Neurodevelopmental Processes in the Prefrontal Cortex Derailed by Chronic HIV-1 Viral Protein Exposure.

Due to the widespread access to, and implementation of, combination antiretroviral therapy, individuals perinatally infected with human immunodeficiency virus type 1 (HIV-1) are living into adolescence and adulthood. Perinatally infected adolescents living with HIV-1 (pALHIV) are plagued by progressive, chronic neurocognitive impairments; the pathophysiological mechanisms underlying these deficits, however, remain understudied. A longitudinal experimental design from postnatal day (PD) 30 to PD 180 was utilized to establish the development of pyramidal neurons, and associated dendritic spines, from layers II-III of the medial prefrontal cortex (mPFC) in HIV-1 transgenic (Tg) and control animals. Three putative neuroinflammatory markers (i.e., IL-1ß, IL-6, and TNF-α) were evaluated early in development (i.e., PD 30) as a potential mechanism underlying synaptic dysfunction in the mPFC. Constitutive expression of HIV-1 viral proteins induced prominent neurodevelopmental alterations and progressive synaptodendritic dysfunction, independent of biological sex, in pyramidal neurons from layers II-III of the mPFC. From a neurodevelopmental perspective, HIV-1 Tg rats exhibited prominent deficits in dendritic and synaptic pruning. With regards to progressive synaptodendritic dysfunction, HIV-1 Tg animals exhibited an age-related population shift towards dendritic spines with decreased volume, increased backbone length, and decreased head diameter; parameters associated with a more immature dendritic spine phenotype. There was no compelling evidence for neuroinflammation in the mPFC during early development. Collectively, progressive neuronal and dendritic spine dysmorphology herald synaptodendritic dysfunction as a key neural mechanism underlying chronic neurocognitive impairments in pALHIV.

Curcumin protects rat hippocampal neurons against pseudorabies virus by regulating the BDNF/TrkB pathway.

Pseudorabies virus (PRV) infection can elicit nervous system disorders. Curcumin has been reported to have neuroprotective effects. However, whether curcumin can protect neurons against PRV infection and the underlying mechanisms remain unclear. In the present study, for the first time, the protective effects of curcumin against PRV-induced oxidative stress, apoptosis, and mitochondrial dysfunction in rat hippocampal neurons and the brain-derived neurotrophic factor (BDNF)/tropomyosin-related kinase B (TrkB) pathway were investigated. Results indicated that PRV with a titer of 3.06 × 106 TCID50 (50% tissue culture infective dose) induced oxidative damage of hippocampal neurons 2 h post-infection and that 10 µM curcumin improved the viability of PRV-infected hippocampal neurons. Blocking the BDNF/TrkB pathway reversed the neuroprotective effects of curcumin, which were imparted by decreasing the PRV-induced upregulation of nitric oxide synthase expression, repressing the PRV-activated mitochondrial apoptotic pathway, and mitochondrial dysfunction. To conclude, curcumin exhibited a neuroprotective role against PRV infection by upregulating the BDNF/TrkB pathway. This study provides insight into the anti-PRV neuroprotective application of curcumin and the underlying mechanism in the prophylaxis and treatment of neurological disorders caused by PRV infection.

Disruption of Timing: NeuroHIV Progression in the Post-cART Era.

The marked increase in life expectancy for HIV-1 seropositive individuals, following the great success of combination antiretroviral therapy (cART), heralds an examination of the progression of HIV-1 associated neurocognitive disorders (HAND). However, since the seminal call for animal models of HIV-1/AIDS in 1988, there has been no extant in vivo animal model system available to provide a truly longitudinal study of HAND. Here, we demonstrate that the HIV-1 transgenic (Tg) rat, resembling HIV-1 seropositive individuals on lifelong cART, exhibits age-related, progressive neurocognitive impairments (NCI), including alterations in learning, sustained attention, flexibility, and inhibition; deficits commonly observed in HIV-1 seropositive individuals. Pyramidal neurons from layers II-III of the medial prefrontal cortex (mPFC) displayed profound synaptic dysfunction in HIV-1 Tg animals relative to controls; dysfunction that was characterized by alterations in dendritic branching complexity, synaptic connectivity, and dendritic spine morphology. NCI and synaptic dysfunction in pyramidal neurons from layers II-III of the mPFC independently identified the presence of the HIV-1 transgene with at least 78.5% accuracy. Thus, even in the absence of sensory or motor system deficits and comorbidities, HAND is a neurodegenerative disease characterized by age-related disease progression; impairments which may be due, at least partly, to synaptic dysfunction in the mPFC. Further, the progression of HAND with age in the HIV-1 Tg rat and associated synaptic dysfunction affords an instrumental model system for the development of therapeutics and functional cure strategies.

IL-10 expression in pyramidal neurons after neuropathogenic coronaviral infection.

The apoptosis of pyramidal neurons in CA2 and CA3 subregions of the hippocampus is induced after infection with Mu-3 virus (Mu-3), a neuropathogenic strain of the JHM virus (JHMV), at 4-5 days post-inoculation (dpi). The viral antigens in the hippocampus are mainly found in the CD11b-positive cells distributed in the stratum oriens located outside the pyramidal layer, and only a few pyramidal neurons are infected. Furthermore, the apoptotic cells, indicated as showing caspase 3 (Cas3) activation, consist of a high number of uninfected cells. Therefore, it is considered that the apoptotic lesions occur through the indirect effects of infection, and not as a result of direct infection with Mu-3, similar to the reported neuronal apoptosis in the hippocampus after other types of infection. The apoptosis in the pyramidal neurons is accompanied by various types of proinflammatory cytokines depending on the causative agents. Thus, the local expression of proinflammatory cytokines was studied, revealing no correlation in the distribution of cytokine expression with the subregions showing apoptosis. However, the anti-inflammatory cytokine IL-10 was produced by pyramidal neurons of CA2 and CA3 at 3 dpi when there is no destructive change or viral invasion in the hippocampus.

Identification of suitable reference genes for BDV-infected primary rat hippocampal neurons.

Borna disease virus (BDV) is a neurotropic RNA virus that infects the limbic system of mammals and results in behavioral disorders. The hippocampus is a core region in the limbic system, which contributes to memory and learning and is important in the regulation of emotion. However, no validated microRNA housekeeping genes have yet been identified in BDV­infected rat primary hippocampal neurons. Proper normalization is key in accurate miRNA expression analysis. The present study used reverse transcription­quantitative polymerase chain reaction (RT­qPCR) to evaluate the expression stability of 10 commonly used reference genes [miR­92a, 5S, U6, miR­103, miR­101a, miR-let-7a, miR­16, E2 small nucleolar RNA (snoRNA), U87 and miR­191] in BDV­infected rat hippocampal neurons and non­infected controls across 12 days post­infection. The data was analyzed by four statistical algorithms: geNorm, NormFinder, BestKeeper, and the comparative Δ­Ct method. Subsequently, the most suitable reference genes (miR­101a and U87) and the least suitable (snoRNA) were determined by the RankAggreg package. miR­155 was selected as a standard by which to evaluate the most and least suitable reference genes. When normalized to the most stable reference gene there were significant differences between the two groups. However, when the data were normalized to the less stably expressed gene, the results were not significant. miR­101a was recommended as a suitable reference gene for BDV-infected rat primary hippocampal neurons.

Combined chronic blockade of hyper-active L-type calcium channels and NMDA receptors ameliorates HIV-1 associated hyper-excitability of mPFC pyramidal neurons.

Human Immunodeficiency Virus type 1 (HIV-1) infection induces neurological and neuropsychological deficits, which are associated with dysregulation of the medial prefrontal cortex (mPFC) and other vulnerable brain regions. We evaluated the impact of HIV infection in the mPFC and the therapeutic potential of targeting over-active voltage-gated L-type Ca(2+) channels (L-channel) and NMDA receptors (NMDAR), as modeled in HIV-1 transgenic (Tg) rats. Whole-cell patch-clamp recording was used to assess the membrane properties and voltage-sensitive Ca(2+) potentials (Ca(2+) influx) in mPFC pyramidal neurons. Neurons from HIV-1 Tg rats displayed reduced rheobase, spike amplitude and inwardly-rectifying K(+) influx, increased numbers of action potentials, and a trend of aberrant firing compared to those from non-Tg control rats. Neuronal hyper-excitation was associated with abnormally-enhanced Ca(2+) influx (independent of NMDAR), which was eliminated by acute L-channel blockade. Combined chronic blockade of over-active L-channels and NMDARs with open-channel blockers abolished HIV effects on spiking, aberrant firing and Ca(2+) potential half-amplitude duration, though not the reduced inward rectification. In contrast, individual chronic blockade of over-active L-channels or NMDARs did not alleviate HIV-induced mPFC hyper-excitability. These studies demonstrate that HIV alters mPFC neuronal activity by dysregulating membrane excitability and Ca(2+) influx through the L-channels. This renders these neurons more susceptible and vulnerable to excitatory stimuli, and could contribute to HIV-associated neuropathogenesis. Combined targeting of over-active L-channels/NMDARs alleviates HIV-induced dysfunction of mPFC pyramidal neurons, emphasizing a potential novel therapeutic strategy that may effectively decrease HIV-induced Ca(2+) dysregulation in the mPFC.

HIV-1 Tat-Mediated Calcium Dysregulation and Neuronal Dysfunction in Vulnerable Brain Regions.

Despite the success of combined antiretroviral therapy, more than half of HIV-1-infected patients in the USA show HIV-associated neurological and neuropsychiatric deficits. This is accompanied by anatomical and functional alterations in vulnerable brain regions of the mesocorticolimbic and nigrostriatal systems that regulate cognition, mood and motivation-driven behaviors, and could occur at early stages of infection. Neurons are not infected by HIV, but HIV-1 proteins (including but not limited to the HIV-1 trans-activator of transcription, Tat) induce Ca(2+) dysregulation, indicated by abnormal and excessive Ca(2+) influx and increased intracellular Ca(2+) release that consequentially elevate cytosolic free Ca(2+) levels ([Ca(2+)]in). Such alterations in intracellular Ca(2+) homeostasis significantly disturb normal functioning of neurons, and induce dysregulation, injury, and death of neurons or non-neuronal cells, and associated tissue loss in HIV-vulnerable brain regions. This review discusses certain unique mechanisms, particularly the over-activation and/or upregulation of the ligand-gated ionotropic glutamatergic NMDA receptor (NMDAR), the voltage-gated L-type Ca(2+) channel (L-channel) and the transient receptor potential canonical (TRPC) channel (a non-selective cation channel that is also permeable for Ca(2+)), which may underlie the deleterious effects of Tat on intracellular Ca(2+) homeostasis and neuronal hyper-excitation that could ultimately result in excitotoxicity. This review also seeks to provide summarized information for future studies focusing on comprehensive elucidation of molecular mechanisms underlying the pathophysiological effects of Tat (as well as some other HIV-1 proteins and immunoinflammatory molecules) on neuronal function, particularly in HIV-vulnerable brain regions.

HIV-1 Transgenic Rat Prefrontal Cortex Hyper-Excitability is Enhanced by Cocaine Self-Administration.

The medial prefrontal cortex (mPFC) is dysregulated in HIV-1-infected humans and the dysregulation is enhanced by cocaine abuse. Understanding mPFC pathophysiology in this comorbid state has been hampered by the dearth of relevant animal models. To help fill this knowledge gap, electrophysiological assessments were made of mPFC pyramidal neurons (PN) from adult male HIV-1 transgenic (Tg) F344 rats (which express seven of the nine HIV-1 toxic proteins) and non-Tg F344 rats that self-administered cocaine for 14 days (COC-SA), as well as saline-yoked controls (SAL-Yoked) and experimentally naive Tg and non-Tg rats. Forebrain slices were harvested and prepared for whole-cell patch-clamp recording, and in treated rats, this occurred after 14-18 days of forced abstinence. Aged-matched rats were used for immunohistochemical detection of the L-channel protein, Cav1.2-α1c. We determined that: (i) the two genotypes acquired the operant task and maintained similar levels of COC-SA, (ii) forced abstinence from COC-SA enhanced mPFC PN excitability in both genotypes, and neurons from Tg rats exhibited the greatest pathophysiology, (iii) neurons from SAL-Yoked Tg rats were more excitable than those from SAL-Yoked non-Tg rats, and in Tg rats (iv) blockade of L-type Ca(2+) channels reduced the enhanced excitability, and (v) Cav1.2-immunoreactivity was increased. These findings provide the first assessment of the mPFC pathophysiology in a rodent model of HIV-1-mediated neuropathology with and without cocaine self-administration. Outcomes reveal an enhanced cortical excitability during chronic exposure to HIV-1 proteins that is excessively exacerbated with cocaine abuse. Such neuropathophysiology may underlie the cognitive dysregulation reported for comorbid humans.

JC virus encephalopathy is associated with a novel agnoprotein-deletion JCV variant.

JC virus encephalopathy (JCVE) is a newly described gray matter disease of the brain caused by productive infection of cortical pyramidal neurons. We characterized the full length sequence of JCV isolated from the brain of a JCVE patient, analyzed its distribution in various compartments by PCR, and determined viral gene expression in the brain by immunohistochemistry(IHC). We identified a novel JCV variant, JCV(CPN1), with a unique 143 bp deletion in the Agno gene encoding a truncated 10 amino acid peptide, and harboring an archetype-like regulatory region. This variant lacked one of three nuclear protein binding regions in the Agno gene. It was predominant in the brain, where it coexisted with an Agno-intact wild-type strain. Double immunostaining with anti-Agno and anti- VP1 antibodies demonstrated that the truncated JCV(CPN1) Agno peptide was present in the majority of cortical cells productively infected with JCV. We then screened 68 DNA samples from 8 brain, 30 CSF and 30 PBMC samples of PML patients, HIV+ and HIV- control subjects. Another JCV(CPN) strain with a different pattern of Agno-deletion was found in the CSF of an HIV+/PML patient, where it also coexisted with wild-type, Agno-intact JCV. These findings suggest that the novel tropism for cortical pyramidal neurons of JCV(CPN1), may be associated with the Agno deletion. Productive and lytic infection of these cells, resulting in fulminant JCV encephalopathy and death may have been facilitated by the co-infection with a wild-type strain of JCV.

Apoptosis of hippocampal pyramidal neurons is virus independent in a mouse model of acute neurovirulent picornavirus infection.

Many viruses, including picornaviruses, have the potential to infect the central nervous system (CNS) and stimulate a neuroinflammatory immune response, especially in infants and young children. Cognitive deficits associated with CNS picornavirus infection result from injury and death of neurons that may occur due to direct viral infection or during the immune responses to virus in the brain. Previous studies have concluded that apoptosis of hippocampal neurons during picornavirus infection is a cell-autonomous event triggered by direct neuronal infection. However, these studies assessed neuron death at time points late in infection and during infections that lead to either death of the host or persistent viral infection. In contrast, many neurovirulent picornavirus infections are acute and transient, with rapid clearance of virus from the host. We provide evidence of hippocampal pathology in mice acutely infected with the Theiler's murine encephalomyelitis picornavirus. We found that CA1 pyramidal neurons exhibited several hallmarks of apoptotic death, including caspase-3 activation, DNA fragmentation, and chromatin condensation within 72 hours of infection. Critically, we also found that many of the CA1 pyramidal neurons undergoing apoptosis were not infected with virus, indicating that neuronal cell death during acute picornavirus infection of the CNS occurs in a non-cell-autonomous manner. These observations suggest that therapeutic strategies other than antiviral interventions may be useful for neuroprotection during acute CNS picornavirus infection.

Brain lesions induced by experimental intranasal infection of Japanese encephalitis virus in piglets.

Non-suppurative encephalitis was experimentally induced in three-week-old piglets by a single intranasal inoculation of Japanese encephalitis virus (JEV) isolated from field pigs. The lesions consisted of glial cell aggregates and perivascular cuffing throughout the olfactory tract and pyriform cortex. JEV antigens were detected in the cytoplasm and neuronal processes of small nerve cells in the granule cell layer of the olfactory bulb, in the neuronal processes of the olfactory tract and in the cytoplasm of neurons in the pyriform cortex. The distribution of the antigens corresponded closely with the distribution of brain lesions. These findings suggest that JEV may enter the brain by the olfactory pathway in addition to via haematogenous spread in piglets.

Fulminant JC virus encephalopathy with productive infection of cortical pyramidal neurons.

The polyomavirus JC (JCV) is the causative agent of progressive multifocal leukoencephalopathy and of JCV granule cell neuronopathy. We present a human immunodeficiency virus-negative patient who experienced development of multiple cortical lesions, aphasia, and progressive cognitive decline after chemotherapy for non-small-cell lung cancer. Brain biopsy and cerebrospinal fluid polymerase chain reaction demonstrated JCV, and she had a rapidly fatal outcome. Postmortem analysis showed diffuse cortical lesions and areas of necrosis at the gray-white junction. Immunostaining showed a productive JCV infection of cortical pyramidal neurons, confirmed by electron microscopy, with limited demyelination. This novel gray matter syndrome expands the scope of JCV clinical presentation and pathogenesis.

Fusion of microglia with pyramidal neurons after retroviral infection.

The neurogenic potential of the postnatal neocortex has not been tested previously with a combination of both retroviral and bromodeoxyuridine (BrdU) labeling. Here we report that injections of enhanced green fluorescent protein (eGFP) retrovirus into 134 postnatal rats resulted in GFP labeling of 642 pyramidal neurons in neocortex. GFP-labeled neocortical pyramidal neurons, however, unlike GFP-labeled glia, did not incorporate BrdU. Closer inspection of retrovirally labeled neurons revealed microglia fused to the apical dendrites of labeled pyramidal neurons. Retroviral infection of mixed cultures of cortical neurons and glia confirmed the presence of specific neuronal-microglial fusions. Microglia did not fuse to other glial cell types, and cultures not treated with retrovirus lacked microglial-neuronal fusion. Furthermore, activation of microglia by lipopolysaccharide greatly increased the virally induced fusion of microglia to neurons in culture. These results indicate a novel form of specific cell fusion between neuronal dendrites and microglia and further illustrate the need for caution when interpreting evidence for neuronogenesis in the postnatal brain.

Herpes simplex virus type 1 inoculation enhances hippocampal excitability and seizure susceptibility in mice.

Herpes simplex virus type 1 (HSV-1) is the major pathogen related to epilepsy. However, little is known about the pathogenesis of HSV-1-associated epilepsy. Here, we report that corneal inoculation of mice with HSV-1 induces acute spontaneous behavioural and electrophysiological seizures and chronically increases hippocampal excitability and seizure susceptibility. In slices from infected mice, the surviving hippocampal CA3 pyramidal neurons exhibited a more depolarizing resting membrane potential concomitant with an increase in membrane input resistance. They also had a lower threshold for generating synchronized bursts and a decrease in the amplitude of afterhyperpolarization (AHP) than did controls. These results suggest that a direct change in the excitability of the hippocampal CA3 neuronal network could play an important role in facilitating the development of acute seizures and subsequent epilepsy.

Brain malformations caused by retroviral vector-mediated gene transfer of hippocampal cholinergic neurostimulating peptide precursor protein into the CNS via embryonic mice ventricles.

Hippocampal cholinergic neurostimulating peptide precursor protein (HCNP-pp) is a unique multifunctional protein, being not only the precursor of HCNP, which promotes the phenotype development of septo-hippocampal cholinergic neurons, but also the binding protein of phosphatidylethanolamine, ATP, Raf-1 kinase (known as "Raf-1 kinase inhibitory factor" in peripheral organs), and serine protease. We obtained a high-titer retroviral vector harboring HCNP-pp cDNA by the use of a modified packaging cell line and centrifugation, and by injecting it into embryonic mouse ventricles, we investigated the function of its gene product within the central nervous system (CNS). We found that efficient transduction into hippocampal pyramidal neurons can be achieved by injecting the vector into embryonic brain ventricles on embryonic day 14 (E14). Three days after receiving the intraventricular injection of the high-titer HCNP-pp retrovirus vector on E14, the tissues around the ventricles showed an overexpression of HCNP-pp. This was accompanied by a reduced amount of activated MEK and Erk (as analyzed by histochemical and Western blot methods), suggesting that HCNP-pp also regulates the MAP-kinase cascade within the CNS. Surprisingly, mouse brains that received the HCNP-pp retroviral vector showed massive malformation of the hippocampus and cerebellum when examined 30 days after birth. This shows that strictly regulated HCNP-pp gene expression is necessary for the normal development of the mouse brain, and that the moderate overexpression achieved by retroviral vector-mediated gene transfer is sufficient to cause severe abnormality of entire brain structures.

Distribution of adenoviral vector in brain after intravenous administration.

The delivery of transgenes to the central nervous system (CNS) can be a valuable tool to treat CNS diseases. Various systems for the delivery to the CNS have been developed; vascular delivery of viral vectors being most recent. Here, we investigated gene transfer to the CNS by intravenous injection of recombinant adenoviral vectors, containing green fluorescence protein (GFP) as a reporter gene. Expression of GFP was first observed 6 days after the gene transfer, peaked at 14 days, and almost diminished after 28 days. The observed expression of GFP in the CNS was highly localized to hippocampal CA regions of cerebral neocortex, inferior colliculus of midbrain, and granular cell and Purkinje cell layers of cerebellum. It is concluded that intravenous delivery of adenoviral vectors can be used for gene delivery to the CNS, and hence the technique could be beneficial to gene therapy.

A novel neurotropic expression vector based on the avirulent A7(74) strain of Semliki Forest virus.

Semliki Forest virus (SFV), an enveloped alphavirus of the family Togaviridae, infects a wide range of mammalian host cells. Most strains are neurotropic but differ in virulence. The authors took advantage of the nonpathogenic properties of SFV strain A7(74), cloned recently in their laboratory, and constructed a replication-proficient expression vector to target the central nervous system (CNS) for heterologous gene expression. The vector, termed VA7, was engineered to drive expression of foreign inserts through a second subgenomic promoter inserted in the viral 3' nontranslated region (NTR). Infectious virus was obtained by in vitro transcription and transfection into BHK cells, and was shown to direct synthesis of heterologous proteins in several mammalian cell lines. Although novel expression vehicle is not applicable for targeting specific cell populations within the CNS in its present form, in cultured rat hippocampal slices, VA7 encoding enhanced green fluorescent protein (EGFP) efficiently transduced pyramidal cells, interneurons, and glial cells. With prolonged time post infection, the number of EGFP-expressing neurons in hippocampal slices increased. Mice infected intraperitoneally with the recombinant virus remained completely asymptomatic but showed CNS expression of EGFP as evidenced by immunohistochemistry. SFV A7(74) is a nonintegrating virus, which gives rise to a randomly distributed, patchy infection of the adult CNS that is cleared within 10 days. With the advantage of noninvasive administration, the expression vector described in this work is thus applicable for short-term gene expression in the CNS.

Semliki Forest virus A7(74) transduces hippocampal neurons and glial cells in a temperature-dependent dual manner.

In central nervous system (CNS) tissue preparations, wild-type Semliki Forest virus (SFV) mainly infects neurons, and in vivo it causes lethal encephalitis in neonatal and adult rodents. The SFV strain A7(74), by contrast, is avirulent in adult rodents, triggering only limited CNS infection. To examine A7(74) infection in hippocampal tissue, the authors constructed a replicon, termed SFV(A774nsP)-GFP, expressing green fluorescent protein. The results were compared to replication-proficient recombinant A7(74) encoding GFP, named VA7-EGFP. As nonstructural gene mutations can confer temperature sensitivity, the authors also tested whether infection was temperature-dependent. Indeed, at 31 degrees C both viral recombinants transduced significantly more baby hamster kidney cells than at 37 degrees C. When rat hippocampal slices and dissociated cells were incubated at 37 degrees C, SFV(A774nsP)-GFP transduced glial cells but virtually no neurons-the opposite of conventional SFV. For VA7-EGFP at 37 degrees C, the preferred GFP-positive cells in hippocampal slices were also non-neuronal cells. At 31 degrees C, however, a more wild-type phenotype was found, with 33% and 94% of the GFP-positive cells being neurons for SFV(A774nsP)-GFP in slices and dissociated cells, respectively, and 94% neurons for VA7-EGFP in slices. Immunochemical and electrophysiological analyses confirmed that at 37 degrees C virtually all cells transduced by SFV(A774nsP)-GFP in slices were astrocytes, while at 31 degrees C they also contained neurons. These results show that in addition to the developmental age, the temperature determines which cell type becomes infected by A7(74). Our data suggest that A7(74) is avirulent in adult animals because it does not readily replicate in mature neurons at body temperature, whereas it still does so at lower temperatures.

Distribution of Adenoviral Vector in Brain after Intravenous Administration

The delivery of transgenes to the central nervous system (CNS) can be a valuable tool to treat CNS diseases. Various systems for the delivery to the CNS have been developed; vascular delivery of viral vectors being most recent. Here, we investigated gene transfer to the CNS by intravenous injection of recombinant adenoviral vectors, containing green fluorescence protein (GFP) as a reporter gene. Expression of GFP was first observed 6 days after the gene transfer, peaked at 14 days, and almost diminished after 28 days. The observed expression of GFP in the CNS was highly localized to hippocampal CA regions of cerebral neocortex, inferior colliculus of midbrain, and granular cell and Purkinje cell layers of cerebellum. It is concluded that intravenous delivery of adenoviral vectors can be used for gene delivery to the CNS, and hence the technique could be beneficial to gene therapy.

Prenatal viral infection leads to pyramidal cell atrophy and macrocephaly in adulthood: implications for genesis of autism and schizophrenia.

We investigated the role of maternal exposure to human influenza virus (H1N1) in C57BL/6 mice on Day 9 of pregnancy on pyramidal and nonpyramidal cell density, pyramidal nuclear area, and overall brain size in Day 0 neonates and 14-week-old progeny and compared them to sham-infected cohorts. Pyramidal cell density increased significantly (p < 0.0038) by 170% in Day 0 infected mice vs. controls. Nonpyramidal cell density decreased by 33% in Day 0 infected progeny vs. controls albeit, nonsignificantly. Pyramidal cell nuclear size decreased significantly (p < 0.0465) by 29% in exposed newborn mice vs. controls. Fourteen-week-old exposed mice continued to show significant increases in both pyramidal and nonpyramidal cell density values vs. controls respectively (p < 0.0085 E1 (exposed group 1), p < 0.0279 E2 (exposed group 2) pyramidal cell density; p < 0.0092 E1, p < 0.0252 E2, nonpyramidal cell density). By the same token, pyramidal cell nuclear size exhibited 37-43% reductions when compared to control values; these were statistically significant vs. controls (p < 0.04 E1, p < 0.0259 E2). Brain and ventricular area measurements in adult exposed mice also showed significant increases and decreases respectively vs. controls. Ventricular brain ratios exhibited 38-50% decreases in exposed mice vs. controls. While the rate of pyramidal cell proliferation per unit area decreased from birth to adulthood in both control and exposed groups, nonpyramidal cell growth rate increased only in the exposed adult mice. These data show for the first time that prenatal exposure of pregnant mice on Day 9 of pregnancy to a sublethal intranasal administration of influenza virus has both short-term and long-lasting deleterious effects on developing brain structure in the progeny as evident by altered pyramidal and nonpyramidal cell density values; atrophy of pyramidal cells despite normal cell proliferation rate and final enlargement of brain. Moreover, abnormal corticogenesis is associated with development of abnormal behavior in the exposed adult mice.