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Klotho (KL) is a glycosyl hydrolase and aging-suppressor gene. Stress is a risk factor for depression and anxiety, which are highly comorbid with each other. The aim of this study is to determine whether KL is regulated by estrogen and plays an important role in sex differences in stress resilience. Our results showed that KL is regulated by estrogen in rat hippocampal neurons in vivo and in vitro and is essential for the estrogen-mediated increase in the number of presynaptic vesicular glutamate transporter 1 (Vglut1)-positive clusters on the dendrites of hippocampal neurons. The role of KL in sex differences in stress response was examined in rats using 3-week chronic unpredictable mild stress (CUMS). CUMS produced a deficit in spatial learning and memory, anhedonic-like behaviors, and anxiety-like behaviors in male but not female rats, which was accompanied by a reduction in KL protein levels in the hippocampus of male but not female rats. This demonstrated the resilience of female rats to CUMS. Interestingly, the knockdown of KL protein levels in the rat hippocampus of both sexes caused a decrease in stress resilience in both sexes, especially in female rats. These results suggest that the regulation of KL by estrogen plays an important role in estrogen-mediated synapse formation and that KL plays a critical role in the sex differences in cognitive deficit, anhedonic-like behaviors, and anxiety-like behaviors induced by chronic stress in rats, highlighting an important role of KL in sex differences in stress resilience. [ABSTRACT FROM AUTHOR]

As a key enzyme for nitrogen assimilation in plant, glutamine synthetase (TaGS) is encoded by 12 genes in common wheat (Triticum aestivum L.), namely TaGS1;1-6A/6B/6D, TaGS1;2-4A/4B/4D, TaGS1;3-4A/4B/4D, and TaGS2-2A/2B/2D. Transcripts of TaGS genes were obtained using the single molecular sequencing technology, and the results showed that TaGS1;1-6A had one alternative splicing (AS) version of transcripts and TaGS1;1-6B had two AS versions. Compared with the TaGS encoded by normal transcript, TaGS1;1-6A-1 lacked part of the Gln_synt_N domain, and TaGS1;1-6B-3 lacked the Gln_synth_gly_rich_ site domain; TaGS1;1-6B-4 was a new AS event, and encoded a truncated GS without Gln_synth_gly_rich_site and Gln_synth_ cat_dom domain. The relative expression level of TaGS alternative splicing transcripts of wheat cultivars with different nitrogen use efficiencies showed that the relative expression level of TaGS1;1-6A-1 in leaves of YM49 (N-efficient) was significantly higher than that of XN509 (N-inefficient), and increased with the increase of nitrogen supply. In roots, the relative expression of TaGS1;1-6A-1 increased with the increase of nitrogen supply in YM49, but decreased in XN509. TaGS1;1-6B-3 showed a similar expression trend in YM49 and XN509, and high concentration of NO3- promoted the expression of TaGS1;1-6B-3 in leaves while inhibited in roots. High concentration of NH4+ inhibited the expression of TaGS1;1-6B-3 in leaves, but had little effect in roots. TaGS1;1-6B-4 was mainly expressed under low nitrogen condition, and its expression in roots of YM49 was not affected by NH4+ supply. Understanding the AS transcript versions of TaGS genes is helpful to illustrate the functions of TaGS isozymes in nitrogen metabolism of wheat. [ABSTRACT FROM AUTHOR]

Abstract Background The human KALRN gene, which encodes a complex, multifunctional Rho GDP/GTP exchange factor, has been linked to cardiovascular disease, psychiatric disorders and neurodegeneration. Examination of existing Kalrn knockout mouse models has focused only on neuronal phenotypes. However, Kalirin was first identified through its interaction with an enzyme involved in the synthesis and secretion of multiple bioactive peptides, and studies in C.elegans revealed roles for its orthologue in neurosecretion. Results We used a broad array of tests to evaluate the effects of ablating a single exon in the spectrin repeat region of Kalrn (KalSRKO/KO); transcripts encoding Kalrn isoforms containing only the second GEF domain can still be produced from the single remaining functional Kalrn promoter. As expected, KalSRKO/KO mice showed a decrease in anxiety-like behavior and a passive avoidance deficit. No changes were observed in prepulse inhibition of acoustic startle or tests of depression-like behavior. Growth rate, parturition and pituitary secretion of growth hormone and prolactin were deficient in the KalSRKO/KO mice. Based on the fact that a subset of Kalrn isoforms is expressed in mouse skeletal muscle and the observation that muscle function in C.elegans requires its Kalrn orthologue, KalSRKO/KO mice were evaluated in the rotarod and wire hang tests. KalSRKO/KO mice showed a profound decrease in neuromuscular function, with deficits apparent in KalSR+/KO mice; these deficits were not as marked when loss of Kalrn expression was restricted to the nervous system. Pre- and postsynaptic deficits in the neuromuscular junction were observed, along with alterations in sarcomere length. Conclusions Many of the widespread and diverse deficits observed both within and outside of the nervous system when expression of Kalrn is eliminated may reflect its role in secretory granule function and its expression outside of the nervous system.

Abstract Background The long-term effects of cocaine on behavior are accompanied by structural changes in excitatory glutamatergic synapses onto the medium spiny neurons of the striatum. The Kalrn gene encodes several functionally distinct isoforms; these multidomain guanine nucleotide exchange factors (GEFs) contain additional domains known to interact with phosphatidylinositides as well as with a number of different proteins. Through their activation of Rho proteins and their interactions with other proteins, the different Kalirin isoforms affect cytoskeletal organization. Chronic exposure of adult male rodents to cocaine increases levels of Kalirin 7 in the striatum. When exposed chronically to cocaine, mice lacking Kalirin 7, the major adult isoform, fail to show an increase in dendritic spine density in the nucleus accumbens, show diminished place preference for cocaine, and exhibit increased locomotor activity in response to cocaine. Results The use of alternate promoters and 3'-terminal exons of the mouse Kalrn gene were investigated using real-time quantitative polymerase chain reaction. While the two most distal full-length Kalrn promoters are used equally in the prefrontal cortex, the more proximal of these promoters accounts for most of the transcripts expressed in the nucleus accumbens. The 3'-terminal exon unique to the Kalirin 7 isoform accounts for a greater percentage of the Kalrn transcripts in prefrontal cortex than in nucleus accumbens. Western blot analyses confirmed these differences. Chronic cocaine treatment increases usage of the promoter encoding the Δ-Kalirin isoforms but does not alter full-length Kalirin promoter usage. Usage of the 3'-terminal exon unique to Kalirin 7 increases following chronic cocaine exposure. Conclusions Kalrn promoter and 3'-terminal exon utilization are region-specific. In the nucleus accumbens, cocaine-mediated alterations in promoter usage and 3'-terminal exon usage favor expression of Kalirin 7 and Δ-Kalirin 7. The Δ-isoform, which lacks a Sec14p domain and four of the nine spectrin-like repeats found in full-length Kalirin isoforms, increases spine headsize without increasing dendritic spine numbers. Thus cocaine-mediated changes in alternative splicing of the Kalrn gene may contribute importantly to the behavioral, morphological and biochemical responses observed.

Neuropeptides and peptide hormones play a crucial role in integrating the many factors that affect physiologic and cognitive processes. The potency of many of these peptides requires an amidated amino acid at the C‐terminus; a single enzyme, peptidylglycine α‐amidating monooxygenase (PAM), catalyzes this modification. Anxiety‐like behavior is known to be altered in mice with a single functional Pam allele (Pam+/−) and in mice unable to express Pam in excitatory forebrain neurons (PamEmx1‐cKO/cKO) or in cardiomyocytes (PamMyh6‐cKO/cKO). Examination of PAM‐positive and glutamic acid decarboxylase 67 (GAD)‐positive cells in the amygdala of PamEmx1‐cKO/cKO mice demonstrated the absence of PAM in pyramidal neurons and its continued presence in GAD‐positive interneurons, suggestive of altered excitatory/inhibitory balance. Additional behavioral tests were used to search for functional alterations in these cell‐type specific knockout mice. PamEmx1‐cKO/cKO mice exhibited a less focused search pattern for the Barnes Maze escape hole than control or PamMyh6‐cKO/cKO mice. While wildtype mice favor interacting with novel objects as opposed to familiar objects, both PamEmx1‐cKO/cKO and PamMyh6‐cKO/cKO mice exhibited significantly less interest in the novel object. Since PAM levels in the central nervous system of PamMyh6‐cKO/cKO mice are unaltered, the behavioral effect observed in these mice may reflect their inability to produce atrial granules and the resulting reduction in serum levels of atrial natriuretic peptide. In the sociability test, male mice of all three genotypes spent more time with same‐sex stranger mice; while control females showed no preference for stranger mice, female PamEmx1‐cKO/cKO mice showed preference for same‐sex stranger mice in all trials. [ABSTRACT FROM AUTHOR]

Estrogen (E2) modulates a wide range of neural functions such as spine formation, synaptic plasticity, and neurotransmission in the hippocampus. Dendritic spines and synapse numbers in hippocampal neurons of female rats cyclically fluctuate across the estrous cycle, but the key genes responsible for these fluctuations are still unknown. In order to address this question, we explore the hippocampal transcriptome via RNA-sequencing (RNA-seq) at the proestrus (PE) and estrus (ES) stages in female rats. At standard fold-change selection criteria, 37 differentially expressed genes (DEGs) were found in PE vs. ES groups (FDR adjusted p-value (q)<0.05). The transcriptional changes identified by RNA-seq were confirmed by quantitative real-time PCR. To gain insight into the function of the DEGs, the E2-regulated genes were annotated by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes database (KEGG). Based on GO and KEGG pathways, the identified DEGs of PE vs. ES stages are involved in extracellular matrix formation, regulation of actin cytoskeleton, oxidative stress, neuroprotection, immune system, oligodendrocyte maturation and myelination, signal transduction pathways, growth factor signaling, retinoid signaling, aging, cellular process, metabolism and transport. The profiles of the gene expression in the hippocampus identified at the PE vs. ES stages were compared with the gene expression profiles in ovariectomized (OVX) rats receiving E2 replacement via RNA-seq and qPCR. The profiles of gene expression between the OVX+E2 and the estrous cycle were different and the possible causes were discussed. [ABSTRACT FROM AUTHOR]

Depression is one of the most prevalent psychiatric disorders. Exercise has been shown to be effective in the amelioration of depression, but the underlying mechanism remains largely unknown. Alterations in the density and morphology of dendritic spines are associated with psychiatric diseases. Chronic unpredictable mild stress (CUMS) is an established animal model of depression. The aim of this study was to determine whether treadmill exercise reverses CUMS-induced both depression-like behaviors and alterations in spine density and morphology of the principal neurons in the brain areas of the mood circuits including the hippocampus, medial prefrontal cortex (mPFC), nucleus accumbens (NAc) and basolateral amygdala (BLA). Male rats were randomly divided into four groups: control, CUMS, exercise, and CUMS+exercise. CUMS-induced depression-like behaviors were evaluated by the sucrose preference test (SPT). Golgi staining was used to visualize dendritic spines. Our results showed that CUMS-induced depression-like behaviors characterized by a decrease in sucrose consumption were accompanied by a decrease in spine density and a change in spine morphology in the pyramidal neurons of both the hippocampal CA3 area and the mPFC, and an increase in spine density and an alteration in spine shape in both the NAc medium spiny neurons (MSNs) and the BLA neurons; exercise reversed both CUMS-induced depression-like behaviors and alterations in dendritic spines. This study provides important information for understanding the mechanism through which exercise ameliorates CUMS-induced depression-like behaviors. [ABSTRACT FROM AUTHOR]

Scaffolds used for soft tissue regeneration are designed to mimic the native extracellular matrix (ECM) structurally and provide adequate mechanical strength and degradation properties. Scaffold architecture, porosity, stiffness and presence of soluble factors have been shown to influence human mesenchymal stem cells (hMSCs) differentiation along neuronal lineage. The present manuscript evaluated the performance of a composite scaffold comprised of electrospun polycaprolactone (PCL) nanofiber lattice coated with sodium alginate (SA) for neural tissue engineering. The nanofiber lattice was included in the scaffold to provide tensile strength and retain suture thread on the nerve graft. Sodium alginate was used to control matrix hydrophilicity, material stiffness and controlled release of biological molecules. The effect of SA molecular weight on the composite scaffold tensile properties, hMSCs adhesion, proliferation and neurogenic differentiation was evaluated. Both random and aligned composite scaffolds showed significantly higher tensile properties as compared to PCL fiber matrix alone indicating the reinforcement of SA hydrogel into fiber lattice. Low molecular weight SA coating because of its low viscosity resulted in uniform penetration into the fiber lattice and resulted in significantly higher tensile strength as compared to high molecular weight SA. Both composite scaffolds showed a controlled SA erosion rate and lost >95% of the SA coating over a period of 10 days under in vitro conditions. Composite scaffolds showed progressive hMSCs growth over 14 days and resulted in significantly higher amount of DNA content (almost double on day 7 and 14) as compared to control PCL fiber matrices. Immunostaining experiments showed higher and uniform expression of the neurotropic protein S-100 on composite scaffolds containing low molecular weight SA. These composite scaffolds may be suitable for peripheral nerve regeneration. Copyright © 2015 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Background: Liquid-state specimen carriers are inadequate for sample transportation in large-scale screening projects in low-resource settings, which necessitates the exploration of novel non-hazardous solid-state alternatives. Studies investigating the feasibility and accuracy of a solid-state human papillomavirus (HPV) sampling medium in combination with different down-stream HPV DNA assays for cervical cancer screening are needed.Methods: We collected two cervical specimens from 396 women, aged 25-65 years, who were enrolled in a cervical cancer screening trial. One sample was stored using DCM preservative solution and the other was applied to a Whatman Indicating FTA Elute® card (FTA card). All specimens were processed using three HPV testing methods, including Hybrid capture 2 (HC2), careHPV™, and Cobas®4800 tests. All the women underwent a rigorous colposcopic evaluation that included using a microbiopsy protocol.Results: Compared to the liquid-based carrier, the FTA card demonstrated comparable sensitivity for detecting high grade Cervical Intraepithelial Neoplasia (CIN) using HC2 (91.7 %), careHPV™ (83.3 %), and Cobas®4800 (91.7 %) tests. Moreover, the FTA card showed a higher specificity compared to a liquid-based carrier for HC2 (79.5 % vs. 71.6 %, P = 0.015), comparable specificity for careHPV™ (78.1 % vs. 73.0 %, P > 0.05), but lower specificity for the Cobas®4800 test (62.4 % vs. 69.9 %, P = 0.032). Generally, the FTA card-based sampling medium's accuracy was comparable with that of liquid-based medium for the three HPV testing assays.Conclusions: FTA cards are a promising sample carrier for cervical cancer screening. With further optimization, it can be utilized for HPV testing in areas of varying economic development. [ABSTRACT FROM AUTHOR]

In neurons, specific RNAs are assembled into granules, which are translated in dendrites, however the functional consequences of granule assembly are not known. Tumor overexpressed gene (TOG) is a granule-associated protein containing multiple binding sites for heterogeneous nuclear ribonucleoprotein (hnRNP) A2, another granule component that recognizes cis-acting sequences called hnRNP A2 response elements (A2REs) present in several granule RNAs. Translation in granules is sporadic, which is believed to reflect monosomal translation, with occasional bursts, which are believed to reflect polysomal translation. In this study, TOG expression was conditionally knocked out (TOG cKO) in mouse hippocampal neurons using cre/lox technology. In TOG cKO cultured neurons granule assembly and bursty translation of activity-regulated cytoskeletal associated (ARC) mRNA, an A2RE RNA, are disrupted. In TOG cKO brain slices synaptic sensitivity and long term potentiation (LTP) are reduced. TOG cKO mice exhibit hyperactivity, perseveration and impaired short term habituation. These results suggest that in hippocampal neurons TOG is required for granule assembly, granule translation and synaptic plasticity, and affects behavior. [ABSTRACT FROM AUTHOR]

Estradiol enhances the formation of dendritic spines and excitatory synapses in hippocampal neurons in vitro and in vivo, but the underlying mechanisms are not fully understood. Kalirin-7 (Kal7), the major isoform of Kalirin in the adult hippocampus, is a Rho GDP/GTP exchange factor localized to postsynaptic densities. In the hippocampus, both Kal7 and estrogen receptor α (ERα) are highly expressed in a subset of interneurons. Over-expression of Kal7 caused an increase in spine density and size in hippocampal neurons. To determine whether Kalirin might play a role in the effects of estradiol on spine formation, Kal7 expression was examined in the hippocampus of ovariectomized rats. Estradiol replacement increased Kal7 staining in both CA1 pyramidal neurons and interneurons in ovariectomized rats. Estradiol treatment of cultured hippocampal neurons increased Kal7 levels at the postsynaptic side of excitatory synapses and increased the number of excitatory synapses along the dendrites of pyramidal neurons. These increases were mediated via ERα because a selective ERα agonist, but not a selective ERβ agonist, caused a similar increase in both Kal7 levels and excitatory synapse number in cultured hippocampal neurons. When Kal7 expression was reduced using a Kal7-specific shRNA, the density of excitatory synapses was reduced and estradiol was no longer able to increase synapse formation. Expression of exogenous Kal7 in hippocampal interneurons resulted in decreased levels of GAD65 staining. Inhibition of GABAergic transmission with bicuculline produced a robust increase in Kal7 expression. These studies suggest Kal7 plays a key role in the mechanisms of estradiol-mediated synaptic plasticity. © 2010 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

The mammalian amygdala expresses various neuropeptides whose signaling has been implicated in emotionality. Many neuropeptides require amidation for full activation by peptidylglycine-amidating monooxygenase (PAM), a transmembrane vesicular cuproenzyme and regulator of the secretory pathway. Mice heterozygous for the Pam gene (PAM/) exhibit physiological and behavioral abnormalities related to specific peptidergic pathways. In the present study, we evaluated emotionality and examined molecular and cellular responses that characterize neurophysiological differences in the PAM/amygdala. PAM/ mice presented with anxiety-like behaviors in the zero maze that were alleviated by diazepam.PAM/animals were deficient in short- and long-term contextual and cued fear conditioning and required higher shock intensities to establish fear-potentiated startle than their wild-type littermates. Immunohistochemical analysis of the amygdala revealed PAM expression in pyramidal neurons and local interneurons that synthesize GABA. We performed whole-cell recordings of pyramidal neurons in the PAM/ amygdala to elucidate neurophysiological correlates of the fear behavioral phenotypes. Consistent with these observations, thalamic afferent synapses in the PAM/ lateral nucleus were deficient in long-term potentiation. This deficit was apparent in the absence and presence of the GABAA receptor antagonist picrotoxin and was abolished when both GABAA and GABAB receptors were blocked. Both evoked and spontaneous excitatory signals were enhanced in the PAM/ lateral nucleus. Phasic GABAergic signaling was also augmented in the PAM/ amygdala, and this difference comprised activity-independent and dependent components. These physiological findings represent perturbations in the PAM/ amygdala that may underlie the aberrant emotional responses in the intact animal. [ABSTRACT FROM AUTHOR]

Loss of endogenous estrogen and dysregulation of the estrogen receptor signaling pathways are associated with an increase in risk for cognitive deficit and depression in women after menopause. Estrogen therapy for menopause increases the risk of breast and ovarian cancers, and stroke. Therefore, it is critical to find an alternate treatment for menopausal women. Osthole (OST), a coumarin, has been reported to have neuroprotective effects. This study examined whether OST improves ovariectomy (OVX)-induced cognitive impairment, and alleviates anxiety- and depression-like behaviors induced by OVX in mice. Adult female C57BL/6J mice were ovariectomized and then treated with OST at a dose of 30 mg/kg for 14 days. At the end of the treatment period, behavioral tests were used to evaluate spatial learning and memory, recognition memory, anxiety- and depression-like behaviors. A cohort of the mice were sacrificed after 14 days of OST treatment and their hippocampi were collected for measurement of the proteins of interest using western blot. OVX-induced alteration in the levels of proteins was accompanied by cognitive deficit, anxiety- and depression-like behaviors. OST treatment improved cognitive deficit, alleviated anxiety- and depression-like behaviors induced by OVX, and reversed OVX-induced alterations in the levels of synaptic proteins and ERα, BDNF, TrKB, p-CREB, p-Akt and Rac1 in the hippocampus. Therefore, reversal of OVX-induced decrease in the levels of hippocampal proteins by OST might contribute to the effects of OST on improving cognitive deficit and alleviating anxiety- and depression-like behaviors induced by OVX. [ABSTRACT FROM AUTHOR]

Background: The long-term effects of cocaine on behavior are accompanied by structural changes in excitatory glutamatergic synapses onto the medium spiny neurons of the striatum. The Kalrn gene encodes several functionally distinct isoforms; these multidomain guanine nucleotide exchange factors (GEFs) contain additional domains known to interact with phosphatidylinositides as well as with a number of different proteins. Through their activation of Rho proteins and their interactions with other proteins, the different Kalirin isoforms affect cytoskeletal organization. Chronic exposure of adult male rodents to cocaine increases levels of Kalirin 7 in the striatum. When exposed chronically to cocaine, mice lacking Kalirin 7, the major adult isoform, fail to show an increase in dendritic spine density in the nucleus accumbens, show diminished place preference for cocaine, and exhibit increased locomotor activity in response to cocaine.Results: The use of alternate promoters and 3'-terminal exons of the mouse Kalrn gene were investigated using real-time quantitative polymerase chain reaction. While the two most distal full-length Kalrn promoters are used equally in the prefrontal cortex, the more proximal of these promoters accounts for most of the transcripts expressed in the nucleus accumbens. The 3'-terminal exon unique to the Kalirin 7 isoform accounts for a greater percentage of the Kalrn transcripts in prefrontal cortex than in nucleus accumbens. Western blot analyses confirmed these differences. Chronic cocaine treatment increases usage of the promoter encoding the Δ-Kalirin isoforms but does not alter full-length Kalirin promoter usage. Usage of the 3'-terminal exon unique to Kalirin 7 increases following chronic cocaine exposure.Conclusions: Kalrn promoter and 3'-terminal exon utilization are region-specific. In the nucleus accumbens, cocaine-mediated alterations in promoter usage and 3'-terminal exon usage favor expression of Kalirin 7 and Δ-Kalirin 7. The Δ-isoform, which lacks a Sec14p domain and four of the nine spectrin-like repeats found in full-length Kalirin isoforms, increases spine headsize without increasing dendritic spine numbers. Thus cocaine-mediated changes in alternative splicing of the Kalrn gene may contribute importantly to the behavioral, morphological and biochemical responses observed. [ABSTRACT FROM AUTHOR]