ATP-P2X7 signaling mediates brain pathology while contributing to viral control in perinatal Zika virus infection.

Zika virus (ZIKV), the causative agent of Zika fever, is a flavivirus transmitted by mosquitoes of the Aedes genus. Zika virus infection has become an international concern due to its association with severe neurological complications such as fetal microcephaly. Viral infection can induce the release of ATP in the extracellular environment, activating receptors sensitized by extracellular nucleotides, such as the P2X7 receptor. This receptor is the primary purinergic receptor involved in neuroinflammation, neurodegeneration, and immunity. In this work, we investigated the role of ATP-P2X7 receptor signaling in Zika-related brain abnormalities. Wild-type mice (WT) and P2X7 receptor-deficient (P2X7-/-) C57BL/6 newborn mice were subcutaneously inoculated with 5 × 106plaque-forming units of ZIKV or mock solution. P2X7 receptor expression increased in the brain of Zika virus-infected mice compared to the mock group. Comparative analyses of the hippocampi from WT and P2X7-/-mice revealed that the P2X7 receptor increased hippocampal damage in CA1/CA2 and CA3 regions. Doublecortin expression decreased significantly in the brains of ZIKV-infected mice. WT ZIKV-infected mice showed impaired motor performance compared to P2X7-/- infected mice. WT ZIKV-infected animals showed increased expression of glial markers GFAP (astrocytes) and IBA-1 (microglia) compared to P2X7-/- infected mice. Although the P2X7 receptor contributes to neuronal loss and neuroinflammation, WT mice were more efficient in controlling the viral load in the brain than P2X7 receptor-deficient mice. This result was associated with higher induction of TNF-α, IFN-ß, and increased interferon-stimulated gene expression in WT mice than P2X7-/-ZIKV-infected. Finally, we found that the P2X7 receptor contributes to inhibiting the neuroprotective signaling pathway AKT/mTOR while stimulating the caspase-3 activation, possibly two distinct pathways contributing to neurodegeneration. These findings suggest that ATP-P2X7 receptor signaling contributes to the antiviral response in the brain of ZIKV-infected mice while increasing neuronal loss, neuroinflammation, and related brain abnormalities.

GABAA overactivation potentiates the effects of NMDA blockade during the brain growth spurt in eliciting locomotor hyperactivity in juvenile mice.

Both NMDA receptor blockade and GABAA receptor overactivation during the brain growth spurt may contribute to the hyperactivity phenotype reminiscent of attention-deficit/hyperactivity disorder. Here, we evaluated the effects of exposure to MK801 (a NMDA antagonist) and/or to muscimol (a GABAA agonist) during the brain growth spurt on locomotor activity of juvenile Swiss mice. This study was carried out in two separate experiments. In the first experiment, pups received a single i.p. injection of either saline solution (SAL), MK801 (MK, 0.1, 0.3 or 0.5 mg/kg) or muscimol (MU, 0.02, 0.1 or 0.5 mg/kg) at the second postnatal day (PND2), and PNDs 4, 6 and 8. In the second experiment, we investigated the effects of a combined injection of MK (0.1 mg/kg) and MU (doses: 0.02, 0.1 or 0.5 mg/kg) following the same injection schedule of the first experiment. In both experiments, locomotor activity was assessed for 15 min at PND25. While MK promoted a dose-dependent increase in locomotor activity, exposure to MU failed to elicit significant effects. The combined exposure to the highest dose of MU and the lowest dose of MK induced marked hyperactivity. Moreover, the combination of the low dose of MK and the high dose of MU resulted in a reduced activity in the center of the open field, suggesting an increased anxiety-like behavior. These findings suggest that, during the brain growth spurt, the blockade of NMDA receptors induces juvenile locomotor hyperactivity whereas hyperactivation of GABAA receptors does not. However, GABAA overactivation during this period potentiates the effects of NMDA blockade in inducing locomotor hyperactivity.

Increased apoptosis and reduced neuronal and glial densities in the hippocampus due to nicotine and ethanol exposure in adolescent mice.

It has been recently shown that nicotine and ethanol interact during adolescence affecting memory/learning and anxiety levels. Considering the role of the hippocampus in both anxiety and memory/learning, we investigated whether adolescent nicotine and/or ethanol administration elicit apoptotic cell death and whether this results in neuronal and/or glial density alterations in the following regions of the hippocampus: granular layer of the dentate gyrus (GrDG), molecular layer (Mol), CA1, CA2 and CA3. From the 30th to the 45th postnatal day, C57BL/6 male and female mice were exposed to nicotine free base (NIC) and/or ethanol (ETOH). Four groups were analyzed: (1) concomitant NIC (50mug/ml in 2% saccharin to drink) and ETOH (25%, 2g/kg i.p. injected every other day) exposure; (2) NIC exposure; (3) ETOH exposure; (4) vehicle. We evaluated cell degeneration (TUNEL assay), neuronal and glial densities (optical disector) and region thicknesses at the end of the period of exposure. Our results demonstrate that ETOH elicited an increase in TUNEL-positive cells relative to the vehicle group in all hippocampal regions. NIC elicited less severe region-dependent effects: the number of TUNEL-positive cells was significantly increased in the Mol and CA1 when compared to the vehicle group. These results were paralleled by reductions in neuronal and glial cells densities, which indicate that both cell types are sensitive to the neurotoxic effects of these drugs. There were no effects on region thicknesses. On the other hand, concomitant NIC and ETOH reduced the adverse effects of the drugs when administered separately. This ability of nicotine and ethanol co-exposure to lessen the adverse effects of nicotine and ethanol may contribute to adolescents co-use and co-abuse of tobacco and alcoholic beverages.