control and NMS pups. Although CX3 CR1 expression was 59% greater in males versus females, NMS had no effect on CX3 CL1 /CX3 CR1 signalling. Given that an amoeboid morphology reflects an immature phenotype in developing microglia, NMS could interfere with synaptic pruning via a different mechanism.

Fluoxetine treatment affects the inflammatory response and microglial function according to the quality of the living environment.

It has been hypothesized that selective serotonin reuptake inhibitors (SSRIs), the most common treatment for major depression, affect mood through changes in immune function. However, the effects of SSRIs on inflammatory response are contradictory since these act either as anti- or pro-inflammatory drugs. Previous experimental and clinical studies showed that the quality of the living environment moderates the outcome of antidepressant treatment. Therefore, we hypothesized that the interplay between SSRIs and the environment may, at least partially, explain the apparent incongruence regarding the effects of SSRI treatment on the inflammatory response. In order to investigate such interplay, we exposed C57BL/6 mice to chronic stress to induce a depression-like phenotype and, subsequently, to fluoxetine treatment or vehicle (21days) while being exposed to either an enriched or a stressful condition. At the end of treatment, we measured the expression levels of several anti- and pro-inflammatory cytokines and inflammatory mediators in the whole hippocampus and in isolated microglia. We also determined microglial density, distribution, and morphology to investigate their surveillance state. Results show that the effects of fluoxetine treatment on inflammation and microglial function, as compared to vehicle, were dependent on the quality of the living environment. In particular, fluoxetine administered in the enriched condition increased the expression of pro-inflammatory markers compared to vehicle, while treatment in a stressful condition produced anti-inflammatory effects. These findings provide new insights regarding the effects of SSRIs on inflammation, which may be crucial to devise pharmacological strategies aimed at enhancing antidepressant efficacy by means of controlling environmental conditions.