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Aim: Chronic intermittent hypoxia (CIH) is a key characteristic of obstructive sleep apnea (OSA) syndrome, a chronic respiratory disorder. The mechanisms of CIH-induced metabolic disturbance and histopathological damage remain unclear. Methods: CIH-induced rats underwent daily 8-h CIH, characterized by oxygen levels decreasing from 21% to 8.5% over 4 min, remaining for 2 min, and quickly returning to 21% for 1 min. The control rats received a continuous 21% oxygen supply. The levels of hypersensitive C reactive protein (h-CRP), tumor necrosis factor-a (TNF-a), interleukin 6 (IL-6), interleukin 8 (IL-8), and nuclear factor kappa-B (NF-kB) were measured by ELISA. Histological analysis of the soft palates was conducted using HE staining. The microbial profiling of fecal samples was carried out by Accu16STM assay. Untargeted metabolomics of serum and soft palate tissue samples were analyzed by UPLC-MS. The protein expression of cAMP-related pathways in the soft palate was determined by Western blot. Results: After 28 h of CIH induction, a significant increase in pro-inflammatory cytokines was observed in the serum, along with mucosal layer thickening and soft palate tissue hypertrophy. CIH induction altered the diversity and composition of fecal microbiota, specifically reducing beneficial bacteria while increasing harmful bacteria/opportunistic pathogens. Notably, CIH induction led to a significant enrichment of genera such as Dorea, Oscillibacter, Enteractinococcus, Paenibacillus, Globicatella, and Flaviflexus genera. Meanwhile, Additionally, CIH induction had a notable impact on 108 serum marker metabolites. These marker metabolites, primarily involving amino acids, organic acids, and a limited number of flavonoids or sterols, were associated with protein transport, digestion and absorption, amino acid synthesis and metabolism, as well as cancer development. Furthermore, these differential serum metabolites significantly affected 175 differential metabolites in soft palate tissue, mainly related to cancer development, signaling pathways, amino acid metabolism, nucleotide precursor or intermediate metabolism, respiratory processes, and disease. Importantly, CIH induction could significantly affect the expression of the cAMP pathway in soft palate tissue. Conclusions: Our findings suggest that targeting differential metabolites in serum and soft palate tissue may represent a new approach to clinical intervention and treatment of OSA simulated by the CIH. [ABSTRACT FROM AUTHOR]

Neutrophils and their production of neutrophil extracellular traps (NETs) significantly contribute to neuroinflammation and brain damage after intracerebral hemorrhage (ICH). Although Akebia saponin D (ASD) demonstrates strong anti‐inflammatory activities and blood–brain barrier permeability, its role in regulating NETs formation and neuroinflammation following ICH is uncharted. Our research focused on unraveling the influence of ASD on neuroinflammation mediated by NETs and the mechanisms involved. We found that increased levels of peripheral blood neutrophils post‐ICH are correlated with worse prognostic outcomes. Through network pharmacology, we identified ASD as a promising therapeutic target for ICH. ASD administration significantly improved neurobehavioral performance and decreased NETs production in neutrophils. Furthermore, ASD was shown to upregulate the membrane protein NTSR1 and activate the cAMP signaling pathway, confirmed through transcriptome sequencing, western blot, and immunofluorescence. Interestingly, the NTSR1 inhibitor SR48692 significantly nullified ASD's anti‐NETs effects and dampened cAMP pathway activation. Mechanistically, suppression of PKAc via H89 negated ASD's anti‐NETs effects but did not affect NTSR1. Our study suggests that ASD may reduce NETs formation and neuroinflammation, potentially involving the NTSR1/PKAc/PAD4 pathway post‐ICH, underlining the potential of ASD in mitigating neuroinflammation through its anti‐NETs properties. [ABSTRACT FROM AUTHOR]

This toxicology study was conducted to assess the impact of formaldehyde, a common air pollutant found in Chinese gymnasiums, on the brain function of athletes. In this research, a total of 24 Balb/c male mice of SPF-grade were divided into four groups, each consisting of six mice. The mice were exposed to formaldehyde at different concentrations, including 0 mg/m3, 0.5 mg/m3, 3.0 mg/m3, and 3.0 mg/m3 in combination with an injection of L-NMMA (NG-monomethyl-L-arginine), which is a nitric oxide synthase antagonist. Following a one-week test period (8 h per day, over 7 days), measurements of biomarkers related to the nitric oxide (NO)/cGMP-cAMP signaling pathway were carried out on the experimental animals post-treatment. The study found that: (1) Exposure to formaldehyde can lead to brain cell apoptosis and neurotoxicity; (2) Additionally, formaldehyde exposure was found to alter the biomarkers of the NO/cGMP-cAMP signaling pathway, with some changes being statistically significant (p < 0.05 or p < 0.01); (3) The use of L-NMMA, an antagonist of the NO/cGMP-cAMP signaling pathway, was found to prevent these biomarker changes and had a protective effect on brain cells. The study suggests that the negative impact of formaldehyde on the brain function of mice is linked to the regulation of the NO/cGMP-cAMP signaling pathway. [ABSTRACT FROM AUTHOR]

Among the most sought after breakthroughs nowadays to combat computational saturation in the electronic hardware realm, neuromorphic and cytomorphic mimetics of biological structures seem potentially promising. Biological circuits are distinguishable due to their minuscule dimensions and immensely low power consumption; yet they achieve extremely complex and magnificent tasks of life, such as, thinking, memorizing, decision making and self-regulating in response to the surroundings. Low power analog circuit solutions are edged over digital ones as they are inherently noisy and fuzzy like bio-systems. In this paper, an analog circuit equivalent for a well-known biological pathway, cyclic adenosine monophosphate (cAMP), has been proposed, exploiting the fabrication characteristics of an analog transistor. The work demonstrates an application of previously published research of the authors, where it was shown that a single transistor operating in analog mode can mimic some fundamental biological circuit processes like receptor-ligand binding, Michaelis Menten and Hill process reactions. Since biological pathways are chain connections of such reactions, same modular approach can be used to build electronic pathways using those basic transistor circuits. Although the idea of creating silicon life seems far-fetched at this stage, this work supplements the idea of cytomorphic chips which is already gaining interest of bio-engineering community.

Introduction: Obesity is an important cause of a range of metabolic diseases. However, the complex mechanisms of obesity and its related diseases make some weight loss methods ineffective or have safety issues. Ginseng, a specialty of Jilin Province in China with both edible andmedicinal value, containsmainly ginsenosides and other components. In order to study the anti-obesity effect of ginseng, network pharmacology was used to predict and screen the active ingredients, action targets and signaling pathways of ginseng. We found (20R)-panaxadiol (PD) is a more desirable active ingredient due to its high drug-like properties and high bioavailability. Moreover, it is closely related to cAMP pathway which is more important in metabolism regulation. The corresponding pharmacodynamic targets of PD include ADRB2 (the gene encoding the ß2-adrenoceptor receptor). Our study aimed to investigate whether Panaxadiol can promote white adipocyte beigeing and increase thermogenesis through modulating the ß2/cAMP pathway to exert antiobesity effects. Methods: In vivo, we established high-fat feeding obesity model, genotypically obese mice (ob/ob) model, and administered PD (10mg/kg). PD treatment in ob/ob mice along with ß2 receptor inhibitor ICI118551. In vitro, differentiated mature 3T3-L1 cells were given palmitate (PA) to induce hypertrophy model along with PD (20 µM). Results: The results of this study demonstrated that PD significantly reduced body weight, improved glucose tolerance and lipid levels in high-fat-induced obese mice and ob/obmice, and also reduced lipid droplet size in PA-treated hypertrophic adipocytes in vitro. Molecular biology assays confirmed that cAMP response element binding protein (CREB) phosphorylation was increased after PD administration, and the expression of thermogenesis-related proteins UCP1, PRDM16 andmitochondrial biosynthesis-related proteins PGC-1a, TFAM and NRF1 were increased. Molecular docking results showed a low binding energy between ß2 receptors and PD, indicating an affinity between the ß2 receptor and PD. In addition, the ß2 receptor inhibition, reversed the anti-obesity effect of PD on the body weight, lipid droplets, the expression of thermogenesis-related proteins and CREB phosphorylation in ob/ob mice. Discussion: These results suggest that PD may promote the expression of thermogenic proteins through phosphorylation of CREB via ß2 receptor activation, and thus exert anti-obesity effects. [ABSTRACT FROM AUTHOR]

Stroke patients show some degree of spontaneous functional recovery, but this is not sufficient to prevent long-term disability. One promising approach is to characterize the dynamics of stroke recovery genes in the lesion and distant areas. We induced sensorimotor cortex lesions in adult C57BL/6J mice using photothrombosis and performed qPCR on selected brain areas at 14, 28, and 56 days post-stroke (P14-56). Based on the grid walk and rotating beam test, the mice were classified into two groups. The expression of cAMP pathway genes Adora2a, Pde10a, and Drd2, was higher in poor- compared to well-recovered mice in contralesional primary motor cortex (cl-MOp) at P14&56 and cl-thalamus (cl-TH), but lower in cl-striatum (cl-Str) at P14 and cl-primary somatosensory cortex (cl-SSp) at P28. Plasticity and axonal sprouting genes, Lingo1 and BDNF, were decreased in cl-MOp at P14 and cl-Str at P28 and increased in cl-SSp at P28 and cl-Str at P14, respectively. In the cl-TH, Lingo1 was increased, and BDNF decreased at P14. Atrx, also involved in axonal sprouting, was only increased in poor-recovered mice in cl-MOp at P28. The results underline the gene expression dynamics and spatial variability and challenge existing theories of restricted neural plasticity. [ABSTRACT FROM AUTHOR]

The follicle-stimulating hormone receptor (FSHR) contains several N-linked glycosylation sites in its extracellular region. We conducted the present study to determine whether conserved glycosylated sites in eel FSHR are necessary for cyclic adenosine monophosphate (cAMP) signal transduction. We used site-directed mutagenesis to induce four mutations (N120Q, N191Q, N272Q, and N288Q) in the N-linked glycosylation sites of eel FSHR. In the eel FSHR wild-type (wt), the cAMP response was gradually increased in a dose-dependent manner (0.01–1500 ng/mL), displaying a high response (approximately 57.5 nM/104 cells) at the Rmax level. Three mutants (N120Q, N272Q, and N288Q) showed a considerably decreased signal transduction as a result of high-ligand treatment, whereas one mutant (N191Q) exhibited a completely impaired signal transduction. The expression level of the N191Q mutant was only 9.2% relative to that of the eel FSHR-wt, indicating a negligible expression level. The expression levels of the N120Q and N272Q mutants were approximately 35.9% and 24% of the FSHG-wt, respectively. The N288Q mutant had an expression level similar to that of the eel FSHR-wt, despite the mostly impaired cAMP responsiveness. The loss of the cell surface agonist-receptor complexes was very rapid in the cells expressing eel FSHR-wt and FSHR-N288Q mutants. Specifically, the N191Q mutant was completely impaired by the loss of cell surface receptors, despite treatment with a high concentration of the agonist. Therefore, we suggest that the N191 site is necessary for cAMP signal transduction. This finding implies that the cAMP response, mediated by G proteins, is directly related to the loss of cell surface receptors as a result of high-agonist treatment. [ABSTRACT FROM AUTHOR]

DNA methylation is an important epigenetic regulatory form that regulates gene expression and tissue development. This study compared the effects of high fiber, low protein (HFLP) and low fiber, high protein (LFHP) diets on the DNA methylation profile of twin lambs' muscles, their effect on glycolysis/gluconeogenesis and related pathways by transcriptome and deep whole-genome bisulfite sequencing (WGBS). Results identified 1,945 differentially methylated regions (DMRs) and 1,471 differentially methylated genes (DMGs). Also, 487 differentially expressed transcripts belonging to 368 differentially expressed genes (DEGs) were discovered between the twin lambs under different diets. Eleven overlapped genes were detected between the DEGs and the DMGs. FKBP5 and FOXO1 were detected to be significantly different. The FOXO1 regulated cAMP and the glycolysis/gluconeogenesis pathways. The glycolysis/gluconeogenesis, and the FOXO pathways were significantly enriched. The expressions of HOMER1 and FOXO1 in the HFLP group were significantly higher than those in the LFHP group. There is a significant correlation between the upregulated gene expression and hypomethylation of HOMER1 and FOXO1 gene in HFLP group. The results showed that FOXO1 induces PDK4 expression in muscle while regulating FKBP5 activity, which stimulates glucose production by activating specific gluconeogenesis target genes. The FOXO1 was able to regulate the glucose metabolism, the cAMP and the occurrence of glycolysis/gluconeogenesis pathways. This study showed that feed type can affect the methylation levels of the glycolysis related gluconeogenesis genes and interaction pathways, providing new ideas for a better understanding of the regulation of muscle energy metabolism and feed development. [ABSTRACT FROM AUTHOR]

Among the most sought after breakthroughs nowadays to combat computational saturation in the electronic hardware realm, neuromorphic and cytomorphic mimetics of biological structures seem potentially promising. Biological circuits are distinguishable due to their minuscule dimensions and immensely low power consumption; yet they achieve extremely complex and magnificent tasks of life, such as, thinking, memorizing, decision making and self-regulating in response to the surroundings. Low power analog circuit solutions are edged over digital ones as they are inherently noisy and fuzzy like bio-systems. In this paper, an analog circuit equivalent for a well-known biological pathway, cyclic adenosine monophosphate (cAMP), has been proposed, exploiting the fabrication characteristics of an analog transistor. The work demonstrates an application of previously published research of the authors, where it was shown that a single transistor operating in analog mode can mimic some fundamental biological circuit processes like receptor-ligand binding, Michaelis Menten and Hill process reactions. Since biological pathways are chain connections of such reactions, same modular approach can be used to build electronic pathways using those basic transistor circuits. Although the idea of creating silicon life seems far-fetched at this stage, this work supplements the idea of cytomorphic chips which is already gaining interest of bio-engineering community. [ABSTRACT FROM AUTHOR]

Endocytosis plays key roles during infection of plant‐pathogenic fungi, but its regulatory mechanisms are still largely unknown. Here, we identified a putative endocytosis‐related gene, PAL1, which was highly expressed in appressorium of Magnaporthe oryzae, and was found to be important for appressorium formation and maturation. Deletion of PAL1 significantly reduced the virulence of M. oryzae due to defects in appressorial penetration and invasive growth in host cells. The Pal1 protein interacted and colocalized with the endocytosis protein Sla1, suggesting it is involved in endocytosis. The Δpal1 mutant was significantly reduced in appressorium formation, which was recovered by adding exogenous cAMP and 3‐isobutyl‐1‐methylxanthine (IBMX). Moreover, the phosphorylation level of Pmk1 in Δpal1 was also reduced, suggesting Pal1 functions upstream of both the cAMP and Pmk1 signalling pathways. As a consequence, the utilization of glycogen and lipid, appressorial autophagy, actin ring formation, localization of septin proteins, as well as turgor accumulation were all affected in the Δpal1 mutant. Taken together, Pal1 regulates cAMP and the Pmk1 signalling pathway for appressorium formation and maturation to facilitate infection of M. oryzae. [ABSTRACT FROM AUTHOR]

Dysregulated platelet aggregation is a fatal condition in many bacterial- and virus-induced diseases. However, classical antithrombotics cannot completely prevent immunothrombosis, due to the unaddressed mechanisms towards inflammation. Thus, targeting platelet hyperactivation together with inflammation might provide new treatment options in diseases, characterized by immunothrombosis, such as COVID-19 and sepsis. The aim of this study was to investigate the antiaggregatory effect and mode of action of 1.8-cineole, a monoterpene derived from the essential oil of eucalyptus leaves, known for its anti-inflammatory proprieties. Platelet activity was monitored by measuring the expression and release of platelet activation markers, i.e., P-selectin, CD63 and CCL5, as well as platelet aggregation, upon treatment with 1.8-cineole and stimulation with several classical stimuli and bacteria. A kinase activity assay was used to elucidate the mode of action, followed by a detailed analysis of the involvement of the adenylyl-cyclase (AC)-cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) pathway by Western blot and ELISA. 1.8-cineole prevented the expression and release of platelet activation markers, as well as platelet aggregation, upon induction of aggregation with classical stimuli and immunological agonists. Mechanistically, 1.8- cineole influences the activation of the AC-cAMP-PKA pathway, leading to higher cAMP levels and vasodilator-stimulated phosphoprotein (VASP) phosphorylation. Finally, blocking the adenosine A 2A receptor reversed the antithrombotic effect of 1.8-cineole. Given the recognized anti-inflammatory attributes of 1.8-cineole, coupled with our findings, 1.8-cineole might emerge as a promising candidate for treating conditions marked by platelet activation and abnormal inflammation. [Display omitted] [ABSTRACT FROM AUTHOR]

Abstract Background Trichoderma reesei represents a model system for investigation of plant cell wall degradation and its connection to light response. The cyclic adenosine monophosphate pathway (cAMP pathway) plays an important role in both physiological outputs, being crucial for regulation of photoreceptor function as well as for cellulase regulation on different carbon sources. Phosphorylation of photoreceptors and of the carbon catabolite repressor CRE1 was shown in ascomycetes, indicating a relevance of protein kinase A in regulation of the target genes of these transcription factors as well as an impact on regulation of induction specific genes. Moreover, the cAMP pathway impacts growth and development. Results Here, we investigated gene regulation by the catalytic subunit of protein kinase A (PKAc1) upon growth on cellulose. We found distinct gene sets for regulation upon growth in light and darkness with an overlap of only 13 genes. PKAc1 regulates metabolic genes as well as transport and defense functions. The overlap of gene regulation by PKAc1 with the genes representing the cAMP dependent regulatory output of the photoreceptor ENV1 indicates an involvement of PKA in this pathway, which counteracts its effects by contrasting regulation. Moreover, we found considerable overlap with the gene sets regulated under cellulase inducing conditions and by the carbon catabolite repressor CRE1. Our analysis also showed that PKAc1 regulates the genes of the SOR cluster associated with the biosynthesis of sorbicillinoids. The homologue of gin4, encoding a CAMK type kinase, which is regulated by PKAc1, CRE1 and YPR2 showed a moderate impact on trichodimerol production. We isolated trichodimerol as representative sorbicillin compound and established a method for its quantification in large sample sets using high performance thin layer chromatography (HPTLC), which can be broadly applied for secondary metabolite screening of mutants or different growth conditions. Due to the high expression levels of the SOR cluster under conditions of sexual development we crosschecked the relevance of PKAc1 under these conditions. We could show that PKAc1 impacts biosynthesis of trichodimerol in axenic growth and upon mating. Conclusions We conclude that PKAc1 is involved in light dependent regulation of plant cell wall degradation, including carbon catabolite repression as well as secondary metabolism and development in T. reesei.

The current study aimed to investigate the mechanism by which miR‐454 influences the progression of heart failure (HF) in relation to the neural precursor cell expressed, developmentally downregulated 4‐2 (NEDD4‐2)/tropomyosin receptor kinase A (TrkA)/cyclic adenosine 3',5'‐monophosphate (cAMP) axis. Sprague‐Dawley rats were used to establish a HF animal model via ligation of the left anterior descending branch of the coronary artery. The cardiomyocyte H9c2 cells were treated with H2O2 to stimulate oxidative stress injury in vitro. RT‐qPCR and Western blot assay were subsequently performed to determine the expression patterns of miR‐454, NEDD4‐2, TrkA, apoptosis‐related proteins and cAMP pathway markers. Dual‐luciferase reporter gene assay coupled with co‐immunoprecipitation was performed to elucidate the relationship between miR‐454, NEDD4‐2 and TrkA. Gain‐ or loss‐of‐function experiments as well as rescue experiments were conducted via transient transfection (in vitro) and adenovirus infection (in vivo) to examine their respective functions on H9c2 cell apoptosis and myocardial damage. Our results suggested that miR‐454 was aberrantly downregulated in the context of HF, while evidence was obtained suggesting that it targeted NEDD4‐2 to downregulate NEDD4‐2 in cardiomyocytes. miR‐454 exerted anti‐apoptotic and protective effects on cardiomyocytes through inhibition of NEDD4‐2, while NEDD4‐2 stimulated ubiquitination and degradation of TrkA protein. Furthermore, miR‐454 activated the cAMP pathway via the NEDD4‐2/TrkA axis, which ultimately suppressed cardiomyocyte apoptosis and attenuated myocardial damage. Taken together, the key findings of the current study highlight the cardioprotective role of miR‐454, which is achieved through activation of the cAMP pathway by impairing NEDD4‐2‐induced TrkA ubiquitination. [ABSTRACT FROM AUTHOR]

Background: Dietary supplementation with L-arginine (Arg) has been shown to increase the volume of fetal fluids in gestating swine. Aquaporins (AQPs), known as water channel proteins, are essential for embryonic growth and development. It was not known if Arg mediates water transport through AQPs in porcine conceptus trophectoderm (pTr2) cells. Methods: pTr2 cells derived from pregnant gilts on day 12 of gestation were cultured in customized Arg-free Dulbecco’s modified Eagle’s Ham medium (DMEM) supplemented with either 0.00, 0.25, or 0.50 mM Arg. Results: Arg treatment increased water transport and the expression of AQP3, which was abundantly expressed in pTr2 cells at both the mRNA and protein levels. Arg also increased the expression of iNOS and the synthesis of nitric oxide (NO) in pTr2 cells. The presence of Nω-nitro-L-arginine methyl ester hydrochloride (L-NAME; an inhibitor of NO synthase) significantly attenuated the Arg-induced expression of AQP3. Furthermore, 0.50 mM Arg increased the concentrations of cAMP and the abundances of phosphorylated cAMP-dependent protein kinase A (PKA), phosphorylated PKA α/β/γ, and phosphorylated CREB. These effects of Arg were mimicked by Forskolin (a cell-permeable activator of adenylyl cyclase), but inhibited by H-89 (an inhibitor of cAMP-dependent protein kinase). Conclusions: The results of this study demonstrate that Arg regulates AQP3 expression and promotes water transport in pTr2 cells through NO- and cAMP-dependent signaling pathways.

Glutathione peroxidases (GPxs) are regarded as important protectors against oxidative stress. Some members of this protein family were reported to play key roles in protecting sperm against oxidative stress. Whether GPx6 a member of the GPx family also plays a role in protection against oxidative stress is not known to date. The objective of the present study was to evaluate the localization and function of glutathione peroxidase 6 (GPx6) in boar accessory sex glands, seminal plasma, and sperm, as well as the effect of GPx6 on vitality and capacitation in boar sperm. qPCR and Western blot analysis demonstrated the presence of GPx6 in testis, epididymis, bulbourethral glands, prostate, seminal vesicle, sperm and seminal plasma. Incubation of sperm with an GPx6 antibody had no significant effect on the viability of boar sperm prior to capacitation. Surprisingly, when capacitated sperm was incubated with the GPx6 antibody for 240 min, sperm vitality was significantly improved. Western blotting showed that in capacitated sperm without prior pretreatment, GPx6 protein content was reduced compared to sperm before capacitation. To further confirm a role for GPx6 in sperm capacitation, we tested sperm acrosome reaction by ACR.2 and FITC-PSA. The results showed that treatment of sperm with the GPx6 antibody significantly increased sperm capacitation and acrosome reaction. Furthermore, we examined the concentration of cAMP in sperm after capacitation. ELISA demonstrated that the cAMP concentration in the sperm exposed to the GPx6 antibody was significantly higher than that of the control group. In addition, the exposure of sperm to the GPx6 antibody significantly increased the concentration of H 2 O 2 , while the expression of SOD3 and CAT were decreased. Based on these observations we would like to postulate that in the boar reproductive tract the GPx6 protein becomes attached to the sperm head preventing the sperm to undergo premature capacitation by affecting components of the antioxidant pathway. How GPx6 expression following ejaculation becomes suppressed to allow sperm capacitation to take place needs further investigation. • Inhibition of GPx6 protein can increase sperm motility after capacitation. • Inhibition of GPx6 protein can promote acrosome reaction of sperm. • GPx6 protein may have the effect of preventing premature capacitation of sperm. [ABSTRACT FROM AUTHOR]

Purpose: There is an increasing agreement that acromegaly caused by growth hormone (GH) cell adenoma has two distinct subtypes: densely granulated (DG) and sparsely granulated (SG). We hypothesized that differential molecular signatures may explain their behavior. Methods: Total transcriptome sequencing was performed on ten DG and seven SG adenomas. The differentially expressed RNAs were identified by bioinformatic analyses, and a candidate RNA was verified by quantitative real-time PCR. Immunohistochemical staining was also performed to detect the protein expression of the candidate. Clinical parameters were correlated with protein expression. Subsequently, cell proliferation, colony formation, and cell cycle progression were analyzed after knockdown of the candidate in pituitary GH3 cells. Activation of the cAMP pathway was assessed by ELISA and Western blot. Results: We confirmed that there were obvious differentially expressed genes between the subtypes. Through gene profiling, we discovered that an orphan adhesion G protein-coupled receptor, GPR64, was overexpressed in more aggressive SG adenomas. Noticeably, GPR64 knockdown significantly inhibited the proliferation of GH3 tumor cells and decreased colony formation. The knockdown also induced cell cycle arrest in GH3 tumor cells. Further studies revealed that GPR64 knockdown decreased cAMP levels and the ratios of p-CREB/CREB, indicating that it suppressed the cAMP/CREB pathway. Conclusions: Our results indicated that GPR64 may promote aggressiveness in SG-type GH cell adenomas and that it is a key factor regulating the cAMP pathway to promote aggressiveness of GH cell adenomas. [ABSTRACT FROM AUTHOR]

Human phosphodiesterases (PDEs) comprise a complex superfamily of enzymes derived from 24 genes separated into 11 PDE gene families (PDEs 1–11), expressed in different tissues and cells, including heart and brain. The isoforms PDE4, PDE7, and PDE8 are specific for the second messenger cAMP, which is responsible for mediating diverse physiological actions involving different hormones and neurotransmitters. The cAMP pathway plays an important role in the development and function of endocrine tissues while phosphodiesterases are responsible for ensuring the appropriate intensity of the actions of this pathway by hydrolyzing cAMP to its inactive form 5'-AMP. PDE1, PDE2, PDE4, and PDE11A are highly expressed in the pituitary, and overexpression of some PDE4 isoforms have been demonstrated in different pituitary adenoma subtypes. This observed over-expression in pituitary adenomas, although of unknown etiology, has been considered a compensatory response to tumorigenesis. PDE4A4/5 has a unique interaction with the co-chaperone aryl hydrocarbon receptor-interacting protein (AIP), a protein implicated in somatotroph tumorigenesis via germline loss-of-function mutations. Based on the association of low PDE4A4 expression with germline AIP-mutation-positive samples, the available data suggest that lack of AIP hinders the upregulation of PDE4A4 protein seen in sporadic somatotrophinomas. This unique disturbance of the cAMP-PDE pathway observed in the majority of AIP-mutation positive adenomas could contribute to their well-described poor response to somatostatin analogs and may support a role in tumorigenesis.

Forskolin is mainly found in the root of a plant called Coleus forskohlii (Willd.) Briq., which has been used in the traditional medicine of Indian Ayurvedic and Southeast Asia since ancient times. Forskolin is responsible for the pharmacological activity of this species. Forskolin is a labdane diterpenoid with a wide biological effect. Several studies suggested a positive role of forskolin on heart complications, respiratory disorders, high blood pressure, obesity, and asthma. There are numerous clinical and pre-clinical studies representing the effect of forskolin on the above-mentioned disorders but more clinical studies need to be performed to support its efficacy. [ABSTRACT FROM AUTHOR]

Human phosphodiesterases (PDEs) comprise a complex superfamily of enzymes derived from 24 genes separated into 11 PDE gene families (PDEs 1–11), expressed in different tissues and cells, including heart and brain. The isoforms PDE4, PDE7, and PDE8 are specific for the second messenger cAMP, which is responsible for mediating diverse physiological actions involving different hormones and neurotransmitters. The cAMP pathway plays an important role in the development and function of endocrine tissues while phosphodiesterases are responsible for ensuring the appropriate intensity of the actions of this pathway by hydrolyzing cAMP to its inactive form 5'-AMP. PDE1, PDE2, PDE4, and PDE11A are highly expressed in the pituitary, and overexpression of some PDE4 isoforms have been demonstrated in different pituitary adenoma subtypes. This observed over-expression in pituitary adenomas, although of unknown etiology, has been considered a compensatory response to tumorigenesis. PDE4A4/5 has a unique interaction with the co-chaperone aryl hydrocarbon receptor-interacting protein (AIP), a protein implicated in somatotroph tumorigenesis via germline loss-of-function mutations. Based on the association of low PDE4A4 expression with germline AIP -mutation-positive samples, the available data suggest that lack of AIP hinders the upregulation of PDE4A4 protein seen in sporadic somatotrophinomas. This unique disturbance of the cAMP-PDE pathway observed in the majority of AIP -mutation positive adenomas could contribute to their well-described poor response to somatostatin analogs and may support a role in tumorigenesis. [ABSTRACT FROM AUTHOR]

Forskolin is mainly found in the root of a plant called Coleus forskohlii (Willd.) Briq., which has been used in the traditional medicine of Indian Ayurvedic and Southeast Asia since ancient times. Forskolin is responsible for the pharmacological activity of this species. Forskolin is a labdane diterpenoid with a wide biological effect. Several studies suggested a positive role of forskolin on heart complications, respiratory disorders, high blood pressure, obesity, and asthma. There are numerous clinical and pre-clinical studies representing the effect of forskolin on the above-mentioned disorders but more clinical studies need to be performed to support its efficacy.

Ethanol's aversive property may limit it's use, but the underlying mechanisms are no well-understood. Emerging evidence suggests a critical role for the lateral habenula (LHb) in the aversive response to various drugs, including ethanol. We previously showed that ethanol enhances glutamatergic transmission and stimulates LHb neurons. GABAergic transmission, a major target of ethanol in many brain regions, also tightly regulates LHb activity. This study assessed the action of ethanol on LHb GABAergic transmission in rat brain slices. Application of ethanol accelerated spontaneous action potential firing of LHb neurons, and LHb activity was increased by the GABA A receptor antagonist gabazine, and ethanol-induced acceleration of LHb firing was further increased by gabazine. Additionally, ethanol potentiated GABAergic transmission (inhibitory postsynaptic currents, IPSCs) with an EC 50 of 1.5 mM. Ethanol-induced potentiation of IPSCs was increased by a GABA B receptor antagonist; it was mimicked by dopamine, dopamine receptor agonists, and dopamine reuptake blocker, and was completely prevented by reserpine, which depletes store of catecholamine. Moreover, ethanol-induced potentiation of IPSCs involved cAMP signaling. Finally, ethanol enhanced simultaneously glutamatergic and GABAergic transmissions to the majority of LHb neurons: the potentiation of the former being greater than that of the latter, the net effect was increased firing. Since LHb excitation may contribute to aversion, ethanol-induced potentiation of GABAergic inhibition tends to reduce aversion. [ABSTRACT FROM AUTHOR]

The remarkable ability of the nervous system to modify its structure and function is mostly experience and activity modulated. The molecular basis of neuronal plasticity has been studied in higher behavioral processes, such as learning and memory formation. However, neuronal plasticity is not restricted to higher brain functions, but may provide a basic feature of adaptation of all neural circuits. The fruit fly Drosophila melanogaster provides a powerful genetic model to gain insight into the molecular basis of nervous system development and function. The nervous system of the larvae is again a magnitude simpler than its adult counter part, allowing the genetic assessment of a number of individual genetically identifiable neurons. We review here recent progress on the genetic basis of neuronal plasticity in developing and functioning neural circuits focusing on the simple visual system of the Drosophila larva.

Three-dimensional (3D) spheroid culture has recently become a valuable tool to evaluate the utility of treatments that better mirror biological features and exhibit high concordance with in vivo conditions. Here, by using the 3D spheroid method, we investigated cell morphology, proliferation, melanin content, tyrosinase activity, and cAMP production. During 3D speroid cultures, melanin biosynthesis was observed and the amount of melanin continuously increased until cell proliferation slowed down. Up-regulation of the cAMP pathway enhanced tyrosinase activity, allowing melanin synthesis. However, after treatment with depigmenting agents, melanin content was reduced. These results indicate that 3D spheroid culture method is an important tool to study melanin biosynthesis, which enables us to overcome several limitations found in the 2D adherent cell culture mehtod. [ABSTRACT FROM AUTHOR]

Hyperalgesia often occurs in opioid-induced withdrawal syndrome. In the present study, we found that three hourly injections of DAMGO (a µ-opioid receptor agonist) followed by naloxone administration at the fourth hour significantly decreased rat paw nociceptive threshold, indicating the induction of withdrawal hyperalgesia. Application of NaHS (a hydrogen sulfide donor) together with each injection of DAMGO attenuated naloxone-precipitated withdrawal hyperalgesia. RT-PCR and Western blot analysis showed that NaHS significantly reversed the gene and protein expression of up-regulated spinal calcitonin gene-related peptide (CGRP) in naloxone-treated animals. NaHS also inhibited naloxone-induced cAMP rebound and cAMP response element-binding protein (CREB) phosphorylation in rat spinal cord. In SH-SY5Y neuronal cells, NaHS inhibited forskolin-stimulated cAMP production and adenylate cyclase (AC) activity. Moreover, NaHS pre-treatment suppressed naloxone-stimulated activation of protein kinase C (PKC) α, Raf-1, and extracellular signal-regulated kinase (ERK) 1/2 in rat spinal cord. Our data suggest that H2S prevents the development of opioid withdrawal-induced hyperalgesia via suppression of synthesis of CGRP in spine through inhibition of AC/cAMP and PKC/Raf-1/ERK pathways. [ABSTRACT FROM AUTHOR]

In the principal neurons of striatum (medium spiny neurons, MSNs), cAMP pathway is primarily activated through the stimulation of dopamine D1 and adenosine A2A receptors, these receptors being mainly expressed in striatonigral and striatopallidal MSNs, respectively. Since cAMP signaling pathway could be altered in various physiological and pathological situations, including drug addiction and Parkinson’s disease, it is of crucial importance to identify the molecular components involved in the activation of this pathway. In MSNs, cAMP pathway activation is not dependent on the classical Gs GTP-binding protein but requires a specific G protein subunit heterotrimer containing Galpha-olf/beta2/gamma7 in particular association with adenylate cyclase type 5. This assembly forms an authentic functional signaling unit since loss of one of its members leads to defects of cAMP pathway activation in response to D1 or A2A receptor stimulation, inducing dramatic impairments of behavioral responses dependent on these receptors. Interestingly, D1 receptor-dependent cAMP signaling is modulated by the neuronal levels of Galpha-olf, indicating that Galpha-olf represents the rate-limiting step in this signaling cascade and could constitute a critical element for regulation of D1 receptor responses. In both Parkinsonian patients and several animal models of Parkinson’s disease, the lesion of dopamine neurons produces a prolonged elevation of Galpha-olf levels. This observation gives an explanation for the cAMP pathway hypersensitivity to D1 stimulation, occurring despite an unaltered D1 receptor density. In conclusion, alterations in the highly specialized assembly of Galpha-olf/beta2/gamma7 subunits can happen in pathological conditions, such as Parkinson’s disease, and it could have important functional consequences in relation to changes in D1 receptor signaling in the striatum.

The remarkable ability of the nervous system to modify its structure and function is mostly experience and activity modulated. The molecular basis of neuronal plasticity has been studied in higher behavioral processes, such as learning and memory formation. However, neuronal plasticity is not restricted to higher brain functions and it may provide a basic feature of adaptation of all neural circuits. The fruit fly Drosophila melanogaster provides a powerful genetic model to gain insight into the molecular basis of nervous system development and function. The nervous system of the larvae is again a magnitude simpler than its adult counter part, allowing the genetic assessment of a number of individual genetically identifiable neurons. We review here recent progress on the genetic basis of neuronal plasticity in developing and functioning neural circuits focusing on the simple visual system of the Drosophila larva. [ABSTRACT FROM AUTHOR]

BACKGROUND: H2S is a novel type of endogenous neural regulatory factor and gaseous mediator. Exogenous H2S can increase heroin-induced learning and memory damage in rat and alleviates heroin-induced rat hippocampus damage through antioxidant and anti-apoptosis effects. OBJECTIVE: Aim of this study was to identify whether hydrogen sulfide (H2S) protects heroin withdrawal rat is related with adenylate cyclase (AC)-cAMP-protein kinase A (PKA)-cAMP response element-binding protein (CREB) signaling pathway in heroin-dependent rat's nucleus accumbens or not. METHODS: Male Spragne-Dawley rats were randomly divided into Saline + Saline group, Saline + sodium hydrosulfide (NaHS) group, Saline + Heroin group, NaHS + Heroin group according to the principle of increasing heroin dosage day by day, with the establishment of heroin-naloxone-induced withdrawal symptoms determined at day 10. Then the levels of H2S and cAMP and AC and PKA activities were assayed, and the level of phosphorylated CREB (p-CREB), the levels of phosphorylated N-methyl-D-aspartate receptor 1 subunit (p-NR1), phosphorylated N-methyl-D-aspartate receptor 2a subunit (p-NR2A) and phosphorylated N-methyl-D-aspartate receptor 2b subunit (p-NR2B) were assayed in nucleus accumbens. RESULTS: Exogenous H2S can alleviate heroin withdrawal symptoms by increasing the level of H2S level in nucleus accumbens. Exogenous H2S can decrease the high activities of AC, PKA and the high levels of cAMP, p-CREB caused by heroin. Furthermore, exogenous H2S can decrease the high level of p-NR1 and can increase the low levels of p-NR2A and p-NR2B caused by heroin. It is surprising that exogenous H2S treatment alone was able to raise the activities of AC and PKA as well as the levels of cAMP, p-CREB, p-NR1, p-NR2A and p-NR2B. CONCLUSIONS: Exogenous H2S decreases naloxone-precipitated withdrawal signs, maybe through decreasing AC/cAMP/PKA/CREB/NMDR signaling pathway in heroin-dependent rats' nucleus accumbens. [ABSTRACT FROM AUTHOR]

BACKGROUND: H2S is a novel type of endogenous neural regulatory factor and gaseous mediator. Exogenous H2S can increase heroin-induced learning and memory damage in rat and alleviates heroin-induced rat hippocampus damage through antioxidant and anti-apoptosis effects. OBJECTIVE: Aim of this study was to identify whether hydrogen sulfide (H2S) protects heroin withdrawal rat is related with adenylate cyclase (AC)-cAMP-protein kinase A (PKA)-cAMP response element-binding protein (CREB) signaling pathway in heroin-dependent rat's nucleus accumbens or not. METHODS: Male Spragne-Dawley rats were randomly divided into Saline + Saline group, Saline + sodium hydrosulfide (NaHS) group, Saline + Heroin group, NaHS + Heroin group according to the principle of increasing heroin dosage day by day, with the establishment of heroin-naloxone-induced withdrawal symptoms determined at day 10. Then the levels of H2S and cAMP and AC and PKA activities were assayed, and the level of phosphorylated CREB (p-CREB), the levels of phosphorylated N-methyl-Daspartate receptor 1 subunit (p-NR1), phosphorylated N-methyl-D-aspartate receptor 2a subunit (p-NR2A) and phosphorylated N-methyl-Daspartate receptor 2b subunit (p-NR2B) were assayed in nucleus accumbens. RESULTS: Exogenous H2S can alleviate heroin withdrawal symptoms by increasing the level of H2S level in nucleus accumbens. Exogenous H2S can decrease the high activities of AC, PKA and the high levels of cAMP, p-CREB caused by heroin. Furthermore, exogenous H2S can decrease the high level of p-NR1 and can increase the low levels of p-NR2A and p-NR2B caused by heroin. It is surprising that exogenous H2S treatment alone was able to raise the activities of AC and PKA as well as the levels of cAMP, p-CREB, p-NR1, p-NR2A and p-NR2B. CONCLUSIONS: Exogenous H2S decreases naloxone-precipitated withdrawal signs, maybe through decreasing AC/cAMP/PKA/CREB/NMDR signaling pathway in heroin-dependent rats' nucleus accumbens. [ABSTRACT FROM AUTHOR]

The gonadotropin-releasing hormone (GnRH) pulsatile pattern is critical for appropriate regulation of gonadotrope activity but only little is known about the signaling mechanisms by which gonadotrope cells decode such pulsatile pattern. Here, we review recent lines of evidence showing that the GnRH receptor (GnRH-R) activates the cyclic AMP (cAMP) pathway in gonadotrope cells, thus ending a long-lasting controversy. Interestingly, coupling of GnRH-R to the cAMP pathway as well as induction of nitric oxide synthase 1 (NOS1) or follistatin through this signaling pathway take place preferentially under high GnRH pulsatility. The preovulatory surge of GnRH in vivo is indeed associated with an important increase of pituitary cAMP and NOS1 expression levels, both being markedly inhibited by treatment with a GnRH antagonist. Altogether, this suggests that due to its atypical structure and desensitization properties, the GnRH-R may continue to signal through the cAMP pathway under conditions inducing desensitization for most other receptors. Such a mechanism may contribute to decode high GnRH pulsatile pattern and enable gonadotrope cell plasticity during the estrus cycle. [ABSTRACT FROM AUTHOR]

Abstract: Sensing of environmental signals is often mediated by G-protein coupled receptors and their cognate heterotrimeric G-proteins. In Trichoderma reesei (Hypocrea jecorina) the signals transmitted via the G-protein alpha subunits GNA1 and GNA3 cause considerable modulation of cellulase transcript levels and the extent of this adjustment is dependent on the light status. We therefore intended to elucidate the underlying mechanism connecting light response and heterotrimeric G-protein signaling. Analysis of double mutant strains showed that constitutive activation of GNA1 or GNA3 in the absence of the PAS/LOV domain protein ENVOY (ENV1) leads to the phenotype of constitutive G-alpha activation in darkness. In light, however the deletion-phenotype of Δenv1 was observed with respect to growth, conidiation and cellulase gene transcription. Additionally deletion of env1 causes decreased intracellular cAMP accumulation, even upon constitutive activation of GNA1 or GNA3. While supplementation of cAMP caused an even more severe growth phenotype of all strains lacking env1 in light, addition of the phosphodiesterase inhibitor caffeine rescued the growth phenotype of these strains. ENV1 is consequently suggested to connect the light response pathway with nutrient signaling by the heterotrimeric G-protein cascade by adjusting transcript levels of gna1 and gna3 and action on cAMP levels – presumably through inhibition of a phosphodiesterase. [Copyright &y& Elsevier]

We compare here the neurodegenerative processes observed in the hippocampus of bitransgenic mice with chronically altered levels of cAMP-response element-binding protein (CREB) function. The combination of genome-wide transcriptional profiling of degenerating hippocampal tissue with microscopy analyses reveals that the sustained inhibition of CREB function in A-CREB mice is associated with dark neuron degeneration, whereas its strong chronic activation in VP16-CREB mice primarily causes excitotoxic cell death and inflammation. Furthermore, the meta-analysis with gene expression profiles available in public databases identifies relevant common markers to other neurodegenerative processes and highlights the importance of the immune response in neurodegeneration. Overall, these analyses define the ultrastructural and transcriptional signatures associated with these two forms of hippocampal neurodegeneration, confirm the importance of fine-tuned regulation of CREB-dependent gene expression for CA1 neuron survival and function, and provide novel insight into the function of CREB in the etiology of neurodegenerative processes. [ABSTRACT FROM AUTHOR]

The melanocortin 1 receptor gene is a main determinant of human pigmentation, and a melanoma susceptibility gene, because its variants that are strongly associated with red hair color increase melanoma risk. To test experimentally the association between melanocortin 1 receptor genotype and melanoma susceptibility, we compared the responses of primary human melanocyte cultures naturally expressing different melanocortin 1 receptor variants to α-melanocortin and ultraviolet radiation. We found that expression of 2 red hair variants abolished the response to α-melanocortin and its photoprotective effects, evidenced by lack of functional coupling of the receptor, and absence of reduction in ultraviolet radiation-induced hydrogen peroxide generation or enhancement of repair of DNA photoproducts, respectively. These variants had different heterozygous effects on receptor function. Microarray data confirmed the observed differences in responses of melanocytes with functional vs. nonfunctional receptor to α-melanocortin and ultraviolet radiation, and identified DNA repair and antioxidant genes that are modulated by α-melanocortin. Our findings highlight the molecular mechanisms by which the melanocortin 1 receptor genotype controls genomic stability of and the mutagenic effect of ultraviolet radiation on human melanocytes. [ABSTRACT FROM AUTHOR]

Abstract: The resistance to thyrotropin (TSH) action is the disease associated with molecular defects hampering the adequate transmission of TSH stimulatory signal into thyroid cells. The defect may in principle affect every step along the cascade of events following the binding of TSH to its receptor (TSHR) on thyroid cell membranes. After the description of the first family affected with loss-of-function (LOF) TSHR mutations in 1995, there is now evidence that TSH resistance is a disease with a broad range of expressivity going from severe congenital hypothyroidism (CH) with thyroid hypoplasia to mild hyperthyrotropinemia (hyperTSH) associated with an apparent euthyroid state. More severe forms occur in patients with disrupting biallelic TSHR mutations and follow a recessive pattern of inheritance. Differential diagnosis in these cases includes the exclusion of other causes of thyroid dysgenesis, such as mutations in thyroid transcription factors. More mild forms may instead occur in patients with monoallelic TSHR defects following a dominant mode of inheritance. In these cases we described the dominant negative effect exerted by some LOF mutants on the activity of the wild-type TSHR. Differential diagnosis involves the exclusion of mild hypothyroidism in autoimmune thyroid disease or pseudohypoparathyroidism associated with genetic or epigenetic defects at the GNAS locus. This review will focus on the prevalence of TSHR mutations, on the molecular mechanisms leading to TSH resistance and on the variable clinical expression of this disease. [Copyright &y& Elsevier]

Abstract: To develop intracellularly within phagocytes and cause chronic infection, Brucella must overcome different steps of the host immune responses. IFNγ is a key mediator of the innate and adaptive responses produced during Brucella infection. Therefore, Brucella would control host defenses by impairing macrophage responses to IFNγ. We first showed that in infected human macrophages (VD3-differentiated THP-1 cells) Brucella escaped the microbicidal environment generated by IFNγ. We then analyzed the IFNγ-mediated signaling in Brucella-infected cells. We observed no decrease in STAT1 tyrosine or serine phosphorylation, or in dimerization of phosphorylated STAT1 (P-STAT1) and P-STAT1 translocation to the nucleus or in P-STAT1 binding to GAS, a minimal IFNγ-response DNA sequence. In contrast, immuno-precipitation experiments indicated that the IFNγ-mediated association of P-STAT1 with CBP/P300 transactivators was markedly reduced in infected macrophages, demonstrating that P-STAT1 was unable to normally recruit these transactivators. The host cell cAMP pathway triggered by Brucella could be responsible for this defect, CBP/P300 mobilization by phosphorylated CREB (P-CREB) disrupting the IFNγ-induced STAT1-CBP/P300 association, required for a normal response of macrophages to IFNγ. In any case, the inhibition of an essential protein–protein interaction probably lead to a deteriorated response to IFNγ and thus participated in the pathogen''s establishment within its host. [Copyright &y& Elsevier]