The possible impairment of respiratory-related neural loops may be associated with the silent pneumonia induced by SARS-CoV-2.

As compared to many other viral pulmonary infections, there existed several peculiar manifestations in the COVID-19 patients, including the "silence" of pneumonia in both mild and severe cases and a long intensive care unit stay for those requiring invasive mechanical ventilation. Similar silent pneumonia has been documented in the infectioninduced by H5N1 influenza virus HK483 and was found to result from the direct attack of the virus on the bronchopulmonary C-fibers at the early stage and the final infection in the brainstem at the late stage. The long stay of critical patients in the intensive care unit is possibly due to the depression of central respiratory drive, which resulted in the failure to wean from the mechanic ventilation. Carotid and aortic bodies and bronchopulmonary C-fibers are two key peripheral components responsible for the chemosensitive responses in the respiratory system, while triggering respiratory reflexes depends predominantly on the putative chemosensitive neurons located in the pontomedullary nuclei. In view of the findings for the H5N1 influenza virus, the silence of pneumonia induced by SARS-CoV-2 may be due to the possible impairment of peripheral chemosensitive reflexes as well as the damage to the respiratory-related central neurons.

Anatomical and physiological foundations of cerebello-hippocampal interaction.

Multiple lines of evidence suggest that functionally intact cerebello-hippocampal interactions are required for appropriate spatial processing. However, how the cerebellum anatomically and physiologically engages with the hippocampus to sustain such communication remains unknown. Using rabies virus as a retrograde transneuronal tracer in mice, we reveal that the dorsal hippocampus receives input from topographically restricted and disparate regions of the cerebellum. By simultaneously recording local field potential from both the dorsal hippocampus and anatomically connected cerebellar regions, we additionally suggest that the two structures interact, in a behaviorally dynamic manner, through subregion-specific synchronization of neuronal oscillations in the 6-12 Hz frequency range. Together, these results reveal a novel neural network macro-architecture through which we can understand how a brain region classically associated with motor control, the cerebellum, may influence hippocampal neuronal activity and related functions, such as spatial navigation.

Zika virus-induced hyper excitation precedes death of mouse primary neuron.

BACKGROUND: Zika virus infection in new born is linked to congenital syndromes, especially microcephaly. Studies have shown that these neuropathies are the result of significant death of neuronal progenitor cells in the central nervous system of the embryo, targeted by the virus. Although cell death via apoptosis is well acknowledged, little is known about possible pathogenic cellular mechanisms triggering cell death in neurons. METHODS: We used in vitro embryonic mouse primary neuron cultures to study possible upstream cellular mechanisms of cell death. Neuronal networks were grown on microelectrode array and electrical activity was recorded at different times post Zika virus infection. In addition to this method, we used confocal microscopy and Q-PCR techniques to observe morphological and molecular changes after infection. RESULTS: Zika virus infection of mouse primary neurons triggers an early spiking excitation of neuron cultures, followed by dramatic loss of this activity. Using NMDA receptor antagonist, we show that this excitotoxicity mechanism, likely via glutamate, could also contribute to the observed nervous system defects in human embryos and could open new perspective regarding the causes of adult neuropathies. CONCLUSIONS: This model of excitotoxicity, in the context of neurotropic virus infection, highlights the significance of neuronal activity recording with microelectrode array and possibility of more than one lethal mechanism after Zika virus infection in the nervous system.

Neurotropism and behavioral changes associated with Zika infection in the vector Aedes aegypti.

Understanding Zika virus infection dynamics is essential, as its recent emergence revealed possible devastating neuropathologies in humans, thus causing a major threat to public health worldwide. Recent research allowed breakthrough in our understanding of the virus and host pathogenesis; however, little is known on its impact on its main vector, Aedes aegypti. Here we show how Zika virus targets Aedes aegypti's neurons and induces changes in its behavior. Results are compared to dengue virus, another flavivirus, which triggers a different pattern of behavioral changes. We used microelectrode array technology to record electrical spiking activity of mosquito primary neurons post infections and discovered that only Zika virus causes an increase in spiking activity of the neuronal network. Confocal microscopy also revealed an increase in synapse connections for Zika virus-infected neuronal networks. Interestingly, the results also showed that mosquito responds to infection by overexpressing glutamate regulatory genes while maintaining virus levels. This neuro-excitation, possibly via glutamate, could contribute to the observed behavioral changes in Zika virus-infected Aedes aegypti females. This study reveals the importance of virus-vector interaction in arbovirus neurotropism, in humans and vector. However, it appears that the consequences differ in the two hosts, with neuropathology in human host, while behavioral changes in the mosquito vector that may be advantageous to the virus.

Resting-state functional magnetic resonance imaging in clade C HIV: within-group association with neurocognitive function.

Neuroimaging abnormalities are common in chronically infected HIV-positive individuals. The majority of studies have focused on structural or functional brain outcomes in samples infected with clade B HIV. While preliminary work reveals a similar structural imaging phenotype in patients infected with clade C HIV, no study has examined functional connectivity (FC) using resting-state functional magnetic resonance imaging (rs-fMRI) in clade C HIV. In particular, we were interested to explore HIV-only effects on neurocognitive function using associations with rs-fMRI. In the present study, 56 treatment-naïve, clade C HIV-infected participants (age 32.27 ± 5.53 years, education 10.02 ± 1.72 years, 46 female) underwent rs-fMRI and cognitive testing. Individual resting-state networks were correlated with global deficit scores (GDS) in order to explore associations between them within an HIV-positive sample. Results revealed ten regions in six resting-state networks where FC inversely correlated with GDS scores (worse performance). The networks affected included three independent attention networks: the default mode network (DMN), sensorimotor network, and basal ganglia. Connectivity in these regions did not correlate with plasma viral load or CD4 cell count. The design of this study is unique and has not been previously reported in clade B. The abnormalities related to neurocognitive performance reported in this study of clade C may reflect late disease stage and/or unique host/viral dynamics. Longitudinal studies will help to clarify the clinical significance of resting-state alterations in clade C HIV.

Motor, cognitive, and affective areas of the cerebral cortex influence the adrenal medulla.

Modern medicine has generally viewed the concept of "psychosomatic" disease with suspicion. This view arose partly because no neural networks were known for the mind, conceptually associated with the cerebral cortex, to influence autonomic and endocrine systems that control internal organs. Here, we used transneuronal transport of rabies virus to identify the areas of the primate cerebral cortex that communicate through multisynaptic connections with a major sympathetic effector, the adrenal medulla. We demonstrate that two broad networks in the cerebral cortex have access to the adrenal medulla. The larger network includes all of the cortical motor areas in the frontal lobe and portions of somatosensory cortex. A major component of this network originates from the supplementary motor area and the cingulate motor areas on the medial wall of the hemisphere. These cortical areas are involved in all aspects of skeletomotor control from response selection to motor preparation and movement execution. The second, smaller network originates in regions of medial prefrontal cortex, including a major contribution from pregenual and subgenual regions of anterior cingulate cortex. These cortical areas are involved in higher-order aspects of cognition and affect. These results indicate that specific multisynaptic circuits exist to link movement, cognition, and affect to the function of the adrenal medulla. This circuitry may mediate the effects of internal states like chronic stress and depression on organ function and, thus, provide a concrete neural substrate for some psychosomatic illness.

Caveats in Transneuronal Tracing with Unmodified Rabies Virus: An Evaluation of Aberrant Results Using a Nearly Perfect Tracing Technique.

Apart from the genetically engineered, modified, strains of rabies virus (RABV), unmodified 'fixed' virus strains of RABV, such as the 'French' subtype of CVS11, are used to examine synaptically connected networks in the brain. This technique has been shown to have all the prerequisite characteristics for ideal tracing as it does not metabolically affect infected neurons within the time span of the experiment, it is transferred transneuronally in one direction only and to all types of neurons presynaptic to the infected neuron, number of transneuronal steps can be precisely controlled by survival time and it is easily detectable with a sensitive technique. Here, using the 'French' CVS 11 subtype of RABV in Wistar rats, we show that some of these characteristics may not be as perfect as previously indicated. Using injection of RABV in hind limb muscles, we show that RABV-infected spinal motoneurons may already show lysis 1 or 2 days after infection. Using longer survival times we were able to establish that Purkinje cells may succumb approximately 3 days after infection. In addition, some neurons seem to resist infection, as we noted that the number of RABV-infected inferior olivary neurons did not progress in the same rate as other infected neurons. Furthermore, in our hands, we noted that infection of Purkinje cells did not result in expected transneuronal labeling of cell types that are presynaptic to Purkinje cells such as molecular layer interneurons and granule cells. However, these cell types were readily infected when RABV was injected directly in the cerebellar cortex. Conversely, neurons in the cerebellar nuclei that project to the inferior olive did not take up RABV when this was injected in the inferior olive, whereas these cells could be infected with RABV via a transneuronal route. These results suggest that viral entry from the extracellular space depends on other factors or mechanisms than those used for retrograde transneuronal transfer. We conclude that transneuronal tracing with RABV may result in unexpected results, as not all properties of RABV seem to be ubiquitously valid.

Intrinsic network connectivity abnormalities in HIV-infected individuals over age 60.

Individuals infected with HIV are living longer due to effective treatment with combination antiretroviral therapy (cART). Despite these advances, HIV-associated neurocognitive disorders (HAND) remain prevalent. In this study, we analyzed resting state functional connectivity (rs-fc) data from HIV-infected and matched HIV-uninfected adults aged 60 years and older to determine associations between HIV status, neuropsychological performance, and clinical variables. HIV-infected participants with detectable plasma HIV RNA exhibited decreased rs-fc within the salience (SAL) network compared to HIV-infected participants with suppressed plasma HIV RNA. We did not identify differences in rs-fc within HIV-infected individuals by HAND status. Our analysis identifies focal deficits in the SAL network that may be mitigated with suppression of plasma virus. However, these findings suggest that rs-fc may not be sensitive as a marker of HAND among individuals with suppressed plasma viral loads.

Revealing the secrets of neuronal circuits with recombinant rabies virus technology.

An understanding of how the brain processes information requires knowledge of the architecture of its underlying neuronal circuits, as well as insights into the relationship between architecture and physiological function. A range of sophisticated tools is needed to acquire this knowledge, and recombinant rabies virus (RABV) is becoming an increasingly important part of this essential toolbox. RABV has been recognized for years for its properties as a synapse-specific trans-neuronal tracer. A novel genetically modified variant now enables the investigation of specific monosynaptic connections. This technology, in combination with other genetic, physiological, optical, and computational tools, has enormous potential for the visualization of neuronal circuits, and for monitoring and manipulating their activity. Here we will summarize the latest developments in this fast moving field and provide a perspective for the use of this technology for the dissection of neuronal circuit structure and function in the normal and diseased brain.

Fortress brain.

Neurodegenerative diseases are associated with neuronal inclusions, comprised of protein aggregates. In Alzheimer's Disease (AD) and Lewy Body Disease (LBD) such lesions are distributed in a hierarchical retrograde transynaptic spatial pattern. This implies a retrograde transynaptic temporal propagation as well. There can be few explanations for this other than infectious agents (prions and viruses). This suggests that AD and LBD (at least) may have infectious origins. Transynaptic infiltration of the CNS along cranial nerve or other major projections, by one or more infectious agents has important implications. The clinical syndrome and natural history of each neurodegenerative disorder will reflect its portal of entry. There may be a different neurodegenerative syndrome for each cranial nerve or other portal of entry, and not all may manifest as "dementia". Each syndrome may be associated with more than one pathological lesion. Each pathology may be associated with several clinical syndromes. Host-parasite interactions are species specific. This may explain the rarity of AD-like pathology in most other older mammals. Over evolutionary timescales, the human brain should be adapted to predation by neurotropic agents. Viewed from this perspective, the prion-like pro-inflammatory and pro-apoptotic properties of ß-amyloid and other proteins may be adaptive, and anti-microbial. Reductions in synaptic density may slow the progress of invading pathogens, while perineuronal nets and other structures may guard the gates. This suggests a defense in depth of a structure, the brain, that is inherently vulnerable to invasion along its neural networks.

The motor cortex communicates with the kidney.

We used retrograde transneuronal transport of rabies virus from the rat kidney to identify the areas of the cerebral cortex that are potential sources of central commands for the neural regulation of this organ. Our results indicate that multiple motor and nonmotor areas of the cerebral cortex contain output neurons that indirectly influence kidney function. These cortical areas include the primary motor cortex (M1), the rostromedial motor area (M2), the primary somatosensory cortex, the insula and other regions surrounding the rhinal fissure, and the medial prefrontal cortex. The vast majority of the output neurons from the cerebral cortex were located in two cortical areas, M1 (68%) and M2 (15%). If the visceromotor functions of M1 and M2 reflect their skeletomotor functions, then the output to the kidney from each cortical area could make a unique contribution to autonomic control. The output from M1 could add precision and organ-specific regulation to descending visceromotor commands, whereas the output from M2 could add anticipatory processing which is essential for allostatic regulation. We also found that the output from M1 and M2 to the kidney originates predominantly from the trunk representations of these two cortical areas. Thus, a map of visceromotor representation appears to be embedded within the classic somatotopic map of skeletomotor representation.

Advances in viral transneuronal tracing.

Powerful transneuronal tracing technologies exploit the ability of some neurotropic viruses to travel across neuronal pathways and to function as self-amplifying markers. Two main classes of viral transneuronal tracers are available, derived from alpha-herpesviruses (Herpes Simplex virus type 1, Pseudorabies) and rabies virus. Depending on the virus type and strain, there are major differences with regard to host range, peripheral uptake, replication mechanisms, transport direction and specificity. While alpha-herpesviruses are the tracers of choice for studying autonomic innervation, rabies virus is the ideal tool for studying motor innervation, since its peripheral uptake occurs exclusively at motor endplates. Rabies virus is the only viral tracer that is entirely specific, as it moves exclusively across chemical synapses by strictly unidirectional (retrograde) transneuronal transfer without altering neuronal metabolism, allowing for the stepwise, time-dependent, identification of neuronal networks across an unlimited number of synapses. This review will highlight and contrast the different properties of these viral tracers, and summarize the methodological issues that are critical for the appropriate execution and interpretation of transneuronal tracing studies. Combinations of viral tracing with other methodologies will be evaluated. Emerging technologies, based on genetically modified herpes and rabies tracers, will be also discussed and put in perspective.

Recent findings on the organization of central nervous system structures involved in the innervation of endocrine glands and other organs; observations obtained by the transneuronal viral double-labeling technique.

This review summarizes the data obtained with the aid of the recently introduced dual viral tracing technique, which uses isogenic recombinants of pseudorabies virus that express unique reporter gene. This approach made possible to explore simultaneously neural circuits of two organs. The results of these studies indicate: (1) there are neurons innervating exclusively a given organ; (2) left-sided predominance in the supraspinal innervation of the endocrine glands (adrenal, ovary) studied, so far; (3) viral co-infection of neurons, i.e., special neuronal populations coexist in different brain areas that are transsynaptically connected with both paired endocrine and non-endocrine organs, endocrine glands and non-endocrine organs, and organs of bodily systems other than the endocrine one. The number of common neurons seems to be related to the need of coordinating action of different systems. The data on co-infection of neurons suggest that the central nervous system has the capacity to coordinate different organ functions via common brain neurons providing supraspinal innervation of the organs.

Loss of connexin36 in rat hippocampus and cerebellar cortex in persistent Borna disease virus infection.

Neonatal Borna disease virus (BDV) infection of the Lewis rat leads to progressive degeneration of dentate gyrus granule cells, and cerebellar Purkinje neurons. Our aim here was to clarify whether BDV interfered with the formation of electrical synapses, and we, therefore, analysed expression of the neuronal gap junction protein connexin36 (Cx36) in the Lewis rat hippocampal formation, and cerebellar cortex, 4 and 8 weeks after neonatal infection. Semiquantitative RT-PCR, revealed a BDV-dependent decrease in Cx36 mRNA in the hippocampal formation 4 and 8 weeks post-infection (p.i.), and in the cerebellar cortex 8 weeks p.i. Correspondingly, immunofluorescent staining revealed reduced Cx36 immunoreactivity in both dentate gyrus, and ammons horn CA3 region, 4 and 8 weeks post-infection. In the cerebellar cortex, Cx36 immunoreactivity was detected only 8 weeks post-infection in the molecular layer, where it was down regulated by BDV. Our findings demonstrate, for the first time, distinct BDV-dependent reductions in Cx36 mRNA and protein in the rat hippocampal formation and cerebellar cortex, suggesting altered neuronal network properties to be an important feature of persistent viral brain infections.

Use of rabies virus as a transneuronal tracer of neuronal connections: implications for the understanding of rabies pathogenesis.

In neurosciences, rabies virus (CVS strain) has become a very powerful tool for studying multisynaptic neuronal connections, due to its ability to function as a self-replicating marker and to propagate exclusively between connected neurons by transneuronal transfer, which is strictly time-dependent. In this laboratory, transneuronal tracing studies of rabies virus propagation in primates and rodent models during the asymptomatic period have provided valuable information on rabies pathogenesis. We have shown that rabies virus propagates by fast axonal transport at similar speeds in primates and rodents, after inoculation into the peripheral or central nervous system (CNS). Intracellulartransport of rabies virus is preferentially addressed to neuronal dendrites rather than axons, since transneuronal transfer occurs only retrogradely, i.e., from dendrites of first infected neurons to presynaptic terminals of connected neurons. Rabies virus propagation occurs at chemical synapses, but not via gap junctions or local spread. The results of our studies show that rabies virus receptors have a ubiquitous distribution on neurons within the CNS. Conversely, in the peripheral nervous system, rabies virus receptors are present only on motor endings, since uptake is restricted to motor endplates and axons, whereas sensory and autonomic endings are not infected. Thus, after peripheral inoculations, motoneurons are the only gateway for rabies virus transmission to the CNS. Infection of sensory and autonomic neurons requires longer incubation times, since it reflects centrifugal propagation of rabies virus from the CNS to the periphery, i.e., it is the result of retrograde transneuronal transfer to sensory and autonomic terminals within the CNS.

The human immunodeficiency virus reduces network capacity: acoustic noise effect.

OBJECTIVE: Increased acoustic noise (AN) during working memory leads to increased brain activation in healthy individuals and may have greater impact in human immunodeficiency virus (HIV) patients. RESULTS: Compared with control subjects, HIV patients showed reduced AN activation and lower neuronal marker N-acetylaspartate in prefrontal and parietal cortices. Competing use of the working memory network between AN and cognitive load showed lower dynamic range of the hemodynamic responses in prefrontal and parietal cortices in HIV patients. INTERPRETATION: These findings suggest that reduced reserve capacity of the working memory network in HIV patients and additional stress (eg, AN) might exhaust the impaired network for more demanding tasks.

Adaptation of the attention network in human immunodeficiency virus brain injury.

Human immunodeficiency virus (HIV)-positive patients commonly have attention and concentration problems. However, it remains unclear how HIV infection affects the attention network. Therefore, blood oxygenation level dependent functional magnetic resonance imaging (BOLD-fMRI) was performed in 36 subjects (18 HIV and 18 seronegative [SN] controls) during a set of visual attention tasks with increasing levels of attentional load. Compared with SN controls, HIV subjects showed similar task performance (accuracies and reaction times) but decreased activation in the normal visual attention network (dorsal parietal, bilateral prefrontal, and cerebellar regions) and increased activation in adjacent or contralateral brain regions. Cognitive performance (assessed with NPZ-8), CD4, and viral load all correlated with activated BOLD signals in brain regions that activated more in HIV subjects. Furthermore, HIV subjects activated more than SN controls in brain regions that showed load-dependent increase in activation (right prefrontal and right parietal regions) but less in regions that showed a saturation effect with increasing load. These findings suggest that HIV-associated brain injury leads to reduced efficiency in the normal attention network, thus requiring reorganization and increased usage of neural reserves to maintain performance during attention-requiring tasks. Exceeding the brain reserve capacity may lead to attention deficits and cognitive impairment in HIV patients.

Suprachiasmatic nucleus input to autonomic circuits identified by retrograde transsynaptic transport of pseudorabies virus from the eye.

Intraocular injection of the Bartha strain of pseudorabies virus (PRV Bartha) results in transsynaptic infection of the hypothalamic suprachiasmatic nucleus (SCN), a retinorecipient circadian oscillator. PRV Bartha infection of a limited number of retinorecipient structures, including the SCN, was initially interpreted as the differential infection of a subpopulation of rat retinal ganglion cells, followed by replication and anterograde transport via the optic nerve. A recent report that used a recombinant strain of PRV Bartha (PRV152) expressing enhanced green fluorescent protein demonstrated that SCN infection actually results from retrograde transneuronal transport of the virus via the autonomic innervation of the eye in the golden hamster. In the present study using the rat, the pattern of infection after intravitreal inoculation with PRV152 was examined to determine if infection of the rat SCN is also restricted to retrograde transsynaptic transport. It was observed that infection in preganglionic autonomic nuclei (i.e., Edinger-Westphal nucleus, superior salivatory nucleus, and intermediolateral nucleus) precedes infection in the SCN. Sympathetic superior cervical ganglionectomy did not abolish label in the SCN after intraocular infection, nor did lesions of parasympathetic preganglionic neurons in the Edinger-Westphal nucleus. However, combined Edinger-Westphal nucleus ablation and superior cervical ganglionectomy eliminated infection of the SCN. This observation allowed a detailed examination of the SCN contribution to descending autonomic circuits afferent to the eye. The results indicate that in the rat, as in the hamster, SCN infection after intraocular PRV152 inoculation is by retrograde transsynaptic transport via autonomic pathways to the eye.