Type I interferon signaling limits reoviral tropism within the brain and prevents lethal systemic infection

In vivo and ex vivo models of reoviral encephalitis were utilized to delineate the contribution of type I interferon (IFN) to the host’s defense against local central nervous system (CNS) viral infection and systemic viral spread. Following intracranial (i.c.) inoculation with either serotype 3 (T3) or serotype 1 (T1) reovirus, increased expression of IFN-α, IFN-β, and myxovirus-resistance protein (Mx1; a prototypical IFN stimulated gene) was observed in mouse brain tissue. Type I IFN receptor deficient mice (IFNAR^−/−) had accelerated lethality, compared to wildtype (B6wt) controls, following i.c. T1 or T3 challenge. Although viral titers in the brain and eyes of reovirus infected IFNAR^−/− mice were significantly increased, these mice did not develop neurologic signs or brain injury. In contrast, increased reovirus titers in peripheral tissues (liver, spleen, kidney, heart, and blood) of IFNAR^−/− mice were associated with severe intestinal and liver injury. These results suggest that reovirus-infected IFNAR^−/− mice succumb to peripheral disease rather than encephalitis per se. To investigate the potential role of type I IFN in brain tissue, brain slice cultures (BSCs) were prepared from IFNAR^−/− mice and B6wt controls for ex vivo T3 reovirus infection. Compared to B6wt controls, reoviral replication and virus-induced apoptosis were enhanced in IFNAR^−/− BSCs indicating that a type I IFN response, initiated by resident CNS cells, mediates innate viral immunity within the brain. T3 reovirus tropism was extended in IFNAR^−/− brains to include dentate neurons, ependymal cells, and meningeal cells indicating that reovirus tropism within the CNS is dependent upon type I interferon signaling.     

A brain slice culture model of viral encephalitis reveals an innate CNS cytokine response profile and the therapeutic potential of caspase inhibition

Viral encephalitis is a significant cause of human morbidity and mortality in large part due to suboptimal diagnosis and treatment. Murine reovirus infection serves as a classic experimental model of viral encephalitis. Infection of neonatal mice with T3 reoviruses results in lethal encephalitis associated with neuronal infection, apoptosis, and CNS tissue injury. We have developed an ex vivo brain slice culture (BSC) system that recapitulates the basic pathological features and kinetics of viral replication seen in vivo. We utilize the BSC model to identify an innate, brain-tissue specific inflammatory cytokine response to reoviral infection, which is characterized by the release of IL6, CXCL10, RANTES, and murine IL8 analog (KC). Additionally, we demonstrate the potential utility of this system as a pharmaceutical screening platform by inhibiting reovirus-induced apoptosis and CNS tissue injury with the pan-caspase inhibitor, Q-VD-OPh. Cultured brain slices not only serve to model events occurring during viral encephalitis, but can also be utilized to investigate aspects of pathogenesis and therapy that are not experimentally accessible in vivo.     

Minocycline delays disease onset and mortality in reovirus encephalitis

Minocycline is neuroprotective in many experimental models of neurodegenerative diseases and central nervous system (CNS) injury but has not previously been tested in a model of viral encephalitis. Experimental infection of neonatal mice with neurotropic reoviruses is a classic model for studying the pathogenesis of viral encephalitis. Intracerebral inoculation of serotype 3 reovirus strain Dearing (T3D) in neonatal mice results in lethal encephalitis caused by neuronal apoptosis throughout the CNS. Minocycline significantly delayed death in mice to 11.6 +/- 0.9 days post-infection vs. 8.6 +/- 0.7 days post-infection in controls (P < 0.01). Virus-induced CNS injury, apoptosis, viral titer and antigen expression were significantly decreased in the brains of minocycline-treated mice on 6 and 8 days post-infection compared to controls. Virus-induced injury and viral titer in minocycline-treated infected mice at 11 days post-infection were similar to those seen in untreated T3D-infected mice at 8 days post-infection. Little microglial or astrocytic invasion of brain regions with viral injury was found at any time-point in untreated or minocycline-treated mice, suggesting that in this model system the neuroprotective effect exerted by minocycline is more likely due to its anti-apoptotic properties rather than its capacity to inhibit microglial activation and limit gliosis. These findings, similar to those reported for neurodegenerative diseases, indicate that minocycline does not prevent development of fatal reovirus encephalitis but delays disease onset and progression, suggesting that minocycline treatment may provide a useful adjunctive therapy in viral CNS infections.     

Reovirus infection of the CNS enhances iNOS expression in areas of virus-induced injury

Nitric oxide (NO) has been implicated as a contributor to the host's innate defense against viral infections including those affecting the CNS. Reovirus infection of the CNS is a classic experimental system for understanding the pathogenesis of neurotropic viral infection. Infection with serotype 3 strains is associated with perturbations in various cellular signaling pathways including NF-kappaB and NO plays a regulatory role in many of these same pathways. We therefore examined whether NO production is dysregulated following reovirus serotype 3 strain Abney (T3A) infection of the mouse CNS. Nitric oxide synthase (NOS) activity was significantly higher in brain homogenates from T3A-infected animals compared to mock infected. Increased NOS activity correlated with inducible NOS (iNOS) expression in brain homogenates of T3A-infected animals. Expression of iNOS was confined to areas of viral infection and injury. T3A infection of primary neuronal and glial cultures was also associated with enhanced expression of iNOS. Immunocytochemical studies of primary glial cultures demonstrated that, in addition to its known neuronotropism, T3A was also capable of infecting immature microglial cells. T3A infection did not alter expression of either neuronal or endothelial NOS isoforms in neuronal or glial cultures or in mouse brain. The NO donor S-Nitroso-N-acetyl penicillamine (SNAP) significantly inhibited T3A growth in neuronal cultures, conversely the NOS inhibitor N-omega-Nitro-L-arginine methyl ester hydrochloride (L-NAME) augmented viral growth. Our findings provide the first evidence of reovirus-induced iNOS expression and the first demonstration that NO inhibits mammalian reovirus replication, suggesting that NO may play an antiviral role during reovirus infection.     

Poliovirus type 1 infection of murine <Emphasis Type="Italic">PRNP</Emphasis>-knockout neuronal cells

Transfection of the prion protein gene (Prnp) into prion-deficient mouse cells was shown to reduce the replication of coxsackievirus B3, an enterovirus. Because mice can be susceptible to poliovirus infection by parenteral routes, the authors tested the susceptibility to poliovirus-1 (PV-1) of a panel of murine neuronal cell lines differing in their ability to express Prnp. The investigated cell lines (prionless HpL3.4 cells, HpL3.4 cells transfected with a Prnp vector, HpL3.4 cells transfected with a void vector, wild-type Hw3.5 Prnp ^+/+ cells) expressed the murine homologue (Tage4) of human poliovirus receptor (CD155/hPVR). PV-1 infection of Prnp ^−/− HpL3.4 cells resulted in the production of high viral titers, though viral antigens could be detected in only 0.5% to 2% of cells. Wild-type Prnp ^+/+ cells and prionless cells transfected with the Prnp gene were not permissive to PV-1. Results of viral titration and immunofluorescence were confirmed by conventional polymerase chain reaction (PCR) and quantitative real-time PCR. Exposure to PV-1 had no influence on the gene expression profile of Prnp ^+/+ cells. In contrast, PV-1 infection was associated with up regulation of several genes in permissive Prnp ^−/− cell cultures: type I interferon (IFN) genes, IFN-related developmental regulator 1 (IFNRD1), tumor necrosis factor superfamily member 13b (TNFSF13b), interleukin (IL) − 7, granulocyte/macrophage colony-stimulating factors (CSFs), hepatocyte growth factor (HGF), vascular endothelial growth factor-A, transforming growth factors beta1 and beta3 (TGFb1, TGFb3), as well as a variety of bone morphogenetic proteins endowed with neuroprotective activity. Distinction of permissive from nonpermissive neuronal cells on the basis of Prnp expression suggests that prion-deficient mice could represent an extraordinarily sensitive animal model for poliovirus infection.     

Reovirus-induced neuronal apoptosis is mediated by caspase 3 and is associated with the activation of death receptors

Reovirus infection of the central nervous system (CNS) is an important experimental system for understanding the pathogenesis of neurotropic viral infection. Infection of neonatal mice with T3 reoviruses causes lethal encephalitis in which injury results from virus-induced apoptosis. We now show that this apoptosis in vivo is associated with activation of caspase 3, and use neuroblastoma and primary neuronal cultures to identify the cellular pathways involved. Reovirus-induced apoptosis in neuronal cultures is initiated by activation of the tumor necrosis factor (TNF) receptor superfamily death receptors and is inhibited by treatment with soluble death receptors (DRs). The DR-associated initiator caspase, caspase 8, is activated following infection, this activation is inhibited by a cell-permeable peptide inhibitor (IETD-CHO). In contrast to our previous findings in non-neuronal cell lines, reovirus-induced neuronal apoptosis is not accompanied by significant release of cytochrome c from the mitochondria or with caspase 9 activation following infection. This suggests that in neuronal cells, unlike their non-neuronal counterparts, the mitochondria-mediated apoptotic pathway associated with cytochrome c release and caspase 9 activation does not play a significant role in augmenting reovirus-induced apoptosis. Consistent with these results, peptide caspase inhibitors show a hierarchy of efficacy in inhibiting reovirus-induced apoptosis, with inhibitors of caspase 3 > caspase 8 > caspase 9. These studies provide a comprehensive profile of the pattern of virus-induced apoptotic pathway activation in neuronal culture.     

The type I interferon response bridles rabies virus infection and reduces pathogenicity

Rabies virus (RABV) is a neurotropic virus transmitted by the bite of an infected animal that triggers a fatal encephalomyelitis. During its migration in the nervous system (NS), RABV triggers an innate immune response, including a type I IFN response well known to limit viral infections. We showed that although the neuroinvasive RABV strain CVS-NIV dampens type I IFN signaling by inhibiting IRF3 phosphorylation and STAT2 translocation, an early and transient type I IFN response is still triggered in the infected neuronal cells and NS. This urged us to investigate the role of type I IFN on RABV infection. We showed that primary mouse neurons (DRGs) of type I IFN(α/β) receptor deficient mice (IFNAR^−/− mice) were more susceptible to RABV than DRGs of WT mice. In addition, exogenous type I IFN is partially efficient in preventing and slowing down infection in human neuroblastoma cells. Intra-muscular inoculation of type I IFNAR deficient mice [IFNAR^−/− mice and NesCre (^+/−) IFNAR (^flox/flox) mice lacking IFNAR in neural cells of neuroectodermal origin only] with RABV reveals that the type I IFN response limits RABV dissemination in the inoculated muscle, slows down invasion of the spinal cord, and delays mortality. Thus, the type I IFN which is still produced in the NS during RABV infection is efficient enough to reduce neuroinvasiveness and pathogenicity and partially protect the host from fatal infection.     

Cerebellar Long-term Depression is Deficient in Niemann�CPick Type C Disease Mice

Niemann–Pick type C disease (NPC) is an autosomal recessive lipidosis characterized by progressive neurodegeneration. Although several studies have revealed unusual accumulation of unesterfied cholesterol in astrocytic lysosome of NPC, pathophysiological basis of cerebellar neuronal dysfunction remains unclear. We compared parallel fiber-Purkinje cell synaptic transmission and long-term depression (LTD) in +/+npc ^nih (npc ^+/+) and −/−npc ^nih (npc ^−/−) mice. Our data showed that adenosine A1 receptor agonists decreased parallel fiber excitatory postsynaptic current (EPSC) amplitude and mEPSC frequency while its antagonists increased EPSC amplitude and mEPSC frequency in wild type and mutant mice. Furthermore, parallel fiber LTD was deficient in npc ^−/− mice and supplement of adenosine triphosphate (ATP) rescued the impaired LTD. Taken together, these experiments suggest that synaptic strength and LTD are altered in npc ^−/− mice due to the decrease of ATP/adenosine release and deactivation of A1 receptors in parallel fiber terminals. The enhanced synaptic transmission and the decreased LTD might result in progressive neurotoxicity of Purkinje cells in npc ^−/− mice.     

Intracellular calcium-binding protein S100A4 influences injury-induced migration of white matter astrocytes

Astrocytes play a crucial role in central nervous system (CNS) pathophysiology. White and gray matter astrocytes are regionally specialized, and likely to respond differently to CNS injury and in CNS disease. We previously showed that the calcium-binding protein S100A4 is exclusively expressed in white matter astrocytes and markedly up-regulated after injury. Furthermore, down-regulation of S100A4 in vitro significantly increases the migration capacity of white matter astrocytes, a property, which might influence their function in CNS tissue repair. Here, we performed a localized injury (scratch) in confluent cultures of white matter astrocytes, which strongly express S100A4, and in cultures of white matter astrocytes, in which S100A4 was down-regulated by transfection with short interference (si) S100A4 RNA. We found that S100A4-silenced astrocytes rapidly migrated into the injury gap, whereas S100A4-expressing astrocytes extended hypertrophied processes toward the gap, but without closing it. To explore the involvement of S100A4 in migration of astrocytes in vivo, we induced focal demyelination and transient glial cell elimination in the spinal cord white matter by ethidium bromide injection in S100A4 (−/−) and (+/+) mice. The results show that astrocyte migration into the demyelinated area is promoted in S100A4 (−/−) compared to (+/+) mice, in which a pronounced glial scar was formed. These data indicate that S100A4 reduces the migratory capacity of reactive white matter astrocytes in the injured CNS and is involved in glial scar formation after injury.     

Dual lentivirus infection potentiates neuroinflammation and neurodegeneration: viral copassage enhances neurovirulence

Infection by multiple lentiviral strains is recognized as a major driving force in the human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS) epidemic, but the neuropathogenic consequences of multivirus infections remain uncertain. Herein, we investigated the neurovirulence and underlying mechanisms of dual lentivirus infections with distinct viral strains. Experimental feline immunodeficiency virus (FIV) infections were performed using cultured cells and an in vivo model of AIDS neuropathogenesis. Dual infections were comprised of two FIV strains (FIV-Ch and FIV-PPR) as copassaged or superinfected viruses, with subsequent outcome analyses of host immune responses, viral load, neuropathological features, and neurobehavioral performance. Dual infections of feline macrophages resulted in greater IL-1β (interleukin-1b), TNF-α (tumor necrosis factor α), and IDO (indoleamine 2,3-dioxygenase) expression and associated neurotoxic properties. FIV coinfection and sequential superinfection in vivo also induced greater IL-1β, TNF-α, and IDO expression in the basal ganglia (BG) and cortex (CTX), compared to the monovirus- and mock-infected groups, although viral loads were similar in single virus and dual virusinfected animals. Immunoblot analyses disclosed lower synaptophysin immunoreactivity in the CTX resulting from FIV super- and coinfections. Cholinergic and GABAergic neuronal injury was evident in the CTX of animals with dual FIV infections. With increased glial activation and neuronal loss in dual FIV infected brains, immunohistochemical analysis also revealed elevated detection of cleaved caspase-3 in dysmorphic neurons, which was associated with worsened neurobehavioral abnormalities among animals infected with the copassaged viruses. Dual lentivirus infections caused an escalation in neuroinflammation and ensuing neurodegeneration, underscoring the contribution of infection by multiple viruses to neuropathogenesis.     

Gene expression in the brain during reovirus encephalitis

Viral encephalitis remains a significant cause of morbidity and mortality throughout the world. We performed microarray analysis to identify genes and pathways that are differentially regulated during reovirus encephalitis and that may provide novel therapeutic targets for virus-induced diseases of the central nervous system (CNS). An increase in the expression of 130 cellular genes was found in the brains of reovirus-infected mice at early times post infection, compared to mock-infected controls. The up-regulation of these genes was consistent with activation of innate immune responses, particularly interferon signaling. At later times post infection, when significant CNS injury is present and mice exhibit signs of severe neurologic disease, many more (1374) genes were up-regulated, indicating that increased gene expression correlates with disease pathology. Virus-induced gene expression at late times post infection was again consistent with the activation of innate immune responses. However, additional significant pathways included those associated with cytokine signaling and apoptosis, both of which can contribute to CNS injury. This is the first report comparing virus-induced cellular gene and pathway regulation at early and late times following virus infection of the brain. The shift of virus-induced gene expression from innate immune responses at early times post infection to cytokine signaling and apoptosis at later times suggests a potential therapeutic strategy that preserves early protective responses whilst inhibiting later responses that contribute to pathogenesis.     

Motor neuronal loss and neurofilament-ubiquitin alteration in MoMuLV-ts1 encephalopathy

A temperature-sensitive mutant of Moloney murine leukemia virus (MoMuLV-ts1) induces immunosuppression and spongiform encephalopathy in susceptible newborn mice. The associated neuronal degeneration primarily involves the motor neurons in specific target areas of the central nervous system (CNS). Neuronal loss occurs in the absence of direct viral infection of neurons and is the most dramatic pathological change in the CNS of infected mice. To quantitatively demonstrate neuronal loss, an unbiased morphometric stereological study was undertaken using the optical disector method. Using highly susceptible FVB/N mice, neuronal loss was quantitated in the tissue sections of brain stem from infected and noninfected mice at 20 and 35 days post inoculation (dpi). Results indicated that there was no significant neuronal loss at 20 dpi, but significant (P < 0.05) at 35 dpi. In addition, histology, transmission electron microscopy and immunohistochemistry revealed Lewy body-like inclusions consisting of aggregates of neurofilaments and cellular organelles. Degenerated neurons and glial cells were heavily ubiquitinated. Together, these results suggest that significant neuronal loss occurs at the end of the disease process and that Lewy body-like formation and protein ubiquitination are part of the pathogenic process in ts1-induced encephalopathy. Received: 8 June 1999 / Revised, accepted: 2 August 1999     

Ghrelin Antagonized 1-Methyl-4-Phenylpyridinium (MPP<Superscript>+</Superscript>)-Induced Apoptosis in MES23.5 Cells

Ghrelin is an endogenous ligand for the growth hormone secretagogue receptor (GHS-R) acting to stimulate growth hormone release. In the previous study, we have observed the neuroprotective effects of ghrelin on dopaminergic neurons in vivo in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine -treated Parkinson’s disease mice. In order to illustrate the underlying mechanisms, in the present study, we conducted our experiment in vitro in 1-methyl-4-phenylpyridinium (MPP⁺)-treated MES23.5 cells that could express GHS-R1a. Ten- to 1,000-μmol/L MPP⁺ treatment caused decreased cell viability, with increased lactate dehydrogenase leakage. A 200-μmol/L MPP⁺ treatment was chosen to do the further experiments. MES23.5 cells treated with 200 μmol/L MPP⁺ showed decreased mitochondrial transmembrane potential, an elevated level of reactive oxidative species production and activation of caspase-3. Additionally, these cells also showed apoptotic morphological changes. Pretreatment with different doses of ghrelin (10⁻¹²–10⁻⁷ mol/L) could abolish the MPP⁺-induced apoptotic changes in a dose-dependent manner. These results suggested that ghrelin could antagonize MPP⁺-induced apoptosis in MES23.5 cells. The protective effects of ghrelin involved the restoration of mitochondria function. Juanjuan Dong and Ning Song contributed equally to this work.     

Neuropathology of wild-type and <Emphasis Type="Italic">nef</Emphasis>-attenuated T cell tropic simian immunodeficiency virus (SIVmac32H) and macrophage tropic neurovirulent SIVmac17E-Fr in cynomolgus macaques

The neuropathology of simian immunodeficiency (SIV) infection in cynomolgus macaques (Macaca fascicularis) was investigated following infection with either T cell tropic SIVmacJ5, SIVmacC8 or macrophage tropic SIVmac17E-Fr. Formalin fixed, paraffin embedded brain tissue sections were analysed using a combination of in situ techniques. Macaques infected with either wild-type SIVmacJ5 or neurovirulent SIVmac17E-Fr showed evidence of neuronal dephosphorylation, loss of oligodendrocyte and CCR5 staining, lack of microglial MHC II expression, infiltration by CD4⁺ and CD8⁺ T cells and mild astrocytosis. SIVmacJ5-infected animals exhibited activation of microglia whilst those infected with SIVmac17E-Fr demonstrated a loss of microglia staining. These results are suggestive of impaired central nervous system (CNS) physiology. Furthermore, infiltration by T cells into the brain parenchyma indicated disruption of the blood brain barrier (BBB). Animals infected with the Δnef-attenuated SIVmacC8 showed microglial activation and astrogliosis indicative of an inflammatory response, lack of MHC II and CCR5 staining and infiltration by CD8⁺ T cells. These results demonstrate that the SIV infection of cynomolgus macaque can be used as a model to replicate the range of CNS pathologies observed following HIV infection of humans and to investigate the pathogenesis of HIV associated neuropathology.     

CSF neurofilament protein (NFL) — a marker of active HIV-related neurodegeneration

Abstract Background and methods The light subunit of the neurofilament protein (NFL), a major structural component of myelinated axons, is a sensitive indicator of axonal injury in the central nervous system (CNS) in a variety of neurodegenerative disorders. Cerebrospinal fluid (CSF) NFL concentrations were measured by ELISA (normal < 250 ng/l) in archived samples from 210 HIV-infected patients not taking antiretroviral treatment: 55 with AIDS dementia complex (ADC), 44 with various CNS opportunistic infections/tumours (CNS OIs), 95 without neurological symptoms or signs, and 16 with primary HIV infection (PHI). The effect of highly active antiretroviral treatment (HAART) was studied by repeated CSF sampling in four of the ADC patients initiating treatment. Results CSF NFL concentrations were significantly higher in patients with ADC (median 2590 ng/l, IQR 780–7360) and CNS OIs (2315 ng/l, 985–7390 ng/l) than in neuroasymptomatic patients (<250 ng/l, <250–300) or PHI (<250 ng/l, <250–280), p < 0.001. Among patients with ADC, those with more severe disease (stage 2–4) had higher levels than those with milder disease (stage 0.5–1), p < 0.01. CSF NFL declined during HAART to the limit of detection in parallel with virological response and neurological improvement in ADC.CSF NFL concentrations were higher in neuroasymptomatic patients with lower CD4-cell strata than higher, p < 0.001. This increase was less marked than in the ADC patients and noted in 26/58 neuroasymptomatic patients with CD4 counts <200/μl compared to 1/37 with CD4-cells ≥200/μl. Conclusions The findings of this study support the value of CSF NFL as a useful marker of ongoing CNS damage in HIV infection. Markedly elevated CSF NFL concentrations in patients without CNS OIs are associated with ADC, follow the grade of severity, and decrease after initiation of effective antiretroviral treatment. Nearly all previously suggested CSF markers of ADC relate to immune activation or HIV viral load that do not directly indicate brain injury. By contrast NFL is a sensitive marker of such injury, and should prove useful in evaluating the presence and activity of ongoing CNS injury in HIV infection. Drs. Abdulle and Mellgren contributed equally to this work.     

Roles of GABA<Subscript>B</Subscript> receptor subtypes in presynaptic auto- and heteroreceptor function regulating GABA and glutamate release

Γ-Aminobutyric acid B (GABA_B) receptors are heterodimers composed of two subunits GABA_B(1) and GABA_B(2), the former existing in two isoforms GABA_B(1a) and GABA_B(1b). The contributions of individual receptor subunits and isoforms to GABA_B auto- and heteroreceptor functions were investigated, using release experiments in cortical slice preparations from corresponding knockout mice. Presynaptic GABA_B autoreceptors are located on GABAergic terminals and inhibit GABA release, whereas presynaptic GABA_B heteroreceptors control the release of other neurotransmitters (e.g. glutamate). Neither baclofen nor the selective antagonist CGP55845 at maximally active concentrations affected [³H]GABA release in slices from GABA_B(1)−/− mice. The amount of [³H]GABA released per pulse was unaffected by the stimulation frequency in slices from GABA_B(1)−/− and GABA_B(2)−/− demonstrating a loss of GABA_B autoreceptor function in these knockout animals. The GABA_B receptor agonist baclofen was ineffective in modulating glutamate release in cortical slices from GABA_B(2)−/− mice, showing that heteroreceptor function was abolished as well. Next we investigated knockout mice for the two predominant GABA_B(1) isoforms expressed in brain, GABA_B(1a) and GABA_B(1b). In cortical, hippocampal and striatal slices from both GABA_B(1a)−/− and GABA_B(1b)−/− mice, the frequency dependence of [³H]GABA released per pulse was maintained, suggesting that both isoforms participate or can substitute for each other in GABA_B autoreceptor function. By contrast, the efficacy of baclofen to inhibit glutamate release was substantially reduced in GABA_B(1a)−/−, but essentially unaltered in GABA_B(1b)−/− mice. Our data suggest that functional GABA_B heteroreceptors regulating glutamate release are predominantly, but not exclusively composed of GABA_B(1a) and GABA_B(2) subunits. An erratum to this article can be found at     

Behavior of mice with mutations in the conserved region deleted in velocardiofacial/DiGeorge syndrome

Velocardiofacial/DiGeorge syndrome (VCFS/DGS) is a developmental disorder caused by a 1.5 to 3-Mb hemizygous 22q11.2 deletion. VCFS/DGS patients display malformations in multiple systems, as well as an increased frequency of neuropsychiatric defects including schizophrenia. Haploinsufficiency of TBX1 appears to be responsible for these physical malformations in humans and mice, but the genes responsible for the neuropsychiatric defects are unknown. In this study, two mouse models of VCFS/DGS, a deletion mouse model (Lgdel/+) and a single gene model (Tbx1 +/−), as well as a third mouse mutant (Gscl −/−) for a gene within the Lgdel deletion, were tested in a large behavioral battery designed to assess gross physical features, sensorimotor reflexes, motor activity nociception, acoustic startle, sensorimotor gating, and learning and memory. Lgdel/+ mice contain a 1.5-Mb hemizygous deletion of 27 genes in the orthologous region on MMU 16 and present with impairment in sensorimotor gating, grip strength, and nociception. Tbx1 +/− mice were impaired in grip strength similar to Lgdel/+ mice and movement initiation. Gscl −/− mice were not impaired in any of the administered tests, suggesting that redundant function of other Gsc family members may compensate for the loss of Gscl. Thus, although deletion of the genes in the Lgdel region in mice may recapitulate some of the behavioral phenotypes seen in humans with VCFS/DGS, these phenotypes are not found in mice with complete loss of Gscl or in mice with heterozygous loss of Tbx1, suggesting that the neuropsychiatric and physical malformations of VCFS/DGS may act by different genetic mechanisms.     

AMPA receptor function is altered in GLUR2-deficient mice

The GluR2 subunit of the α-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid (AMPA) receptor determines many of the biophysical properties of native AMPA receptors, including Ca⁺⁺ permeability. Genetically engineered mice unable to edit the Q to R site of the GluR2 subunit die within 3 wk postpartum, presumably due to toxicity associated with enhanced Ca⁺⁺ influx through AMPA receptors. In contrast, disruption of the gene encoding GluR2 is not necessarily lethal. The objective of this study was to explore potential mechanisms that permit survival of GluR2 (−/−) mice despite AMPA receptors that are highly Ca⁺⁺ permeable. Whole-cell, patch-clamp recording of AMPA receptor responses in cortical pyramidal cells revealed that the kinetics of recovery from desensitization were significantly slower for receptors from GluR2 (−/−) mice compared to receptors from GluR2 (+/+) mice. The recovery time constants for AMPA receptors from GluR2 (−/−) and GluR2 (+/+) mice were 109.8±17 ms and 54.4±7.1 ms, respectively. The slower recovery kinetics would be expected to reduce Ca⁺⁺ influx during repetitive stimulation. Because both RNA editing at the R/G site and alternative splicing of the flip and flp module affect AMPA receptor desensitization recovery rates, the possibility that these mechanisms were changed in GluR2 (−/−) mice was investigated. On a macroscopic level, neither editing nor splicing of the GluR-1, 3 or 4 subunits were changed in GluR2 (−/−) mice compared to GluR2 (+/+) mice. In summary, an increase in the time constant for recovery from desensitization may contribute to the ability of GluR2 (−/−) to survive.