Unwanted Exacerbation of the Immune Response in Neurodegenerative Disease: A Time to Review the Impact.

The COVID-19 pandemic imposed a series of behavioral changes that resulted in increased social isolation and a more sedentary life for many across all age groups, but, above all, for the elderly population who are the most vulnerable to infections and chronic neurodegenerative diseases. Systemic inflammatory responses are known to accelerate neurodegenerative disease progression, which leads to permanent damage, loss of brain function, and the loss of autonomy for many aged people. During the COVID-19 pandemic, a spectrum of inflammatory responses was generated in affected individuals, and it is expected that the elderly patients with chronic neurodegenerative diseases who survived SARSCoV-2 infection, it will be found, sooner or later, that there is a worsening of their neurodegenerative conditions. Using mouse prion disease as a model for chronic neurodegeneration, we review the effects of social isolation, sedentary living, and viral infection on the disease progression with a focus on sickness behavior and on the responses of microglia and astrocytes. Focusing on aging, we discuss the cellular and molecular mechanisms related to immunosenescence in chronic neurodegenerative diseases and how infections may accelerate their progression.

Virus Infections on Prion Diseased Mice Exacerbate Inflammatory Microglial Response.

We investigated possible interaction between an arbovirus infection and the ME7 induced mice prion disease. C57BL/6, females, 6-week-old, were submitted to a bilateral intrahippocampal injection of ME7 prion strain (ME7) or normal brain homogenate (NBH). After injections, animals were organized into two groups: NBH (n = 26) and ME7 (n = 29). At 15th week after injections (wpi), animals were challenged intranasally with a suspension of Piry arbovirus 0.001% or with NBH. Behavioral changes in ME7 animals appeared in burrowing activity at 14 wpi. Hyperactivity on open field test, errors on rod bridge, and time reduction in inverted screen were detected at 15th, 19th, and 20th wpi respectively. Burrowing was more sensitive to earlier hippocampus dysfunction. However, Piry-infection did not significantly affect the already ongoing burrowing decline in the ME7-treated mice. After behavioral tests, brains were processed for IBA1, protease-resistant form of PrP, and Piry virus antigens. Although virus infection in isolation did not change the number of microglia in CA1, virus infection in prion diseased mice (at 17th wpi) induced changes in number and morphology of microglia in a laminar-dependent way. We suggest that virus infection exacerbates microglial inflammatory response to a greater degree in prion-infected mice, and this is not necessarily correlated with hippocampal-dependent behavioral deficits.

Compartmentalization of cerebral cortical germinal zones in a lissencephalic primate and gyrencephalic rodent.

Previous studies of macaque and human cortices identified cytoarchitectonically distinct germinal zones; the ventricular zone inner subventricular zone (ISVZ), and outer subventricular zone (OSVZ). To date, the OSVZ has only been described in gyrencephalic brains, separated from the ISVZ by an inner fiber layer and considered a milestone that triggered increased neocortical neurogenesis. However, this observation has only been assessed in a handful of species without the identification of the different progenitor populations. We examined the Amazonian rodent agouti (Dasyprocta agouti) and the marmoset monkey (Callithrix jacchus) to further understand relationships among progenitor compartmentalization, proportions of various cortical progenitors, and degree of cortical folding. We identified a similar cytoarchitectonic distinction between the OSVZ and ISVZ at midgestation in both species. In the marmoset, we quantified the ventricular and abventricular divisions and observed similar proportions as previously described for the human and ferret brains. The proportions of radial glia, intermediate progenitors, and outer radial glial cell (oRG) populations were similar in midgestation lissencephalic marmoset as in gyrencephalic human or ferret. Our findings suggest that cytoarchitectonic subdivisions of SVZ are an evolutionary trend and not a primate specific feature, and a large population of oRG can be seen regardless of cortical folding.

Early behavioral changes and quantitative analysis of neuropathological features in murine prion disease: stereological analysis in the albino Swiss mice model.

Behavioral and neuropathological changes have been widely investigated in murine prion disease but stereological based unbiased estimates of key neuropathological features have not been carried out. After injections of ME7 infected (ME7) or normal brain homogenates (NBH) into dorsal CA1 of albino Swiss mice and C57BL6, we assessed behavioral changes on hippocampal-dependent tasks. We also estimated by optical fractionator at 15 and 18 weeks post-injections (w.p.i.) the total number of neurons, reactive astrocytes, activated microglia and perineuronal nets (PN) in the polymorphic layer of dentate gyrus (PolDG), CA1 and septum in albino Swiss mice. On average, early behavioral changes in albino Swiss mice start four weeks later than in C57BL6. Cluster and discriminant analysis of behavioral data in albino Swiss mice revealed that four of nine subjects start to change their behavior at 12 w.p.i. and reach terminal stage at 22 w.p.i and the remaining subjects start at 22 w.p.i. and reach terminal stage at 26 w.p.i. Biotinylated dextran-amine BDA-tracer experiments in mossy fiber pathway confirmed axonal degeneration, and stereological data showed that early astrocytosis, microgliosis and reduction in the perineuronal nets are independent of a change in the number of neuronal cell bodies. Statistical analysis revealed that the septal region had greater levels of neuroinflammation and extracellular matrix damage than CA1. This stereological and multivariate analysis at early stages of disease in an outbred model of prion disease provided new insights connecting behavioral changes and neuroinflammation and seems to be important to understand the mechanisms of prion disease progression.

Enriched environment contributes to recovery of visual acuity and increases perineuronal nets in monocular-deprived animals

The aim of the present study was to analyze the influence of enriched environment on the distribution of perineuronal nets (PNNs) using a stereogically based unbiased protocol and visual acuity in adult Swiss albino mice that underwent monocular deprivation during the critical period of postnatal development. Eight female Swiss albino mice were monocular deprived on postnatal day 10 and divided into two groups at weaning: standard environment (SE group, n = 4) and enriched environment (EE group, n = 4). After 3 months, all of the mice were subjected to grating visual acuity tests, sacrificed, and perfused with aldehyde fixative. The brains were removed and cut at 70 µm thickness in a vibratome and processed for lectin histochemical staining with Wisteria floribunda agglutinin (WFA). Architectonic limits of area 17 were conspicuously defined by WFA histochemical staining, and the optical fractionator stereological method was applied to estimate the total number of PNNs in the supragranular, granular, and infragranular layers. All groups were compared using Student's t-test at a 95 percent confidence level. Comparative analysis of the average PNN estimations revealed that the EE group had higher PNNs in the supragranular layer (2726.33 ± 405.416, mean ± standard deviation) compared with the SE group (1543.535 ± 260.686; Student's t-test, p = .0495). No differences were found in the other layers. Visual acuity was significantly lower in the SE group (0.55 cycles/degree) than in the EE group (1.06 cycles/degree). Our results suggest that the integrity of the specialized extracellular matrix PNNs of the supragranular layer may be essential for normal visual acuity development.(AU)

Enriched environment contributes to recovery of visual acuity and increases perineuronal nets in monocular-deprived animals

The aim of the present study was to analyze the influence of enriched environment on the distribution of perineuronal nets (PNNs) using a stereogically based unbiased protocol and visual acuity in adult Swiss albino mice that underwent monocular deprivation during the critical period of postnatal development. Eight female Swiss albino mice were monocular deprived on postnatal day 10 and divided into two groups at weaning: standard environment (SE group, n = 4) and enriched environment (EE group, n = 4). After 3 months, all of the mice were subjected to grating visual acuity tests, sacrificed, and perfused with aldehyde fixative. The brains were removed and cut at 70 µm thickness in a vibratome and processed for lectin histochemical staining with Wisteria floribunda agglutinin (WFA). Architectonic limits of area 17 were conspicuously defined by WFA histochemical staining, and the optical fractionator stereological method was applied to estimate the total number of PNNs in the supragranular, granular, and infragranular layers. All groups were compared using Student's t-test at a 95 percent confidence level. Comparative analysis of the average PNN estimations revealed that the EE group had higher PNNs in the supragranular layer (2726.33 ± 405.416, mean ± standard deviation) compared with the SE group (1543.535 ± 260.686; Student's t-test, p = .0495). No differences were found in the other layers. Visual acuity was significantly lower in the SE group (0.55 cycles/degree) than in the EE group (1.06 cycles/degree). Our results suggest that the integrity of the specialized extracellular matrix PNNs of the supragranular layer may be essential for normal visual acuity development.

Influence of enriched environment on viral encephalitis outcomes: behavioral and neuropathological changes in albino Swiss mice.

An enriched environment has previously been described as enhancing natural killer cell activity of recognizing and killing virally infected cells. However, the effects of environmental enrichment on behavioral changes in relation to virus clearance and the neuropathology of encephalitis have not been studied in detail. We tested the hypothesis that environmental enrichment leads to less CNS neuroinvasion and/or more rapid viral clearance in association with T cells without neuronal damage. Stereology-based estimates of activated microglia perineuronal nets and neurons in CA3 were correlated with behavioral changes in the Piry rhabdovirus model of encephalitis in the albino Swiss mouse. Two-month-old female mice maintained in impoverished (IE) or enriched environments (EE) for 3 months were behaviorally tested. After the tests, an equal volume of Piry virus (IEPy, EEPy)-infected or normal brain homogenates were nasally instilled. Eight days post-instillation (dpi), when behavioral changes became apparent, brains were fixed and processed to detect viral antigens, activated microglia, perineuronal nets, and T lymphocytes by immuno- or histochemical reactions. At 20 or 40 dpi, the remaining animals were behaviorally tested and processed for the same markers. In IEPy mice, burrowing activity decreased and recovered earlier (8-10 dpi) than open field (20-40 dpi) but remained unaltered in the EEPy group. EEPy mice presented higher T-cell infiltration, less CNS cell infection by the virus and/or faster virus clearance, less microgliosis, and less damage to the extracellular matrix than IEPy. In both EEPy and IEPy animals, CA3 neuronal number remained unaltered. The results suggest that an enriched environment promotes a more effective immune response to clear CNS virus and not at the cost of CNS damage.

Hippocampus and dentate gyrus of the Cebus monkey: architectonic and stereological study.

Behavioral, electrophysiological, and anatomical assays of non-human primates have provided substantial evidence that the hippocampus and dentate gyrus are essential for memory consolidation. However, a single anatomical and stereological investigation of these regions has been done in New World primates to complement those assays. The aim of the present study was to describe the cyto-, myelo-, and histochemical architecture of the hippocampus and dentate gyrus, and to use the optical fractionator method to estimate the number of neurons in the hippocampal pyramidal and granular neurons in the dentate gyrus of the Cebus monkey. NeuN immunolabeling, lectin histochemical staining with Wisteria floribunda agglutinin (WFA), enzyme-histochemical detection of NADPH-diaphorase activity and Gallyas silver staining were used to define the layers and limits of the hippocampal fields and dentate gyrus. A comparative analysis of capuchin (Cebus apella) and Rhesus (Macaca mulatta) monkeys revealed similar structural organization of these regions but significant differences in the regional distribution of neurons. C. apella were found to have 1.3 times fewer pyramidal and 3.5 times fewer granular neurons than M. mulatta. Taken together the architectonic and stereological data of the present study suggest that hippocampal and dentate gyrus neural networks in the C. apella and M. mulatta may contribute to hippocampal-dentate gyrus-dependent tasks in different proportions.

Adult brain nitrergic activity after concomitant prenatal exposure to ethanol and methyl mercury.

Pregnant rats were exposed to ethanol (EtOH) and/or methyl mercury (MeHg) during fetal brain development. Nitrergic activity was quantified by densitometric measurement of formazan deposits in the hippocampus, cerebellum and striatum of two-month-old offspring following histochemical assay for NADPH-diaphorase (NADPH-d) activity. Compared to control subjects, an increase in nitrergic activity was found in the molecular layer of dentate gyrus and in the lacunosum molecular and stratum radiatum of CA1 (cornus amoni 1) in the EtOH+MeHg group, whereas a single administration of EtOH increased the activity in all striatal segments. The cerebellum seems to be less sensitive at this time-point to intoxication, and presented an increase only at the molecular layer of EtOH-exposed animals when compared to the MeHg and EtOH+MeHg groups (ANOVA, one-way followed by Tukey's test, p<0.05 or p<0.01). Taken together, results suggest that developmental exposure to EtOH and MeHg, singularly or in combination, alters nitrergic activity in adult rat in different ways depending on the region and layer of the central nervous system (CNS), and that these alterations might be related to different local metabolic properties.

Number and distribution of neurons in the retinal ganglion cell layer in relation to foraging behaviors of tyrant flycatchers.

The tyrant flycatchers represent a monophyletic radiation of predominantly insectivorous passerine birds that exhibit a plethora of stereotyped prey capture techniques. However, little is known about their retinal organization. Using retinal wholemounts, we estimated the total number and topography of neurons in the ganglion cell layer in the generalist yellow-bellied elaenia (Elaenia flavogaster) and the up-hover-gleaner mouse-colored tyrannulet (Phaeomyias murina) with the optical fractionator method. The mean estimated total number of neurons in the ganglion cell layer was 4,152,416 +/- 189,310 in E. flavogaster and 2,965,132 +/- 354,359 in P. murina. Topographic maps of isocounting lines revealed a similar distribution for both species: a central fovea and a temporal area surrounded by a poorly defined horizontal streak. In addition, both species had increased numbers of giant ganglion cells in the dorsotemporal retina forming an area giganto cellularis. In E. flavogaster, these giant ganglion cells were also distributed across the nasal and ventral retinal peripheries, which is in agreement with the generalist habits of this species. However, in P. murina these cells were rarely seen along the nasal and ventral peripheries, possibly reflecting a lesser need to perceive movement because this species captures stationary insects resting on foliage. Thus, we suggest that the retinas of the tyrant flycatchers in the present study show a general common pattern of neuron distribution in the ganglion cell layer irrespective of their foraging habits. We also suggest that the distribution of giant ganglion cells is indicative of the visual requirements of the species.

Exercise and food ad libitum reduce the impact of early in life nutritional inbalances on nitrergic activity of hippocampus and striatum.

Nutritional imbalances were produced by varying litter size pups per dam: 3 (small), 6 (medium), and 12 (large). On the 21st day, 4 subjects of each litter, were sacrificed and the remaining were grouped, 2 per cage, with or without running wheels, with food and water ad libitum. Adult subjects were tested in water maze, their brains processed for NADPH-diaphorase histochemistry and quantified by densitometry. No differences were detected in water maze. At 21st day, S and L compared with M presented reduced NADPH-d in the stratum molecular of dentate gyrus (DG), stratum lacunosum of CA1 and in all CA3 layers but not in the striatum. On the 58th day, actvity remained low in S and L in CA3 and striatum and L in CA1 and DG. Voluntary exercise increased NADPH-d in DG, CA1, CA3, and striatum in S, and in the stratum lacunosum of CA1 and CA3 in L.

NADPH-diaphorase histochemical changes in the hippocampus, cerebellum and striatum are correlated with different modalities of exercise and watermaze performances.

Nitric oxide is involved in memory and motor learning. We investigated possible influences of exercise on spatial memory and NADPH-diaphorase (NADPH-d) histochemical activity in the hippocampus, striatum and cerebellum. Fifteen albino Swiss mice between the 22nd and 55th post-natal days were exercised in the following modalities: voluntary (V), acrobatic (A), acrobatic/voluntary (AV) and forced (F) and compared to inactive group (I). After the exercise period, all subjects were tested in the water maze for 3 days. Animal brains were processed for NADPH-d histochemistry. Densitometry of the neuropil of the hippocampus, striatum and cerebellum and morphometric analysis of NADPHd+ type I neurons of the striatum were done. Exercise groups presented higher levels of NADPH-d activity in the molecular and polymorphic layers of dentate gyrus and lacunosum molecular layer of CA1. The A group presented higher NADPH-d activity in the cerebellar granular layer than all other groups. Branching points and dendritic segment densities of NADPH-d type I neurons were higher in V, A and AV than in F and I groups. Exercise groups revealed best performances on water maze tests. Thus, different modalities of exercise increases in different proportions for the nitrergic activity in the hippocampus, striatum and cerebellum, and these changes seem to be beneficial to spatial memory.