Parvalbumin Interneurons and Perineuronal Nets in the Hippocampus and Retrosplenial Cortex of Adult Male Mice After Early Social Isolation Stress and Perinatal NMDA Receptor Antagonist Treatment

Abstract

Both early life aversive experiences and intrinsic alterations in early postnatal neurodevelopment are considered predisposing factors for psychiatric disorders, such as schizophrenia. The prefrontal cortex and the hippocampus have protracted postnatal development and are affected in schizophrenic patients. Interestingly, similar alterations have been observed in the retrosplenial cortex (RSC). Studies in patients and animal models of schizophrenia have found alterations in cortical parvalbumin (PV) expressing interneurons, making them good candidates to study the etiopathology of this disorder. Some of the alterations observed in PV+ interneurons may be mediated by perineuronal nets (PNNs), specialized regions of the extracellular matrix, which frequently surround these inhibitory neurons. In this study, we have used a double hit model (DHM) combining a single perinatal injection of an NMDAR antagonist (MK801) to disturb early postnatal development and post-weaning social isolation as an early life aversive experience. We have investigated PV expressing interneurons and PNNs in the hippocampus and the RSC of adult male mice, using unbiased stereology. In the CA1, but not in the CA3 region, of the hippocampus, the number of PNNs and PV + PNN+ cells was affected by the drug treatment, and a significant decrease of these parameters was observed in the groups of animals that received MK801. The percentage of PNNs surrounding PV+ cells was significantly decreased after treatment in both hippocampal regions; however, the impact of isolation was observed only in CA1, where isolated animals presented lower percentages. In the RSC, we observed significant effects of isolation, MK801 and the interaction of both interventions on the studied parameters; in the DHM, we observed a significantly lower number of PV+, PNNs, and PV+PNN+cells when compared to control mice. Similar significant decreases were observed for the groups of animals that were just isolated or treated with MK801. To our knowledge, this is the first report on such alterations in the RSC in an animal model combining neurodevelopmental alterations and aversive experiences during infancy/adolescence. These results show the impact of early-life events on different cortical regions, especially on the structure and plasticity of PV+ neurons and their involvement in the emergence of certain psychiatric disorders.

Keywords: early aversive experience; interneuron; parvalbumin; schizophrenia; social isolation.

Figure 1

Description of the regions and cells under study. (A) Scheme depicting the location of the different areas under study (modified from Paxinos and Franklin, 2008) (B) Panoramic confocal microphotograph showing the general distribution of PV+ interneurons (red) and PNNs (blue) in a representative section coinciding with the scheme shown in (A). (C) High magnification confocal images from CA1 stratum pyramidale, showing a PV-immunoreactive neuron surrounded by PNNs. Scale bars: 500 μm (B), 20 μm (C).

Figure 2

Analysis of parvalbumin (PV) expressing cells and perineuronal nets (PNNs) in the CA1 region of the hippocampus. (A1–A4) Confocal images from the CA1 showing PV expressing neurons (red), PNNs (blue), and their co-localization (PNN+ PV+). (B–F)Histograms indicating changes in the total numbers of PV+ somata (B), PNNs (C), PV+ cells surrounded by PNNs (D), PNNs not surrounding PV⁺ cells (E), and PV+ cells not surrounded by PNNs (F). (G,H) Histograms indicating the percentages of PNNs surrounding PV+ somata (G) and PV+ somata surrounded by PNNs (H). *p < 0.05; **p < 0.01; ***p < 0.001. Scale bar: 20 μm.

Figure 3

Analysis of parvalbumin (PV) expressing cells and perineuronal nets (PNNs) in the CA3 region of the hippocampus. (A1–A4) Confocal images from the CA3, showing PV expressing neurons (red), PNNs (blue), and their co-localization (PNN+PV+). B-F: Histograms indicating changes in the total numbers of PV+ somata (B), PNNs (C), PV+ cells surrounded by PNNs (D), PNNs not surrounding PV⁺ cells (E), and PV+ cells not surrounded by PNNs (F). G&H: Histograms indicating the percentages of PNNs surrounding PV+ somata (G) and PV+ somata surrounded by PNNs (H). *p < 0.05. Scale bar: 20 μm.

Figure 4

Analysis of parvalbumin (PV) expressing cells and perineuronal nets (PNNs) in the retrosplenial cortex. (A1–A4) Confocal images from the retrosplenial cortex, showing PV expressing neurons (red), PNNs (blue), and their co-localization (PNN+ PV+). (B–F) Histograms indicating changes in the total numbers of PV+ somata (B), PNNs (C), PV+ cells surrounded by PNNs (D), PNNs not surrounding PV⁺ cells (E), and PV+ cells not surrounded by PNNs (F). (G,H) Histograms indicating the percentages of PNNs surrounding PV+ somata (G) and PV+ somata surrounded by PNNs (H). Dashed lines indicate significant interactions and the presence of significant differences after post hoc analysis. *p < 0.05; **p < 0.01; ***p < 0.001. Scale bar: 20 μm.