Beneficial Effects of Dinitrosyl Iron Complexes on Wound Healing Compared to Commercial Nitric Oxide Plasma Generator.

Nitric oxide (NO) is a gaseous molecule which plays a key role in wound healing. Previously, we identified the optimal conditions for wound healing strategies using NO donors and an air plasma generator. The aim of this study was to compare the wound healing effects of binuclear dinitrosyl iron complexes with glutathione (B-DNIC-GSH) and NO-containing gas flow (NO-CGF) at their optimal NO doses (0.04 mmol for B-DNIC-GSH and 1.0 mmol for NO-CGF per 1 cm2) in a rat full-thickness wound model over a 3-week period. Excised wound tissues were studied by light and transmission electron microscopy and immunohistochemical, morphometrical and statistical methods. Both treatments had an identical stimulating impact on wound healing, which indicated a higher dosage effectiveness of B-DNIC-GSH compared to the NO-CGF. B-DNIC-GSH spray application reduced inflammation and promoted fibroblast proliferation, angiogenesis and the growth of granulation tissue during the first 4 days after injury. However, prolonged NO spray effects were mild compared to NO-CGF. Future studies should determine the optimal B-DNIC-GSH solution course for a more effective wound healing stimulation.

Proteomic and electron microscopy study of myogenic differentiation of alveolar mucosa multipotent mesenchymal stromal cells in three-dimensional culture.

Myocyte differentiation is featured by adaptation processes, including mitochondria repopulation and cytoskeleton re-organization. The difference between monolayer and spheroid cultured cells at the proteomic level is uncertain. We cultivated alveolar mucosa multipotent mesenchymal stromal cells in spheroids in a myogenic way for the proper conditioning of ECM architecture and cell morphology, which induced spontaneous myogenic differentiation of cells within spheroids. Electron microscopy analysis was used for the morphometry of mitochondria biogenesis, and proteomic was used complementary to unveil events underlying differences between two-dimensional/three-dimensional myoblasts differentiation. The prevalence of elongated mitochondria with an average area of 0.097 µm2 was attributed to monolayer cells 7 days after the passage. The population of small mitochondria with a round shape and area of 0.049 µm2 (p < 0.05) was observed in spheroid cells cultured under three-dimensional conditions. Cells in spheroids were quantitatively enriched in proteins of mitochondria biogenesis (DNM1L, IDH2, SSBP1), respiratory chain (ACO2, ATP5I, COX5A), extracellular proteins (COL12A1, COL6A1, COL6A2), and cytoskeleton (MYL6, MYL12B, MYH10). Most of the Rab-related transducers were inhibited in spheroid culture. The proteomic assay demonstrated delicate mechanisms of mitochondria autophagy and repopulation, cytoskeleton assembling, and biogenesis. Differences in the ultrastructure of mitochondria indicate active biogenesis under three-dimensional conditions.

Lack of synaptic protein, calsyntenin-2, impairs morphology of synaptic complexes in mice.

Calsyntenin-2 (Clstn2) is the synaptic protein, which belongs to the superfamily of cadherins, playing an important role in learning and memory. We recently reported that Clstn2 knockout mice (Clstn2-KO) have a deficit of GABAergic interneurons, associated with hyperactivity, deficient spatial memory, and social behavior. Therefore, we sought to characterize morphometric features of the ultrastructure of synaptic complexes of hippocampal and cortical neurons in Clstn2-KO mice, using high magnification electron microscopy. Morphometric analysis revealed a reduction of symmetric (inhibitory) synaptic density, length of synaptic contacts, and postsynaptic density in neurons of Clstn2-KO mice. Moreover, cortical neurons of Clstn2-KO mice were characterized by the predominance of the simplified type of synapses with the emergence of negative curvature of the synaptic zone in Clstn2-KO mice. Notably, presynaptic zones of cortical neurons of Clstn2-KO mice were characterized by the increased number of synaptic vesicles in opposite to the decreased number of synaptic vesicles in the presynaptic zones of hippocampal neurons. Overall, we found that lack of calsyntenin-2 leads to the striking architectonic alterations of synaptic complexes in the mouse brain, disrupting synaptic density, shape, and connectivity.

Maternal immune activation during gestation interacts with Disc1 point mutation to exacerbate schizophrenia-related behaviors in mice.

Schizophrenia is thought to result from interactions between susceptible genotypes and environmental risk factors. DISC1 is an important gene for schizophrenia and mood disorders based on both human and animal studies. In the present study we sought to investigate interactions between two distinct point mutations in the mouse Disc1 gene (L100P and Q31L) and maternal immune activation (MIA) during pregnancy with polyinosinic:polycytidylic acid (polyI:C). PolyI:C given at 5 mg/kg impaired cognitive and social behavior in both wild-type (WT) and Disc1-Q31L(+/-) offspring, and reduced prepulse inhibition at 16 but not 8 weeks of age. Disc1-L100P(+/-) mutants were more sensitive to MIA than WT or Disc1-Q31L(+/-) mice. Interleukin-6 (IL-6) is a critical cytokine for mediating the behavioral and transcriptional effects of polyI:C. We found a more pronounced increase of IL-6 in response to polyI:C in fetal brain in Disc1-L100P(+/-) mice compared with WT or Disc1-Q31L(+/-) mice. Coadministration of an anti-IL-6 antibody with polyI:C reversed schizophrenia-related behavioral phenotypes in Disc1-L100P(+/-) mice. In summary, we found specific interactions between discrete genetic (Disc1-L100P(+/-)) and environmental factors (MIA) that exacerbate schizophrenia-related phenotypes. IL-6 may be important in the pathophysiology of this interaction.

Tricyclic and tetracyclic antidepressants upregulate VMAT2 activity and rescue disease-causing VMAT2 variants.

Vesicular monoamine transporter 2 (VMAT2) is an essential transporter that regulates brain monoamine transmission and is important for mood, cognition, motor activity, and stress regulation. However, VMAT2 remains underexplored as a pharmacological target. In this study, we report that tricyclic and tetracyclic antidepressants acutely inhibit, but persistently upregulate VMAT2 activity by promoting VMAT2 protein maturation. Importantly, the VMAT2 upregulation effect was greater in BE(2)-M17 cells that endogenously express VMAT2 as compared to a heterologous expression system (HEK293). The net sustained effect of tricyclics and tetracyclics is an upregulation of VMAT2 activity, despite their acute inhibitory effect. Furthermore, imipramine and mianserin, two representative compounds, also demonstrated rescue of nine VMAT2 variants that cause Brain Vesicular Monoamine Transport Disease (BVMTD). VMAT2 upregulation could be beneficial for disorders associated with reduced monoamine transmission, including mood disorders and BVMTD, a rare but often fatal condition caused by a lack of functional VMAT2. Our findings provide the first evidence that small molecules can upregulate VMAT2 and have potential therapeutic benefit for various neuropsychiatric conditions.

The habenular volume and PDE7A allelic polymorphism in major depressive disorder: preliminary findings.

OBJECTIVES: The habenula is a brain structure implicated in depression, yet with unknown molecular mechanisms. Several phosphodiesterases (PDEs) have been associated with a risk of depression. Although the role of PDE7A in the brain is unknown, it has enriched expression in the medial habenula, suggesting that it may play a role in depression. METHODS: We analysed: (1) habenula volume assessed by 3-T magnetic resonance imaging (MRI) in 84 patients with major depressive disorder (MDD) and 41 healthy controls; (2) frequencies of 10 single nucleotide polymorphisms (SNPs) in PDE7A gene in 235 patients and 41 controls; and (3) both indices in 80 patients and 27 controls. The analyses considered gender, age, body mass index and season of the MRI examination. RESULTS: The analysis did not reveal habenula volumetric changes in MDD patients regardless of PDE7A SNPs. However, in the combined group, the carriers of one or more mutations among 10 SNPs in the PDE7A gene had a lower volume of the left habenula (driven mainly by rs972362 and rs138599850 mutations) and consequently had the reduced habenular laterality index in comparison with individuals without PDE7A mutations. CONCLUSIONS: Our findings suggest the implication of the PDE7A gene into mechanisms determining the habenula structure.

Neuronal lack of PDE7a disrupted working memory, spatial learning, and memory but facilitated cued fear memory in mice.

BACKGROUND: PDEs regulate cAMP levels which is critical for PKA activity-dependent activation of CREB-mediated transcription in learning and memory. Inhibitors of PDEs like PDE4 and Pde7 improve learning and memory in rodents. However, the role of PDE7 in cognition or learning and memory has not been reported yet. METHODS: Therefore, we aimed to explore the cognitive effects of a PDE7 subtype, PDE7a, using combined pharmacological and genetic approaches. RESULTS: PDE7a-nko mice showed deficient working memory, impaired novel object recognition, deficient spatial learning & memory, and contextual fear memory, contrary to enhanced cued fear memory, highlighting the potential opposite role of PDE7a in the hippocampal neurons. Further, pharmacological inhibition of PDE7 by AGF2.20 selectively strengthens cued fear memory in C57BL/6 J mice, decreasing its extinction but did not affect cognitive processes assessed in other behavioral tests. The further biochemical analysis detected deficient cAMP in neural cell culture with genetic excision of the PDE7a gene, as well as in the hippocampus of PDE7a-nko mice in vivo. Importantly, we found overexpression of PKA-R and the reduced level of pPKA-C in the hippocampus of PDE7a-nko mice, suggesting a novel mechanism of the cAMP regulation by PDE7a. Consequently, the decreased phosphorylation of CREB, CAMKII, eif2a, ERK, and AMPK, and reduced total level of NR2A have been found in the brain of PDE7a-nko animals. Notably, genetic excision of PDE7a in neurons was not able to change the expression of NR2B, BDNF, synapsin1, synaptophysin, or snap25. CONCLUSION: Altogether, our current findings demonstrated, for the first time, the role of PDE7a in cognitive processes. Future studies will untangle PDE7a-dependent neurobiological and molecular-cellular mechanisms related to cAMP-associated disorders.

Lung inflammation is associated with lipid deposition.

Lung inflammation, pneumonia, is an acute respiratory disease of varying etiology that has recently drawn much attention during the COVID-19 pandemic as lungs are among the main targets for SARS-CoV-2. Multiple other etiological agents are associated with pneumonias. Here, we describe a newly-recognized pathology, namely abnormal lipid depositions in the lungs of patients who died from COVID-19 as well as from non-COVID-19 pneumonias. Our analysis of both semi-thin and Sudan III-stained lung specimens revealed extracellular and intracellular lipid depositions irrespective of the pneumonia etiology. Most notably, lipid depositions were located within vessels adjacent to inflamed regions, where they apparently interfere with the blood flow. Structurally, the lipid droplets in the inflamed lung tissue were homogeneous and lacked outer membranes as assessed by electron microscopy. Morphometric analysis of lipid droplet deposition area allowed us to distinguish the non-pneumonia control lung specimens from the macroscopically intact area of the pneumonia lung and from the inflamed area of the pneumonia lung. Our measurements revealed a gradient of lipid deposition towards the inflamed region. The pattern of lipid distribution proved universal for all pneumonias. Finally, lipid metabolism in the lung tissue was assessed by the fatty acid analysis and by expression of genes involved in lipid turnover. Chromato-mass spectrometry revealed that unsaturated fatty acid content was elevated at inflammation sites compared to that in control non-inflamed lung tissue from the same individual. The expression of genes involved in lipid metabolism was altered in pneumonia, as shown by qPCR and in silico RNA-seq analysis. Thus, pneumonias of various etiologies are associated with specific lipid abnormalities; therefore, lipid metabolism can be considered to be a target for new therapeutic strategies.

Neto1 is a novel CUB-domain NMDA receptor-interacting protein required for synaptic plasticity and learning.

The N-methyl-D-aspartate receptor (NMDAR), a major excitatory ligand-gated ion channel in the central nervous system (CNS), is a principal mediator of synaptic plasticity. Here we report that neuropilin tolloid-like 1 (Neto1), a complement C1r/C1s, Uegf, Bmp1 (CUB) domain-containing transmembrane protein, is a novel component of the NMDAR complex critical for maintaining the abundance of NR2A-containing NMDARs in the postsynaptic density. Neto1-null mice have depressed long-term potentiation (LTP) at Schaffer collateral-CA1 synapses, with the subunit dependency of LTP induction switching from the normal predominance of NR2A- to NR2B-NMDARs. NMDAR-dependent spatial learning and memory is depressed in Neto1-null mice, indicating that Neto1 regulates NMDA receptor-dependent synaptic plasticity and cognition. Remarkably, we also found that the deficits in LTP, learning, and memory in Neto1-null mice were rescued by the ampakine CX546 at doses without effect in wild-type. Together, our results establish the principle that auxiliary proteins are required for the normal abundance of NMDAR subunits at synapses, and demonstrate that an inherited learning defect can be rescued pharmacologically, a finding with therapeutic implications for humans.

Abnormal sensory perception masks behavioral performance of Grin1 knockdown mice.

The development and function of sensory systems require intact glutamatergic neurotransmission. Changes in touch sensation and vision are common symptoms in autism spectrum disorders, where altered glutamatergic neurotransmission is strongly implicated. Further, cortical visual impairment is a frequent symptom of GRIN disorder, a rare genetic neurodevelopmental disorder caused by pathogenic variants of GRIN genes that encode NMDA receptors. We asked if Grin1 knockdown mice (Grin1KD), as a model of GRIN disorder, had visual impairments resulting from NMDA receptor deficiency. We discovered that Grin1KD mice had deficient visual depth perception in the visual cliff test. Since Grin1KD mice are known to display robust changes in measures of learning, memory, and emotionality, we asked whether deficits in these higher-level processes could be partly explained by their visual impairment. By changing the experimental conditions to improve visual signals, we observed significant improvements in the performance of Grin1KD mice in tests that measure spatial memory, executive function, and anxiety. We went further and found destabilization of the outer segment of retina together with the deficient number and size of Meissner corpuscles (mechanical sensor) in the hind paw of Grin1KD mice. Overall, our findings suggest that abnormal sensory perception can mask the expression of emotional, motivational and cognitive behavior of Grin1KD mice. This study demonstrates new methods to adapt routine behavioral paradigms to reveal the contribution of vision and other sensory modalities in cognitive performance.

Behavioral phenotypes of Disc1 missense mutations in mice.

To support the role of DISC1 in human psychiatric disorders, we identified and analyzed two independently derived ENU-induced mutations in Exon 2 of mouse Disc1. Mice with mutation Q31L showed depressive-like behavior with deficits in the forced swim test and other measures that were reversed by the antidepressant bupropion, but not by rolipram, a phosphodiesterase-4 (PDE4) inhibitor. In contrast, L100P mutant mice exhibited schizophrenic-like behavior, with profound deficits in prepulse inhibition and latent inhibition that were reversed by antipsychotic treatment. Both mutant DISC1 proteins exhibited reduced binding to the known DISC1 binding partner PDE4B. Q31L mutants had lower PDE4B activity, consistent with their resistance to rolipram, suggesting decreased PDE4 activity as a contributory factor in depression. This study demonstrates that Disc1 missense mutations in mice give rise to phenotypes related to depression and schizophrenia, thus supporting the role of DISC1 in major mental illness.

Genetic and pharmacological evidence for schizophrenia-related Disc1 interaction with GSK-3.

Recent studies have identified disrupted-in-schizophrenia-1 (DISC1) as a strong genetic risk factor associated with schizophrenia. Previously, we have reported that a mutation in the second exon of the DISC1 gene [leucine to proline at amino acid position 100, L100P] leads to the development of schizophrenia-related behaviors in mice. Glycogen synthase kinase-3 (GSK-3) is a serine/threonine protein kinase that interacts with the N-terminal region of DISC1 (aa 1-220) and has been implicated as an important downstream component in the etiology of schizophrenia. Here, for the first time, we show that pharmacological and genetic inactivation of GSK-3 reverse prepulse inhibition and latent inhibition deficits as well as normalizing the hyperactivity of Disc1-L100P mutants. In parallel to these observations, interaction between DISC1 and GSK-3α and ß is reduced in Disc1-L100P mutants. Our data provide genetic, biochemical, and behavioral evidence for a molecular link between DISC1 and GSK-3 in relation to psychopathology and highlights the value of missense mutations in dissecting the underlying and complex molecular mechanisms of neurological disorders.

COVID-19: Multiorgan Dissemination of SARS-CoV-2 Is Driven by Pulmonary Factors.

Multi-organ failure is one of the common causes of fatal outcome in COVID-19 patients. However, the pathogenetic association of the SARS-CoV-2 viral load (VL) level with fatal dysfunctions of the lungs, liver, kidneys, heart, spleen and brain, as well as with the risk of death in COVID-19 patients remains poorly understood. SARS-CoV-2 VL in the lungs, heart, liver, kidneys, brain, spleen and lymph nodes have been measured by RT qPCR using the following formula: NSARS-CoV-2/NABL1 × 100. Dissemination of SARS-CoV-2 in 30.5% of cases was mono-organ, and in 63.9% of cases, it was multi-organ. The average SARS-CoV-2 VL in the exudative phase of diffuse alveolar damage (DAD) was 60 times higher than in the proliferative phase. The SARS-CoV-2 VL in the lungs ranged from 0 to 250,281 copies. The "pulmonary factors" of SARS-CoV-2 multi-organ dissemination are the high level of SARS-CoV-2 VL (≥4909) and the exudative phase of DAD. The frequency of SARS-CoV-2 dissemination to lymph nodes was 86.9%, heart-56.5%, spleen-52.2%, liver-47.8%, kidney-26%, and brain-13%. We found no link between the SARS-CoV-2 VL level in the liver, kidneys, and heart and the serum level of CPK, LDH, ALP, ALT, AST and Cr of COVID-19 patients. Isolated detection of SARS-CoV-2 RNA in the myocardium of COVID-19 patients who died from heart failure is possible. The pathogenesis of COVID-19-associated multi-organ failure requires further research in a larger cohort of patients.

Viral Load and Patterns of SARS-CoV-2 Dissemination to the Lungs, Mediastinal Lymph Nodes, and Spleen of Patients with COVID-19 Associated Lymphopenia.

Lymphopenia is a frequent hematological manifestation, associated with a severe course of COVID-19, with an insufficiently understood pathogenesis. We present molecular genetic immunohistochemical, and electron microscopic data on SARS-CoV-2 dissemination and viral load (VL) in lungs, mediastinum lymph nodes, and the spleen of 36 patients who died from COVID-19. Lymphopenia <1 × 109/L was observed in 23 of 36 (63.8%) patients. In 12 of 36 cases (33%) SARS-CoV-2 was found in lung tissues only with a median VL of 239 copies (range 18-1952) SARS-CoV-2 cDNA per 100 copies of ABL1. Histomorphological changes corresponding to bronchopneumonia and the proliferative phase of DAD were observed in these cases. SARS-CoV-2 dissemination into the lungs, lymph nodes, and spleen was detected in 23 of 36 patients (58.4%) and was associated with the exudative phase of DAD in most of these cases. The median VL in the lungs was 12,116 copies (range 810-250281), lymph nodes-832 copies (range 96-11586), and spleen-71.5 copies (range 0-2899). SARS-CoV-2 in all cases belonged to the 19A strain. A immunohistochemical study revealed SARS-CoV-2 proteins in pneumocytes, alveolar macrophages, and bronchiolar epithelial cells in lung tissue, sinus histiocytes of lymph nodes, as well as cells of the Billroth pulp cords and spleen capsule. SARS-CoV-2 particles were detected by transmission electron microscopy in the cytoplasm of the endothelial cell, macrophages, and lymphocytes. The infection of lymphocytes with SARS-CoV-2 that we discovered for the first time may indicate a possible link between lymphopenia and SARS-CoV-2-mediated cytotoxic effect.

Vocal and physical phenotypes of calsyntenin2 knockout mouse pups model early-life symptoms of the autism spectrum disorder.

This study discovered a novel acoustic phenotype in Calsyntenin2 deficient knockout (Clstn2-KO) pups in the neurodevelopment period of 5-9 postnatal days (PND 5-9). The narrowband ultrasonic calls (nUSVs) were less complex (mostly one-note, shorter in duration and higher in peak frequency) in Clsnt2-KO than in wild-type (WT) C57BL/6 J pups. The wideband ultrasonic calls (wUSVs) were produced substantially more often by Clstn2-KO than WT pups. The clicks were longer in duration and higher in peak frequency and power quartiles in Clstn2-KO pups. The elevated discomfort due to additional two-minute maternal separation coupled with experimenter's touch, resulted in significantly higher call rates of both nUSVs and clicks in pups of both genotypes and sexes compared to the previous two-minute maternal separation, whereas the call rate of wUSVs was not affected. In Clstn2-KO pups, the prevalence of emission of wUSVs retained at both sex and both degrees of discomfort, thus providing a reliable quantitative acoustic indicator for this genetic line. Besides the acoustic differences, we also detected the increased head-to-body ratio in Clstn2-KO pups. Altogether, this study demonstrated that lack of such synaptic adhesion protein as calsyntenin2 affects neurodevelopment of vocalization in a mouse as a model organism.

The increased density of the habenular neurons, high impulsivity, aggression and resistant fear memory in Disc1-Q31L genetic mouse model of depression.

Mood disorders affect nearly 300 million humans worldwide, and it is a leading cause of death from suicide. In the last decade, the habenula has gained increased attention due to its major role to modulate emotional behavior and related psychopathologies, including depression and bipolar disorder, through the modulation of monoamines' neurotransmission. However, it is still unclear which genetic factors may directly affect the function of the habenula and hence, could contribute to the psychopathological mechanisms of mood disorders. Disrupted-In-Schizophrenia-1 (DISC1) gene is among robust gene-candidates predisposing to major depression, bipolar disorder and schizophrenia in humans. DISC1-Q31L, a well-established genetic mouse model of depression, offers a unique opportunity for translational studies. The current study aimed to probe morphological features of the habenula in the DISC1-Q31L mouse line and detect novel behavioral endophenotypes, including the increased emotionality in mutant females, high aggression in mutant males and deficient extinction of fear memory in DISC1 mutant mice of both sexes. The histological analysis found the increased neural density in the lateral and medial habenula in DISC1-Q31L mice regardless of sex, hence, excluding direct association between the habenular neurons and emotionality in mutant females. Altogether, our findings demonstrated, for the first time, the direct impact of the DISC1 gene on the habenular neurons and affective behavior in the DISC1-Q31L genetic mouse line. These new findings suggest that the combination of the DISC1 genetic analysis together with habenular neuroimaging may improve diagnostics of mood disorder in clinical studies.

Conditioned stimulus presentations alter anxiety level in fear-conditioned mice.

It is generally believed that fear is rapidly triggered by a distinct cue while anxiety onset is less precise and not associated with a distinct cue. Although it has been claimed that both processes can be measured with certain independence of each other, it is unclear how exactly they differ. In this study, we measured anxiety in mice that received discriminative fear conditioning using behavioral, heart rate and calcium (Ca2+) responses in the ventral hippocampal CA1 (vCA1) neurons. We found that the occurrence of fear significantly interfered with anxiety measurements under various conditions. Diazepam reduced basal anxiety level but had no effect during the presentation of conditioned stimulus (CS). Injection of an inhibitory peptide of PKMzeta (ZIP) into the basolateral amygdala almost entirely abolished CS-triggered fear expression and reduced anxiety to basal level. Heart rate measures suggested a small reduction in anxiety during CS-. Calcium responses in the lateral hypothalamus-projecting vCA1 neurons showed a steady decay during CS suggesting a reduced anxiety. Thus, under our experimental conditions, CS presentations likely reduce anxiety level in the fear-conditioned mice.

Kainate receptor auxiliary subunit NETO2 is required for normal fear expression and extinction.

NETO1 and NETO2 are auxiliary subunits of kainate receptors (KARs). They interact with native KAR subunits to modulate multiple aspects of receptor function. Variation in KAR genes has been associated with psychiatric disorders in humans, and in mice, knockouts of the Grik1 gene have increased, while Grik2 and Grik4 knockouts have reduced anxiety-like behavior. To determine whether the NETO proteins regulate anxiety and fear through modulation of KARs, we undertook a comprehensive behavioral analysis of adult Neto1-/- and Neto2-/- mice. We observed no differences in anxiety-like behavior. However, in cued fear conditioning, Neto2-/-, but not Neto1-/- mice, showed higher fear expression and delayed extinction compared to wild type mice. We established, by in situ hybridization, that Neto2 was expressed in both excitatory and inhibitory neurons throughout the fear circuit including the medial prefrontal cortex, amygdala, and hippocampus. Finally, we demonstrated that the relative amount of synaptosomal KAR GLUK2/3 subunit was 20.8% lower in the ventral hippocampus and 36.5% lower in the medial prefrontal cortex in Neto2-/- compared to the Neto2+/+ mice. The GLUK5 subunit abundance was reduced 23.8% in the ventral hippocampus and 16.9% in the amygdala. We conclude that Neto2 regulates fear expression and extinction in mice, and that its absence increases conditionability, a phenotype related to post-traumatic stress disorder and propose that this phenotype is mediated by reduced KAR subunit abundance at synapses of fear-associated brain regions.

Mice with reduced NMDA receptor glycine affinity model some of the negative and cognitive symptoms of schizophrenia.

RATIONALE: Schizophrenic patients demonstrate prominent negative and cognitive symptoms that are poorly responsive to antipsychotic treatment. Abnormal glutamatergic neurotransmission may contribute to these pathophysiological dimensions of schizophrenia. OBJECTIVE: We examined the involvement of the glycine coagonist site on the N-methyl-D: -aspartate receptor (NMDAR) glycine coagonist site in the modulation of negative and cognitive endophenotypes in mice. MATERIALS AND METHODS: Behavioral phenotypes relevant to schizophrenia were assessed in Grin1(D481N) mice that have reduced NMDAR glycine affinity. RESULTS: Grin1(D481N) mutant mice showed abnormally persistent latent inhibition (LI) that was reversed by two agents that enhance NMDAR glycine site function, D: -serine (600 mg/kg) and ALX-5407 (1 mg/kg), and by the classical atypical antipsychotic clozapine (3 mg/kg). Similarly, blockade of the NMDAR glycine site with the antagonist L-701,324 (5 mg/kg) induced persistent LI in C57BL6/J mice. In a social affiliations task, Grin1(D481N) mutant animals showed reduced social approach behaviors that were normalized by D: -serine (600 mg/kg). During a nonassociative spatial object recognition task, mutant mice demonstrated impaired reactivity to a spatial change that was reversible by D: -serine (300 and 600 mg/kg) and clozapine (0.75 mg/kg). In contrast, responses to social novelty and nonspatial change remained unaffected, indicating that the Grin1(D481N) mutation induces selective deficits in sociability and spatial discrimination, while leaving intact the ability to react to novelty. CONCLUSIONS: Genetic and pharmacologically induced deficiencies in glycine binding appear to model the impairments in behavioral flexibility, sociability, and spatial recognition related to the negative and cognitive symptoms of schizophrenia. Antipsychotics that target the NMDAR glycine site may be beneficial in treating such psychiatric symptoms.

Uncoupling DISC1 × D2R Protein-Protein Interactions Facilitates Latent Inhibition in Disc1-L100P Animal Model of Schizophrenia and Enhances Synaptic Plasticity via D2 Receptors.

Both Disrupted-In-Schizophrenia-1 (DISC1) and dopamine receptors D2R have significant contributions to the pathogenesis of schizophrenia. Our previous study demonstrated that DISC1 binds to D2R and such protein-protein interaction is enhanced in patients with schizophrenia and Disc1-L100P mouse model of schizophrenia (Su et al., 2014). By uncoupling DISC1 × D2R interaction (trans-activator of transcription (TAT)-D2pep), the synthesized TAT-peptide elicited antipsychotic-like effects in pharmacological and genetic animal models, without motor side effects as tardive dyskinesia commonly seen with typical antipsychotic drugs (APDs), indicating that the potential of TAT-D2pep of becoming a new APD. Therefore, in the current study, we further explored the APD-associated capacities of TAT-D2pep. We found that TAT-D2pep corrected the disrupted latent inhibition (LI), as a hallmark of schizophrenia associated endophenotype, in Disc1-L100P mutant mice-a genetic model of schizophrenia, supporting further APD' capacity of TAT-D2pep. Moreover, we found that TAT-D2pep elicited nootropic effects in C57BL/6NCrl inbred mice, suggesting that TAT-D2pep acts as a cognitive enhancer, a desirable feature of APDs of the new generation. Namely, TAT-D2pep improved working memory in T-maze, and cognitive flexibility assessed by the LI paradigm, in C57BL/6N mice. Next, we assessed the impact of TAT-D2pep on hippocampal long-term plasticity (LTP) under basal conditions and upon stimulation of D2 receptors using quinpirole. We found comparable effects of TAT-D2pep and its control TAT-D2pep-scrambled peptide (TAT-D2pep-sc) under basal conditions. However, under stimulation of D2R by quinpirole, LTP was enhanced in hippocampal slices incubated with TAT-D2pep, supporting the notion that TAT-D2pep acts in a dopamine-dependent manner and acts as synaptic enhancer. Overall, our experiments demonstrated implication of DISC1 × D2R protein-protein interactions into mechanisms of cognitive and synaptic plasticity, which help to further understand molecular-cellular mechanisms of APD of the next generation.