A20/TNFAIP3 heterozygosity predisposes to behavioral symptoms in a mouse model for neuropsychiatric lupus.

Background: Neuropsychiatric lupus (NPSLE) refers to the neurological and psychiatric manifestations that are commonly observed in patients with systemic lupus erythematosus (SLE). An important question regarding the pathogenesis of NPSLE is whether the symptoms are caused primarily by CNS-intrinsic mechanisms or develop as a consequence of systemic autoimmunity. Currently used spontaneous mouse models for SLE have already contributed significantly to unraveling how systemic immunity affects the CNS. However, they are less suited when interested in CNS primary mechanisms. In addition, none of these models are based on genes that are associated with SLE. In this study, we evaluate the influence of A20, a well-known susceptibility locus for SLE, on behavior and CNS-associated changes in inflammatory markers. Furthermore, given the importance of environmental triggers for disease onset and progression, the influence of an acute immunological challenge was evaluated. Methods: Female and male A20 heterozygous mice (A20+/-) and wildtype littermates were tested in an extensive behavioral battery. This was done at the age of 10±2weeks and 24 â€‹± â€‹2 weeks to evaluate the impact of aging. To investigate the contribution of an acute immunological challenge, LPS was injected intracerebroventricularly at the age of 10±2weeks followed by behavioral analysis. Underlying molecular mechanisms were evaluated in gene expression assays on hippocampus and cortex. White blood cell count and blood-brain barrier permeability were analyzed to determine whether peripheral inflammation is a relevant factor. Results: A20 heterozygosity predisposes to cognitive symptoms that were observed at the age of 10 â€‹± â€‹2 weeks and 24 â€‹± â€‹2 weeks. Young A20+/- males and females showed a subtle cognitive phenotype (10±2weeks) with distinct neuroinflammatory phenotypes. Aging was associated with clear neuroinflammation in female A20+/- mice only. The genetic predisposition in combination with an environmental stimulus exacerbates the behavioral impairments related to anxiety, cognitive dysfunction and sensorimotor gating. This was predominantly observed in females. Furthermore, signs of neuroinflammation were solely observed in female A20+/- mice. All above observations were made in the absence of peripheral inflammation and of changes in blood-brain barrier permeability, thus consistent with the CNS-primary hypothesis. Conclusions: We show that A20 heterozygosity is a predisposing factor for NPSLE. Further mechanistic insight and possible therapeutic interventions can be studied in this mouse model that recapitulates several key hallmarks of the disease.

Specific suppression of microgliosis cannot circumvent the severe neuropathology in peroxisomal ß-oxidation-deficient mice.

An important hallmark of various neurodegenerative disorders is the proliferation and activation of microglial cells, the resident immune cells of the central nervous system (CNS). Mice that lack multifunctional protein-2 (MFP2), the key enzyme in peroxisomal ß-oxidation, develop excessive microgliosis that positively correlates with behavioral deficits whereas no neuronal loss occurs. However, the precise contribution of neuroinflammation to the fatal neuropathology of MFP2 deficiency remains largely unknown. Here, we first attempted to suppress the inflammatory response by administering various anti-inflammatory drugs but they failed to reduce microgliosis. Subsequently, Mfp2-/- mice were treated with the selective colony-stimulating factor 1 receptor (CSF1R) inhibitor PLX5622 as microglial proliferation and survival is dependent on CSF1R signaling. This resulted in the elimination of >95% of microglia from control mice but only 70% of the expanded microglial population from Mfp2-/- mice. Despite microglial diminution in Mfp2-/- brain, inflammatory markers remained unaltered and residual microglia persisted in a reactive state. CSF1R inhibition did not prevent neuronal dysfunction, cognitive decline and clinical deterioration of Mfp2-/- mice. Collectively, the unaltered inflammatory profile despite suppressed microgliosis concurrent with persevering clinical decline strengthens our hypothesis that neuroinflammation importantly contributes to the Mfp2-/- phenotype.

A2A adenosine receptor deletion is protective in a mouse model of Tauopathy.

Consumption of caffeine, a non-selective adenosine A2A receptor (A2AR) antagonist, reduces the risk of developing Alzheimer's disease (AD) in humans and mitigates both amyloid and Tau burden in transgenic mouse models. However, the impact of selective A2AR blockade on the progressive development of AD-related lesions and associated memory impairments has not been investigated. In the present study, we removed the gene encoding A2AR from THY-Tau22 mice and analysed the subsequent effects on both pathological (Tau phosphorylation and aggregation, neuro-inflammation) and functional impairments (spatial learning and memory, hippocampal plasticity, neurotransmitter profile). We found that deleting A2ARs protect from Tau pathology-induced deficits in terms of spatial memory and hippocampal long-term depression. These effects were concomitant with a normalization of the hippocampal glutamate/gamma-amino butyric acid ratio, together with a global reduction in neuro-inflammatory markers and a decrease in Tau hyperphosphorylation. Additionally, oral therapy using a specific A2AR antagonist (MSX-3) significantly improved memory and reduced Tau hyperphosphorylation in THY-Tau22 mice. By showing that A2AR genetic or pharmacological blockade improves the pathological phenotype in a Tau transgenic mouse model, the present data highlight A2A receptors as important molecular targets to consider against AD and Tauopathies.

Object memory enhancement by combining sub-efficacious doses of specific phosphodiesterase inhibitors.

The second messengers cGMP and cAMP have a vital role in synaptic plasticity and memory processes. As such, phosphodiesterases inhibitors (PDE-Is), which prevent the breakdown of these cyclic nucleotides, represent a potential treatment strategy in memory decline. Recently it has been demonstrated that cGMP and cAMP signaling act in sequence during memory consolidation, with early cGMP signaling requiring subsequent cAMP signaling. Here, we sought to confirm this relationship, and to evaluate its therapeutic implications. Combining sub-efficacious doses of the cGMP-specific PDE type 5 inhibitor vardenafil (0.1 mg/kg) and cAMP-specific PDE type 4 inhibitor rolipram (0.01 mg/kg) during the early and late memory consolidation phase, respectively, led to improved memory performance in a 24 h interval object recognition task. Similarly, such a sub-efficacious combination treatment enhanced the transition of early-phase long-term potentiation (LTP) to late-phase LTP in hippocampal slices. In addition, both object memory and LTP were improved after administration of two sub-efficacious doses of the dual substrate PDE type 2 inhibitor BAY60 7550 (0.3 mg/kg) at the early and late consolidation phase, respectively. Taken together, combinations of sub-efficacious doses of cAMP- and cGMP-specific PDE-Is have an additive effect on long-term synaptic plasticity and memory formation and might prove a superior alternative to single PDE-I treatment.

Improved spatial learning is associated with increased hippocampal but not prefrontal long-term potentiation in mGluR4 knockout mice.

Although much information about metabotropic glutamate receptors (mGluRs) and their role in normal and pathologic brain function has been accumulated during the last decades, the role of group III mGluRs is still scarcely documented. Here, we examined mGluR4 knockout mice for types of behavior and synaptic plasticity that depend on either the hippocampus or the prefrontal cortex (PFC). We found improved spatial short- and long-term memory in the radial arm maze, which was accompanied by enhanced long-term potentiation (LTP) in hippocampal CA1 region. In contrast, LTP in the PFC was unchanged when compared with wild-type controls. Changes in paired-pulse facilitation that became overt in the presence of the GABAA antagonist picrotoxin indicated a function of mGluR4 in maintaining the excitation/inhibition balance, which is of crucial importance for information processing in the brain and the deterioration of these processes in neuropsychological disorders such as autism, epilepsy and schizophrenia.

Loss of survivin in neural precursor cells results in impaired long-term potentiation in the dentate gyrus and CA1-region.

In adult mammals, newborn neural precursor cells (NPCs) derived from either the subventricular zone (SVZ) or the subgranular zone (SGZ) migrate into the olfactory bulb and the dentate gyrus (DG), respectively, where some of them mature into excitatory and inhibitory neurons. There is increasing evidence that this neurogenesis process is important for some types of learning and synaptic plasticity and vice versa. Survivin, a member of the inhibitor-of-apoptosis protein (IAP) family, has been suggested to have a central role in the regulation of neurogenesis. The protein is abundantly expressed in nervous tissue during embryonic development while being restricted postnatally to proliferating and migrating NPCs in SVZ and SGZ. Here we examined adult Survivin(Camcre) mice with a conditional deletion of the survivin gene in embryonic neurogenic regions. Although the deletion of survivin had no effect on basic excitability in DG and CA1-region, there was a marked impairment of long-term potentiation (LTP) in these areas. Our data support a function of survivin in hippocampal synaptic plasticity and learning and underline the importance of adult brain neurogenesis for proper operation of the hippocampal tri-synaptic circuit and the physiological functions that depend on it.

An N-methyl-D-aspartate-receptor dependent, late-phase long-term depression in middle-aged mice identifies no GluN2-subunit bias.

Late-phase long-term depression (L-LTD) in middle-aged mice has been difficult to achieve and maintain. Here we report an electrically induced, homosynaptic, input-specific form of LTD that could be stably maintained for at least 4 h in the CA1 area of hippocampal slices of 10-14 months old mice. This form of L-LTD was similar in magnitude in aged, middle-aged and young mice and was blocked by high concentrations of broad-spectrum N-methyl-d-aspartate receptor (NMDAR) antagonists such as d(-)-2-amino-5-phospho-pentanoic acid (d-AP5) and (R)-3-(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid (CPP). Extracellular and whole cell recordings revealed a decreased sensitivity to d-AP5 with age, without any differences in NMDAR conductance between the age groups tested. This L-LTD could be inhibited neither by common doses of NMDA-subunit specific antagonists like zinc, ifenprodil and Ro-25-6981, nor by various co-applications of these compounds. In addition to the lack of any GluN2 subunit bias, L-LTD did not show any discernible involvement of L-type voltage-gated calcium channels. In conclusion, our results do not support any specific role of NMDAR subunits in LTD.

Deficiency of Aph1B/C-gamma-secretase disturbs Nrg1 cleavage and sensorimotor gating that can be reversed with antipsychotic treatment.

Regulated intramembrane proteolysis by gamma-secretase cleaves proteins in their transmembrane domain and is involved in important signaling pathways. At least four different gamma-secretase complexes have been identified, but little is known about their biological role and specificity. Previous work has demonstrated the involvement of the Aph1A-gamma-secretase complex in Notch signaling, but no specific function could be assigned to Aph1B/C-gamma-secretase. We demonstrate here that the Aph1B/C-gamma-secretase complex is expressed in brain areas relevant to schizophrenia pathogenesis and that Aph1B/C deficiency causes pharmacological and behavioral abnormalities that can be reversed by antipsychotic drugs. At the molecular level we find accumulation of Nrg1 fragments in the brain of Aph1BC(-/-) mice. Our observations gain clinical relevance by the demonstration that a Val-to-Leu mutation in the Nrg1 transmembrane domain, associated with increased risk for schizophrenia, affects gamma-secretase cleavage of Nrg1. This finding suggests that dysregulation of intramembrane proteolysis of Nrg1 could increase risk for schizophrenia and related disorders.

FVB.129P2-Pde6b(+) Tyr(c-ch)/Ant, a sighted variant of the FVB/N mouse strain suitable for behavioral analysis.

Mice of the FVB/N strain are severely visual impaired as a result of tyrosinase gene defects, leading to a deficiency of the key enzyme for melanin synthesis in skin and eye and of cyclic guanosine monophosphate phosphodiesterase gene defects, which results in albinism (Tyr(c/c)) and retinal degeneration (Pde6b(rd1/rd1)), respectively. Nevertheless, FVB/N mice are commonly used for the generation of transgenic animals because of their large, strong pronuclei and high breeding performance. However, due to visual impairment of the FVB/N animals, the resulting transgenic animals cannot be used in tests that depend on vision, including tests of cognitive behavior. Therefore, we have bred a sighted version of the FVB/N strain by an outcross between FVB/N and 129P2/OlaHsd, followed by repeated backcrosses to FVB/N mice while selecting against albinism and homozygosity of the retinal degeneration mutation. After 11 generations of backcrossing, sighted animals were intercrossed to generate the congenic FVB.129P2-Pde6b(+) Tyr(c-ch)/Ant strain, which is pigmented (Tyr(c-ch)/(c-ch)) and devoid of the genetic predisposition to retinal degeneration. The accurate visual abilities of the FVB.129P2-Pde6b(+) Tyr(c-ch)/Ant mice, for which we propose the name FVBS/Ant, demonstrated a clear visual evoked potential in the presence of normal eye histology and improved performance in the Morris water maze test.

Modest phenotypic improvements in ASA-deficient mice with only one UDP-galactose:ceramide-galactosyltransferase gene.

BACKGROUND: Arylsulfatase A (ASA)-deficient mice are a model for the lysosomal storage disorder metachromatic leukodystrophy. This lipidosis is characterised by the lysosomal accumulation of the sphingolipid sulfatide. Storage of this lipid is associated with progressive demyelination. We have mated ASA-deficient mice with mice heterozygous for a non-functional allele of UDP-galactose:ceramide-galactosyltransferase (CGT). This deficiency is known to lead to a decreased synthesis of galactosylceramide and sulfatide, which should reduce sulfatide storage and improve pathology in ASA-deficient mice. RESULTS: ASA-/- CGT+/- mice, however, showed no detectable decrease in sulfatide storage. Neuronal degeneration of cells in the spiral ganglion of the inner ear, however, was decreased. Behavioural tests showed small but clear improvements of the phenotype in ASA-/- CGT+/- mice. CONCLUSION: Thus the reduction of galactosylceramide and sulfatide biosynthesis by genetic means overall causes modest improvements of pathology.

The normal phenotype of Pmm1-deficient mice suggests that Pmm1 is not essential for normal mouse development.

Phosphomannomutases (PMMs) are crucial for the glycosylation of glycoproteins. In humans, two highly conserved PMMs exist: PMM1 and PMM2. In vitro both enzymes are able to convert mannose-6-phosphate (mannose-6-P) into mannose-1-P, the key starting compound for glycan biosynthesis. However, only mutations causing a deficiency in PMM2 cause hypoglycosylation, leading to the most frequent type of the congenital disorders of glycosylation (CDG): CDG-Ia. PMM1 is as yet not associated with any disease, and its physiological role has remained unclear. We generated a mouse deficient in Pmm1 activity and documented the expression pattern of murine Pmm1 to unravel its biological role. The expression pattern suggested an involvement of Pmm1 in (neural) development and endocrine regulation. Surprisingly, Pmm1 knockout mice were viable, developed normally, and did not reveal any obvious phenotypic alteration up to adulthood. The macroscopic and microscopic anatomy of all major organs, as well as animal behavior, appeared to be normal. Likewise, lectin histochemistry did not demonstrate an altered glycosylation pattern in tissues. It is especially striking that Pmm1, despite an almost complete overlap of its expression with Pmm2, e.g., in the developing brain, is apparently unable to compensate for deficient Pmm2 activity in CDG-Ia patients. Together, these data point to a (developmental) function independent of mannose-1-P synthesis, whereby the normal knockout phenotype, despite the stringent conservation in phylogeny, could be explained by a critical function under as-yet-unidentified challenge conditions.

Effect of NaCN on currents evoked by uremic retention solutes in dissociated mouse neurons.

Uremic retention solutes possibly contribute to neuronal hypoxia/ischemia and its consequences in patients with renal failure. We examined the in vitro effects of several uremic retention solutes on murine central neurons under chemically induced metabolic hypoxia by application of sodium cyanide (NaCN). Whole cell currents were recorded using the tight-seal whole-cell voltage clamp technique. Application of NaCN caused an inward whole-cell current. From all tested toxins, which included several indoles, guanidino compounds, polyamines, purines, phenols, DL-homocysteine, orotate and myoinositol, only creatinine (CTN), guanidine (G) and guanidinosuccinic acid (GSA) produced a significant current in control and hypoxic neurons. Current evoked by GSA was significantly increased in the chemical hypoxic condition, and a synergistic effect of GSA and spermine was observed in hypoxic neurons.

Nitric oxide in uremia: effects of several potentially toxic guanidino compounds.

Vascular and neurologic impairment remain an important source of morbidity in patients with chronic renal failure (CRF). A portion of CRF patients still suffers from uremic encephalopathy or other signs of nervous system impairment. Several reports demonstrate increased incidence of cardiac infarction and cerebrovascular accidents in CRF patients, even in those with otherwise adequate dialysis treatment [1]. Premature vascular disease, including myocardial infarction, stroke, and peripheral vascular disorder, are the leading causes of death in this population. Although several traditional risk factors for vascular disease and endothelial dysfunction, including smoking, diabetes, dyslipidemia, and hypertension, are often increased in CRF, these factors can only partly explain the high vasculopathy-related morbidity and mortality. Several authors have postulated that CRF-associated atherosclerosis and endothelial dysfunction result from accumulation of certain 'uremic factors,' the identities of which are still a matter of debate. These factors include a variety of guanidino compounds (GCs), which have been shown to be nitric oxide synthase (NOS) modulators both in vitro and in vivo. However, other effects of accumulated uremic GCs have been identified.

Metachromatic leukodystrophy: consequences of sulphatide accumulation.

UNLABELLED: Metachromatic leukodystrophy is a lysosomal lipid storage disorder. It is caused by mutations in the gene for arylsulphatase A, an enzyme involved in the degradation of the sphingolipid 3'-O-sulphogalactosylceramide (sulphatide). This membrane lipid can be found in various cell types, but in particularly high concentrations in the myelin of the nervous system. Patients suffer from progressive, finally lethal, demyelination due to accumulation of sulphatide. In the nervous system, lipid storage not only affects oligodendrocytes but also neurons and, in addition, leads to astrogliosis and activation of microglia. At the cellular level, lysosomal sulphatide storage also affects the lipid composition of myelin itself and has consequences for the amount and localization of particular myelin membrane-associated proteins. Here we review data, largely based on an arylsulphatase A knock-out mouse model of metachromatic leukodystrophy. CONCLUSION: The knock-out mouse model of metachromatic leukodystrophy has provided insights into the histopathological and cellular consequences of sulphatide storage.

Bone marrow stem cell-based gene transfer in a mouse model for metachromatic leukodystrophy: effects on visceral and nervous system disease manifestations.

Arylsulfatase A (ASA) knockout mice represent an animal model for the lysosomal storage disease metachromatic leukodystrophy (MLD). Stem cell gene therapy with bone marrow overexpressing the human ASA cDNA from a retroviral vector resulted in the expression of high enzyme levels in various tissues. Treatment partially reduces sulfatide storage in livers exceeding 18 ng ASA/mg tissue, while complete reduction was observed in livers exceeding 50 ng ASA/mg tissue. This corresponds to about 80% and 200% of normal enzyme activity. Similar values seem to apply for kidney. A partial correction of the lipid metabolism was detectable in the brain where the galactoerebroside/sulfatide ratio, which is diminished in ASA-deficient mice, increased upon treatment. This partial correction was accompanied by amelioration of neuropathology; axonal cross-sectional areas, which are reduced in deficient mice, were significantly increased in the saphenic and sciatic nerve but not in the optic nerve. Behavioral tests suggest some improvement of neuromotor abilities. The gene transfer did not delay the degeneration occurring in the acoustic ganglion of ASA-deficient animals. The limited success of the therapy appears to be due to the requirement of unexpected high levels of ASA for correction of the metabolic defect.

Long-term effect of partial nephrectomy on biological parameters, kidney histology, and guanidino compound levels in mice.

The long-term adverse consequences of early renal mass reduction in mice have not yet been investigated. The effects of partial surgical nephrectomy (NX) in 2-month-old mice on some biological parameters, on histopathologic and morphometric features of the kidney, and on urea and guanidino compound (GC) levels in plasma, urine, and brain were examined at 10 days, and 1, 2, 4, and 12 months postsurgery. Body weight, urinary volume, and plasma urea were most affected at 10 days and 12 months post-NX. NX-induced changes in the remaining renal tissue (including hypertrophy, glomerular mesangial expansion, and presence of protein casts) increased with age. As in human renal insufficiency, NX mice showed significantly higher plasma guanidinosuccinic acid (GSA) and creatinine (CTN) levels at all studied periods. The same tendency could be seen for most other plasma GCs examined, except for arginine (Arg), guanidinoacetic acid (GAA), and homoarginine (HA). As seen in human pathobiochemistry, the latter 2 compounds tended to be lower in NX mice in our follow-up study. Remarkably, and also similar to humans, NX mice excreted less GAA and more GSA than controls during the entire follow-up study. During the follow-up, excretion levels of GAA were unchanged in NX and sham-operated mice. In brain, GAA and gamma-guanidinobutyric acid (GBA) levels were always higher in NX mice with a tendency to respectively increase or decrease over time in NX as well as sham-operated mice. Although urea and GC metabolism were influenced by time post-NX and aging, the model was confirmed to display a mild stable chronic impairment of renal function. Histopathologic and morphometric changes of the kidney increased with age.

Overexpression of arginase alters circulating and tissue amino acids and guanidino compounds and affects neuromotor behavior in mice.

Arginine is an intermediate of the ornithine cycle and serves as a precursor for the synthesis of nitric oxide, creatine, agmatine and proteins. It is considered to be a conditionally essential amino acid because endogenous synthesis only barely meets daily requirements. In rapidly growing suckling neonates, endogenous arginine biosynthesis is crucial to compensate for the insufficient supply of arginine via the milk. Evidence is accumulating that the intestine rather than the kidney plays a major role in arginine synthesis in this period. Accordingly, ectopic expression of hepatic arginase in murine enterocytes by genetic modification induces a selective arginine deficiency. The ensuing phenotype, whose severity correlates with the level of transgene expression in the enterocytes, could be reversed with arginine supplementation. We analyzed the effect of arginine deficiency on guanidine metabolism and neuromotor behavior. Arginine-deficient transgenic mice continued to suffer from an arginine deficiency after the arginine biosynthetic enzymes had disappeared from the enterocytes. Postweaning catch-up growth in arginine-deficient mice was characterized by increased levels of all measured amino acids except arginine. Furthermore, plasma total amino acid concentration, including arginine, was significantly lower in adult male than in adult female transgenic mice. Decreases in the concentration of plasma and tissue arginine led to significant decreases in most metabolites of arginine. However, the accumulation of the toxic guanidino compounds, guanidinosuccinic acid and methylguanidine, corresponded inversely with circulating arginine concentration, possibly reflecting a higher oxidative stress under hypoargininemic conditions. In addition, hypoargininemia was associated with disturbed neuromotor behavior, although brain levels of toxic guanidino compounds and ammonia were normal.

Applications of the Morris water maze in the study of learning and memory.

The Morris water maze (MWM) was described 20 years ago as a device to investigate spatial learning and memory in laboratory rats. In the meanwhile, it has become one of the most frequently used laboratory tools in behavioral neuroscience. Many methodological variations of the MWM task have been and are being used by research groups in many different applications. However, researchers have become increasingly aware that MWM performance is influenced by factors such as apparatus or training procedure as well as by the characteristics of the experimental animals (sex, species/strain, age, nutritional state, exposure to stress or infection). Lesions in distinct brain regions like hippocampus, striatum, basal forebrain, cerebellum and cerebral cortex were shown to impair MWM performance, but disconnecting rather than destroying brain regions relevant for spatial learning may impair MWM performance as well. Spatial learning in general and MWM performance in particular appear to depend upon the coordinated action of different brain regions and neurotransmitter systems constituting a functionally integrated neural network. Finally, the MWM task has often been used in the validation of rodent models for neurocognitive disorders and the evaluation of possible neurocognitive treatments. Through its many applications, MWM testing gained a position at the very core of contemporary neuroscience research.

Biochemical, histological and behavioral consequences of nephrectomy in young and aged mice.

BACKGROUND: This study investigates the effect of nephrectomy in young and aged mice on some biochemical, histological and behavioural aspects. METHODS: Each age group, 2- and 12-months-old, comprised a sham-operated group, a unilaterally nephrectomized group and a subtotally nephrectomized group. Consequences of nephrectomy were examined 10 days postsurgery on urea and guanidino compound levels in body fluids and brain; the remaining kidney by light-microscopic examination; and learning and memory abilities using the Morris water maze task. RESULTS: Effect of nephrectomy on urea and guanidino compound levels in plasma, urine and brain was significantly more pronounced in the young age group. Some guanidino compounds show a tendency to decrease with aging in the sham-operated group and the two nephrectomized groups. Higher compensatory kidney hypertrophy was found in younger nephrectomized mice whereas in older mice glomerular mesangial expansion was a common feature. Finally, young mice with subtotal nephrectomy displayed a slight but significant impairment in memory and learning; whilst old nephrectomized mice manifested no impairment. CONCLUSIONS: Nephrectomy induces more changes in younger mice than in older mice as observed in higher variation of urea and guanidino compound levels, glomerular volume and kidney hypertrophy and decline in spatial learning and memory.