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Delta opioid receptors (DORs) are widely expressed throughout the central nervous system, including the cerebellum, where they play a regulatory role in neurogenesis. In the cerebellar cortex, Purkinje cells (PCs), the sole output neurons, receive glutamatergic synaptic input from parallel fibers (PFs)-the axonal extensions of granule cells-forming PF-PC synapses. However, the precise distribution of DORs within these synapses and their impact on synaptic transmission remain unclear. In this study, we utilized whole-cell patch-clamp recordings and neuropharmacological approaches to explore the effects of DORs activation on PF-PC synaptic transmission in the mouse cerebellar cortex and to elucidate the underlying mechanisms. We found that the selective DORs agonist DPDPE significantly reduced the amplitude and area under the curve (AUC) of PF-PC evoked excitatory postsynaptic currents (eEPSCs), accompanied by an increase in the paired-pulse ratio (PPR). This inhibitory effect was blocked by the DORs antagonist Naltrindole. Additionally, DPDPE decreased the frequency of PF-PC miniature excitatory postsynaptic currents (mEPSCs) without affecting their amplitude, indicating a presynaptic site of action. When the protein kinase A (PKA) inhibitor PKI was added to the internal solution of the recording electrode, it did not alter the DPDPE-induced suppression of PF-PC mEPSC frequency. However, this suppression was reversed by KT5720, a cell-permeable PKA-specific inhibitor. These findings suggest that DPDPE inhibits PF-PC synaptic transmission through the preferential activation of presynaptic DORs, with this process being dependent on the cyclic adenosine monophosphate (cAMP)-PKA signaling pathway., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Ethnopharmacological Relevance: Syringa oblata Lindl (ZDX) is a plant in the Oleaceae family that is the primary ingredient in the classic Tibetan medicine AKARU sinensis. The plant's stem is used as a medicine, and Tibetan doctors often use it as a sedative, a use with a history of nearly 100 years. Tibetan medicine mainly uses lilac to treat headache, forgetfulness, insomnia, irritability and other symptoms. Depression is a chronic mental disorder characterized by low mood, cognitive impairments, and physical discomfort, and it has become a significant public health issue. Given the limitations of existing treatments, interest in alternative therapies, including herbal medicines, is increasing., Aim: To elucidate the mechanism of ZDX extract in the treatment of depression., Materials and Methods: A depression-like mouse model was established via the subcutaneous injection of corticosterone (CORT) into the groin, and a model of PC12 cell injury was established via CORT treatment. The antidepressant effect of the ZDX extract was subsequently evaluated via weight measurements, the sucrose preference test (SPT), the forced swimming test (FST), the open field test(OFT), the tail suspension test (TST), HE staining, Nissl staining and ELISA. Moreover, immunofluorescence staining, qRT‒PCR and Western blotting were used to determine whether ZDX extract can regulate the cAMP/PKA-CREB-BDNF pathway to prevent depression and neuronal apoptosis., Results: ZDX extract significantly improved depression-like behaviours; inhibited decreases in the protein levels of cAMP, PKA, CREB and BDNF; and increased proliferative activity in the hippocampus and cortex. In addition, in vitro, ZDX extract attenuated CORT-induced injury and apoptosis in hippocampal neurons and inhibited CORT-induced decreases in the mRNA expression levels of cAMP, PKA, CREB and BDNF., Conclusions: These findings suggest that ZDX extract has potential as a novel antidepressant therapeutic agent, offering a complementary approach to current treatments by targeting multiple pathways involved in the pathogenesis of depression. Further research is warranted to explore the clinical applications of ZDX extract in the treatment of depression., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Objective: Hydrolyzed conchiolin protein (HCP) derived from pearl and nacre extracts exerts skin-lightening effects; however, the underlying molecular mechanisms are not fully understood. Herein, we investigated the effect of HCP on melanogenesis and the signalling pathways involved., Methods: B16F10 cells and PIG cells were treated with HCP to verify its ability to inhibit melanin. Western Blot, immunofluorescence, and flow cytometry methods were performed to investigate the effect of HCP on melanogenesis signalling pathway proteins. The inhibitors were used to further validate the effect of HCP on PKA/CREB and MEK/ERK signalling pathways. To further evaluate the whitening ability of HCP, changes in melanin were detected using 3D melanin skin model and zebrafish model., Results: HCP was found to significantly inhibit melanin synthesis and decrease the expression of melanogenesis-related proteins, such as microphthalmia-associated transcription factor (MITF), tyrosinase, and tyrosinase-related protein-2, in a dose-dependent manner. Additionally, we revealed that HCP suppresses melanogenesis via the regulation of the PKA/cAMP response element-binding (CREB) and MEK/extracellular signalling-regulated kinase (ERK) signalling pathways. Using 3D melanin skin models, we demonstrated that HCP can achieve skin-lightening effects by improving apparent chroma, increasing apparent brightness, and inhibiting melanin synthesis. Furthermore, HCP exhibits skin-whitening effects in a zebrafish model., Conclusion: These results suggest that HCP suppresses the melanogenesis signalling cascade by inhibiting the PKA/CREB, MEK/ERK signalling pathway and downregulating MITF and its downstream signalling pathways, resulting in decreased melanin synthesis. In summary, HCP is a potential anti-pigmentation agent with promising applications in cosmetics and pharmaceutical products., (© 2024 The Author(s). International Journal of Cosmetic Science published by John Wiley & Sons Ltd on behalf of Society of Cosmetic Scientists and Societe Francaise de Cosmetologie.)

AKAPs (A-kinase-anchoring proteins) act as scaffold proteins that anchor the regulatory subunits of the cAMP-dependent PKA (protein kinase A) to coordinate and compartmentalize signaling elements and signals downstream of Gs-coupled GPCRs (G protein-coupled receptors). The β 2 AR (β-2-adrenoceptor), as well as the Gs-coupled EP2 and EP4 (E-prostanoid) receptor subtypes of the EP receptor subfamily, are effective regulators of multiple airway smooth muscle (ASM) cell functions whose dysregulation contributes to asthma pathobiology. Here, we identify specific roles of the AKAPs Ezrin and Gravin in differentially regulating PKA substrates downstream of the β 2 AR, EP2R (EP2 receptor) and EP4R. Knockdown of Ezrin, Gravin, or both in primary human ASM cells caused differential phosphorylation of the PKA substrates VASP (vasodilator-stimulated phosphoprotein) and HSP20 (heat shock protein 20). Ezrin knockdown, as well as combined Ezrin and Gravin knockdown, significantly reduced the induction of phospho-VASP and phospho-HSP20 by β 2 AR, EP2R, and EP4R agonists. Gravin knockdown inhibited the induction of phospho-HSP20 by β 2 AR, EP2R, and EP4R agonists. Knockdown of Ezrin, Gravin, or both also attenuated histamine-induced phosphorylation of MLC20. Moreover, knockdown of Ezrin, Gravin, or both suppressed the inhibitory effects of Gs-coupled receptor agonists on cell migration in ASM cells. These findings demonstrate the role of AKAPs in regulating Gs-coupled GPCR signaling and function in ASM and suggest the therapeutic utility of targeting specific AKAP family members in the management of asthma.

Leydig cells (LCs) injury is often irreversible upon discovery; hence, early identification of risk factors for injury is crucial. The ubiquitous plasticizer di-2-ethylhexyl phthalate (DEHP) in the environment has been shown to potentially cause damage to LCs. However, the underlying mechanisms remain unclear. The present study utilized scRNA-seq analysis, the advantage of which is the ability to explore the characteristics of various testicular cells, combined with studies in vitro and in vivo, to assay the changes in and damage processes of LCs during DEHP exposure. We found that DEHP disrupted the structure and function of LCs. GO analysis suggested that a series of pathways changed, among which the most significant were the "steroid synthesis" and "oxidative stress" pathways. Moreover, DEHP dramatically changed the manner of interaction between LCs and other cells, and the most significant type was the cell-cell contact, which included NECTIN, APP, CADM, and CD39. In addition, the activity of multiple transcription factors (TFs) decreased after DEHP exposure, and the activity of steroidogenic factor 1 (SF-1, Nr5a1) was the most obviously altered. Next, we found that the LCs region indeed experienced oxidative stress, including increased ROS signals, the decreased SOD activity and T-AOC, and increased concentration of 8-OHdG and MDA content. The testosterone level, as well as the expression of StAR, P450scc, and 3β-HSD, was also reduced. To study the association between testosterone synthesis and oxidative stress, the antioxidants N-acetyl-L-cysteine (NAC) and H 2 O 2 were used, and we found that mono-2-ethylhexyl ester (MEHP, a major biometabolite of DEHP) disrupted testosterone synthesis through the inhibition of the cAMP/PKA/SF-1/StAR pathway by inducing oxidative stress. Our study provides new insights into the role and mechanisms of DEHP in LCs injury., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Patients with diabetes mellitus show an elevated prevalence of psychiatric disorders such as anxiety. We have reported that fear memory, a model related to anxiety as reflected in the freezing response, is enhanced in diabetic mice and was ameliorated by an AMPA receptor antagonist. The present study investigated whether functions of AMPA receptors in the amygdala and hippocampus are altered in streptozotocin (STZ)-induced diabetic mice. While protein levels of the GluA1 subunit of AMPA receptors were not altered in the amygdala and hippocampus, protein levels of GluA1 phosphorylated at serine 845 in the amygdala and hippocampus and of GluA1 phosphorylated at serine 831 in the hippocampus were increased in STZ-induced diabetic mice. L-lactate, which is increased in the amygdala and hippocampus of STZ-induced diabetic mice, did not alter these protein levels in either brain area. In contrast, protein levels of phosphorylated protein kinase A (PKA) catalytic subunit and phosphorylated calcium calmodulin kinase II (CaMKII), which are known to phosphorylate serine 845 and serine 831 of GluA1, respectively, were increased in the amygdala and hippocampus of STZ-induced diabetic mice. In the fear memory test, the PKA inhibitor H-89 injected before test sessions and the CaMKII inhibitor KN-62 injected before conditioning or test sessions each reduced the increase in freezing in STZ-induced diabetic mice. These results indicate that the functions of AMPA receptors in the amygdala and hippocampus are enhanced due to increased phosphorylation by PKA and CaMKII, which enhances fear memory in diabetic mice., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Stress can alter behavior and contributes to psychiatric disorders by regulating the expression of the GluA2 AMPA receptor subunit. We have previously shown in mice that exposure to predator odor stress elevates GluA2 transcription in cerebellar molecular layer interneurons (MLIs), and MLI activity is required for fear memory consolidation. Here, we identified the critical involvement of adenylyl cyclase 5, in both the stress-induced increase in GluA2 in MLIs and the enhancement of fear memory. We found that noradrenaline release during predator odor stress activates AC5 and downstream PKA-CREB signaling. This pathway interacts synergistically with α1-adrenergic receptors to promote synaptic GluA2 expression in MLIs. At a behavioral level, predator odor stress potentiates associative fear memory, and this is abolished in AC5 knockout mice, suggesting that AC5-dependent plasticity is required for enhanced memory formation. Therefore, AC5 is a promising pharmacological target for preventing stress-enhanced fear memory., Competing Interests: Declaration of interests The authors declare no competing interests., (Published by Elsevier Inc.)

Neurotransmitters and neuromodulators can be released via either action potential (AP)-evoked transient or AP-independent continuous neurotransmission. The elevated AP-evoked neurotransmission in the primary sensory neurons plays crucial roles in hyperalgesia. However, whether and how the AP-independent continuous neurotransmission contributes to hyperalgesia remains largely unknown. Here, we show that primary sensory dorsal root ganglion (DRG) neurons exhibit frequent spontaneous microdomain Ca 2+ (smCa) activities independent of APs across the cell bodies and axons, which are mediated by the spontaneous opening of TRPA1 channels and trigger continuous neurotransmission via the cyclic adenosine monophosphate-protein kinase A signaling pathway. More importantly, the frequency of smCa activity and its triggered continuous neurotransmission in DRG neurons increased dramatically in mice experiencing inflammatory pain, inhibition of which alleviates hyperalgesia. Collectively, this work revealed the AP-independent continuous neurotransmission triggered by smCa activities in DRG neurons, which may serve as a unique mechanism underlying the nociceptive sensitization in hyperalgesia and offer a potential target for the treatment of chronic pain., Competing Interests: Competing interests statement:The authors declare no competing interest.

This study aimed to investigate the role of transforming growth factor-beta 3 (TGF-β3) secreted by adipose-derived stem cells (ADSCs) in suppressing melanin synthesis during the wound healing process, particularly in burn injuries, and to explore the underlying mechanisms involving the cAMP/PKA signaling pathway. ADSCs were isolated from C57BL/6 mice and characterized using flow cytometry and differentiation assays. A burn injury model was established in mice, followed by UVB irradiation to induce hyperpigmentation. Mice were divided into four groups: control (PBS), ADSCs, ADSCs with TGF-β3 overexpression, and ADSCs with TGF-β3 knockdown. Melanin deposition was assessed via Fontana Masson staining, while ELISA measured the expression of MC1R and α-MSH. Western blotting was used to examine TGF-β3 and the activation of cAMP/PKA signaling pathway. Mice treated with ADSCs overexpressing TGF-β3 showed significant reductions in melanin deposition and suppressed expression of MC1R and α-MSH compared to controls. Western blot results revealed that the cAMP/PKA signaling pathway was downregulated in the TGF-β3-overexpressing ADSC group. In contrast, the knockdown of TGF-β3 led to increased melanin deposition and higher cAMP/PKA pathway activity, correlating with greater expression of MC1R and α-MSH. TGF-β3 secreted by ADSCs effectively inhibits melanin synthesis during wound healing, with potential effects on the cAMP/PKA signaling pathway. These findings suggest that TGF-β3 could serve as a potential therapeutic target for managing post-burn hyperpigmentation., Competing Interests: Declarations. Ethics approval and consent to participate: This study followed international guidelines for animal research projects and was approved by the Animal Ethics Committee of the Third Hospital of Hebei Medical University. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Aims: Posttraumatic stress disorder (PTSD) is a debilitating neuropsychiatric illness caused by traumatic or life-threatening events and manifesting as various symptoms, including intrusive re-experiences of trauma, avoidance behaviors, hyperarousal, and negative changes in perception and mood., Main Methods: Current monoamine-based medications commonly exhibit limited efficacy and significant side effects, which hamper their clinical utility. To address this unmet need, we explored 4-methoxycinnamic acid (4-MCA) as a potential novel treatment for PTSD in a single prolonged stress (SPS)-induced animal model., Key Findings: Administration of 4-MCA (3 and 10 mg/kg, p.o.) significantly mitigated anxiety-like behaviors, alleviated depression-like behaviors, and improved cognitive function in an SPS-treated PTSD mouse model. Further, 4-MCA treatment effectively rectified the fear extinction deficits in the fear conditioning test. Molecular analyses revealed that 4-MCA normalized the elevated corticotropin-releasing hormone (CRH) levels as well as the phosphorylation of protein kinase A (PKA) and cAMP response element-binding protein (CREB) in the amygdala, a pivotal region for fear memory formation. Co-administration of 4-MCA and the CRFR1 antagonist antalarmin at subeffective doses facilitated fear memory extinction., Significance: These findings suggest that 4-MCA alleviates SPS-induced PTSD-like behaviors by regulating the CRH-CRFR1-PKA-CREB signaling pathway in the amygdala, and that 4-MCA may be a potential candidate for future PTSD treatment., Competing Interests: Declaration of competing interest The authors have no conflicts of interest relevant to this study to disclose., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Obesity-associated cardiac remodeling is characterized by cardiac sympathetic nerve over-activation and pro-inflammatory macrophage infiltration. We identified norepinephrine (NE), a sympathetic neurotransmitter, as a pro-inflammatory effector to activate macrophage NLRP3 inflammasome, which contributed to cardiac inflammation. In vivo, Sprague-Dawley (SD) rats were fed a high-fat diet (HFD) for 12 weeks to establish obese rat models. Obese rats exhibited marked cardiac hypertrophy compared to normal rats. The expression of NLRP3 and interleukin (IL)-1β was upregulated, accompanied by CD68 + NLRP3 + macrophage infiltration in the hearts of the obese rats. The obese rats also showed increased sympathetic nerve activity. β-adrenergic receptor (AR) inhibition mitigated these changes. In vitro, sympathetic neurotransmitter NE significantly exacerbated palmitic acid (PA)-induced macrophage polarization toward pro-inflammatory type and NLRP3 inflammasome activation in THP-1 macrophages. It was further found that the pro-inflammatory role of NE is dependent on the activation of protein kinase A (PKA) and subsequently inhibition of β-arrestin2, which is an important regulator of the nuclear factor-kappa B (NF-κB) pathway. This study identifies the neuro-immune axis as an important mediator in obesity-associated cardiac remodeling. Targeting the neuro-immune system may open therapeutic opportunities for the treatment of cardiac remodeling in obesity., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Middle-aged obesity, characterized by excessive fat accumulation and systemic energy imbalance, often precedes various health complications. Recent research has unveiled a surprising link between DNA damage response and energy metabolism. Here, we explore the role of Eepd1, a DNA repair enzyme, in regulating adipose tissue function and obesity onset. Eepd1 is primarily expressed in adipose tissue, where its downregulation or deletion accelerates obesity development. We show that Eepd1 ablation hinders PKA activation, thereby inhibiting lipolysis and thermogenesis in adipose tissue. Notably, cold exposure enhances Eepd1's myristoylation, facilitating its anchorage to adipocyte membranes and subsequent activation of PKA, while a mutation at the myristoylation site of Eepd1 disrupts this process. Moreover, individuals with obesity exhibit reduced Eepd1 expression. Pharmacological restoration of Eepd1 with Retigabine dihydrochloride effectively mitigates obesity. This study reveals Eepd1's unexpected role in promoting adipose lipolysis and thermogenesis, underscoring its potential as a promising therapeutic target to combat obesity., Competing Interests: Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

The energetic demands of proliferating cells during tumorigenesis require close coordination between the cell cycle and metabolism. While CDK4 is known for its role in cell proliferation, its metabolic function in cancer, particularly in triple-negative breast cancer (TNBC), remains unclear. Our study, using genetic and pharmacological approaches, reveals that CDK4 inactivation only modestly impacts TNBC cell proliferation and tumor formation. Notably, CDK4 depletion or long-term CDK4/6 inhibition confers resistance to apoptosis in TNBC cells. Mechanistically, CDK4 enhances mitochondria-endoplasmic reticulum contact (MERCs) formation, promoting mitochondrial fission and ER-mitochondrial calcium signaling, which are crucial for TNBC metabolic flexibility. Phosphoproteomic analysis identified CDK4's role in regulating PKA activity at MERCs. In this work, we highlight CDK4's role in mitochondrial apoptosis inhibition and suggest that targeting MERCs-associated metabolic shifts could enhance TNBC therapy., Competing Interests: Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

A synergistic combination of cationic styrylpyridinium dyes and water-soluble deep cavitand hosts can recognize phosphorylated peptides with both site- and state-selectivity. Two mechanisms of interaction are dominant: either the cationic dye interacts with Trp residues in the peptide or the host:dye pair forms a heteroternary complex with the peptide, driven by both strong dye-peptide and cavitand-peptide binding ( K d values up to 4 μM). The presence of multiple recognition mechanisms results in varying fluorescence responses dependent on the phosphorylation state and position, eliminating the need for covalent modification of the peptide target. Differential sensing aided by machine learning algorithms permits full discrimination between differently positioned serine phosphorylations with a minimal 3-component array. The array is fully functional in the presence of protein kinase A (PKA) and its required cofactors and capable of site-selective monitoring of serine phosphorylation at the privileged PKA motif, in the presence of serine residues that do not undergo reaction, illustrating the potential of the system in kinase-based drug screening.

The pituitary gonadotropin luteinizing hormone (LH) is the primary stimulus for ovulation, luteal formation, and progesterone synthesis, regardless of species. Despite increased awareness of intracellular signaling events initiating the massive production of progesterone during the reproductive cycle and pregnancy, critical gaps exist in our knowledge of the metabolic and lipidomic pathways required for initiating and maintaining luteal progesterone synthesis. Using untargeted metabolomics and metabolic flux analysis in primary steroidogenic luteal cells, evidence is provided for rapid LHCGR-stimulation of metabolic pathways leading to increased glycolysis and oxygen consumption. Treatment with LH stimulated posttranslational modifications of enzymes involved in de novo lipogenesis. Mechanistic studies implicated a crucial role for de novo fatty acid synthesis and fatty acid oxidation in energy homeostasis, LHCGR/PKA signaling, and, consequently, progesterone production. These findings reveal novel hormone-sensitive metabolic pathways essential for maintaining LHCGR/PKA signaling and steroidogenesis. Understanding hormonal control of metabolic pathways in steroidogenic cells may help elucidate approaches for improving ovarian function and successful reproduction or identifying metabolic targets for developing nonhormonal contraceptives., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Published by Elsevier Inc.)

Taste receptor type 1 subunit 3 (T1R3) is a G protein-coupled receptor encoded by the TAS1R3 gene that can be specifically activated by certain sweeteners or umami agents for sweet/umami recognition. T1R3 is a potential target for regulating male reproduction. However, studies on the impact of non-nutritive sweeteners on reproduction are limited. In the present study, we evaluated the impact of the non-nutritive sweeteners (saccharin sodium, sucralose and acesulfame-K) on testosterone synthesis in testicular Leydig cells of Xiang pigs by comparing the relative abundance of mRNA transcripts and protein expression of T1R3, steroidogenic related factors, and intracellular cyclic adenosine monophosphate (cAMP), protein kinase A (PKA), as well as testosterone levels using Western blotting, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA). To clarify the specific mechanism, a dual luciferase assay was used to uncover the relationship between the transcription factors and steroidogenic enzyme. The acute intratesticular injection of a typical non-nutritive sweeteners was conducted to verify this impact in mouse. The results showed that saccharin sodium not only enhanced T1R3 expression in Leydig cells of Xiang pigs, but also caused significant increases in testosterone, cAMP, PKA, phosphorylation of specificity protein 1 (p-SP1), total protein of specificity protein 1 (SP1), steroidogenic acute regulatory protein (StAR), and 3β-hydroxysteroid dehydrogenase type 1 (3β-HSD1) (P < 0.05). Similarly, treatment of Leydig cells with sucralose and acesulfame-K also increased testosterone level, protein expression of T1R3, 17-α-hydroxylase/17, 20-lyase (CYP17A1), and 3β-HSD1 (P < 0.05). Treatment with SQ22536 (an adenylate cyclas inhibitor) or H89 (a PKA inhibitor) significantly reduced saccharin sodium-induced protein levels of p-SP1, StAR, CYP17A1, and 3β-HSD1 (P < 0.05). In addition, a dual luciferase assay further demonstrated that SP1 significantly increased the promoter activity of CYP17A1 (P < 0.05). When mouse testes were injected with saccharin sodium, T1R3, p-SP1, CYP17A1, and 3β-HSD1 were upregulated, leading to a significant testicular increase in testosterone and cAMP levels (P < 0.05). These results suggest a mechanism by which the taste receptor T1R3 regulates testosterone production, and this mechanism may be linked to the cAMP-PKA pathway. Understanding the interrelationship between T1R3 and the cAMP-PKA-SP1 pathway contributes to clarify the regulatory mechanisms of male reproduction., (Copyright © 2024 Elsevier Inc. All rights reserved.)

β-Adrenergic signaling activates cAMP-dependent PKA, which regulates the activity of L-type voltage-gated calcium channels such as Ca V 1.2. Several PKA target sites in the C-terminal tail of Ca V 1.2 have been identified, and their phosphorylation has been suggested to increase currents in specific tissues or heterologous expression systems. However, augmentation of Ca V 1.2 currents in the heart is instead mediated by phosphorylation of Rad, a small GTPase that can inhibit Ca V 1.2. It is unclear how each of the proposed target sites in Ca V 1.2 and Rad rank toward their recognition by PKA, which could reveal a preferential phosphorylation. Here, we used quantitative assays on three Ca V 1.2 and four Rad sites. Isothermal titration calorimetry and enzyme kinetics show that there are two Tiers of targets, with Ca V 1.2 residue Ser1981 and Rad residues Ser25 and Ser272 forming tier one substrates for PKA. These share a common feature with two Arginine residues at specific positions that can anchor the peptide into the substrate binding cleft of PKA. In contrast, PKA shows minimal activity for the other, tier two substrates, characterized by low k cat values and undetectable binding via isothermal titration calorimetry. The existence of two tiers suggests that PKA regulation of the Ca V 1.2 complex may occur in a graded fashion. We report crystal structures of the PKA catalytic subunit with and without a Ca V 1.2 and test the importance of several anchoring residues via mutagenesis. Different target sites utilize different anchors, highlighting the plasticity of PKAc to recognize substrates., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Low phosphorus (LP) diets perturb hepatic energy metabolism homeostasis in fish. However, the specific mechanisms in LP-induced hepatic energy metabolism disorders remain to be fully elucidated., Objectives: This study sought to elucidate the underlying mechanisms of mitochondria involved in LP-induced energy metabolism disorders., Methods: Spotted seabass were fed diets with 0.72% (S-AP, control) or 0.36% (S-LP) available phosphorus for 10 wk. Drp1 was knocked down or protein kinase (PK) A was activated using 8Br-cAMP (5 μM, a PKA activator) in spotted seabass hepatocytes under LP medium. Zebrafish were fed Z-LP diets (0.30% available phosphorus) containing Mdivi-1 (5 mg/kg, a Drp1 inhibitor) or 8Br-cAMP (0.5 mg/kg) for 6 wk. Biochemical and molecular parameters, along with transmission electron microscopy and immunofluorescence, were used to assess hepatic glycolipid metabolism, mitochondrial function, and morphology., Results: Spotted seabass fed S-LP diets showed reduced ATP (52%) and cAMP (52%) concentrations, along with reduced Drp1 (s582) (38%) and PKA (61%) phosphorylation concentrations in the liver compared with those fed S-AP diets (P < 0.05). Drp1 knockdown elevated ATP concentrations (1.99-fold), decreased mitochondrial DRP1 protein amounts (45%), and increased mitochondrial aspect ratio (1.82-fold) in LP-treated hepatocytes (P < 0.05). Furthermore, 8Br-cAMP-treated hepatocytes exhibited higher PKA phosphorylation (2.85-fold), ATP concentrations (1.60-fold), and mitochondrial aspect ratio (2.00-fold), along with decreased mitochondrial DRP1 protein concentrations (29%) under LP medium (P < 0.05). However, mutating s582 to alanine mimic Drp1 dephosphorylation decreased ATP concentrations (63%) and mitochondrial aspect ratio (53%) in 8Br-cAMP-treated hepatocytes (P < 0.05). In addition, zebrafish fed Z-LP diets containing Mdivi-1 or 8Br-cAMP had higher ATP concentrations (3.44-fold or 1.98-fold) than those fed Z-LP diets (P < 0.05)., Conclusions: These findings provide a potential mechanistic elucidation for LP-induced energy metabolism disorders through the cAMP/PKA/Drp1-mediated mitochondrial fission signaling pathway., Competing Interests: Conflicts of interest The authors report no conflict of interest., (Copyright © 2024 American Society for Nutrition. Published by Elsevier Inc. All rights reserved.)

Autism is a complex neurodevelopmental condition characterized by repetitive behaviors, impaired social communication, and various associated conditions such as depression and anxiety. Its multifactorial etiology includes genetic, environmental, dietary, and gastrointestinal contributions. Pathologically, Autism is linked to mitochondrial dysfunction, oxidative stress, neuroinflammation, and neurotransmitter imbalances involving GABA, glutamate, dopamine, and oxytocin. Propionic acid (PRPA) is a short-chain fatty acid produced by gut bacteria, influencing central nervous system functions. Elevated PRPA levels can exacerbate Autism-related symptoms by disrupting metabolic processes and crossing the blood-brain barrier. Our research investigates the neuroprotective potential of Genistein (GNT), an isoflavone compound with known benefits in neuropsychiatric and neurodegenerative disorders, through modulation of the AC/cAMP/CREB/PKA signaling pathway and mitochondrial ETC complex (I-IV) function. In silico analyses revealed GNT's high affinity for these targets. Subsequent in vitro and in vivo experiments using a PRPA-induced rat model of autism demonstrated that GNT (40 and 80 mg/kg., orally) significantly improves locomotion, neuromuscular coordination, and cognitive functions in PRPA-treated rodents. Behavioral assessments showed reduced immobility in the forced swim test, enhanced Morris water maze performance, and restored regular locomotor activity. On a molecular level, GNT restored levels of key signaling molecules (AC, cAMP, CREB, PKA) and mitochondrial complexes (I-V), disrupted by PRPA exposure. Additionally, GNT reduced neuroinflammation and apoptosis, normalized neurotransmitter levels, and improved the complete blood count profile. Histopathological analyses confirmed that GNT ameliorated PRPA-induced brain injuries, restored normal brain morphology, reduced demyelination, and promoted neurogenesis. The study supports GNT's potential in autism treatment by modulating neural pathways, reducing inflammation, and restoring neurotransmitter balance., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Post-stroke depression (PSD) is a significant complication in stroke patients, increases long-term mortality, and exaggerates ischemia-induced brain injury. However, the underlying molecular mechanisms and effective therapeutic targets related to PSD have remained elusive. Here, we employed an animal behavioral model of PSD by combining the use of middle cerebral artery occlusion (MCAO) followed by spatial restraint stress to study the molecular underpinnings and potential therapies of PSD. Interestingly, we found that sub-chronic application of gastrodin (Gas), a traditional Chinese medicinal herb Gastrodia elata extraction, relieved depression-related behavioral deficits, increased the impaired expression of synaptic transmission-associated proteins, and restored the altered spine density in hippocampal CA1 of PSD animals. Furthermore, our results indicated that the anti-PSD effect of Gas was dependent on membrane cannabinoid-1 receptor (CB1R) expression. The contents of phosphorated protein kinase A (p-PKA) and phosphorated Ras homolog gene family member A (p(ser188)-RhoA) were decreased in the hippocampus of PSD-mice, which was reversed by Gas treatment, and CB1R depletion caused a diminished efficacy of Gas on p-PKA and p-RhoA expression. In addition, the anti-PSD effect of Gas was partially blocked by PKA inhibition or RhoA activation, indicating that the anti-PSD effect of Gas is associated with the CB1R-mediated PKA/RhoA signaling pathway. Together, our findings revealed that Gas treatment possesses protective effects against the post-stroke depressive-like state; the CB1R-involved PKA/RhoA signaling pathway is critical in mediating Gas's anti-PSD potency, suggesting that Gas application may be beneficial in the prevention and adjunctive treatment of PSD., Competing Interests: Declarations. Competing Interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Sertoli cells (SCs) are essential for appropriate spermatogenesis. From a metabolic standpoint, they catabolize glucose and provide germ cells with lactate, which is their main energy source. SCs also oxidize fatty acids (FAs), which are stored as triacylglycerides (TAGs) within lipid droplets (LDs), to fulfill their own energy requirements. On the other hand, it has been demonstrated that FSH regulates some of SCs functions, but little is known about its effect on lipid metabolism. In the present study, we aimed to analyze FSH-mediated regulation of (1) lipid storage in LDs and (2) the expression of genes involved in FAs activation and TAG synthesis and storage in SCs. SCs obtained from 20-day-old rats were cultured for different incubation periods with FSH (100 ng/ml). It was observed that FSH increased LD content and TAG levels in SCs. There were also increments in the expression of Plin1, Fabp5, Acsl1, Acsl4, Gpat3, and Dgat1, which suggests that these proteins may mediate the increase in TAGs and LDs elicited by FSH. Regarding the signaling involved in FSH actions, it was observed that dbcAMP increased LD, and H89, a PKA inhibitor, inhibited FSH stimulus. Also, dbcAMP increased Plin2, Fabp5, Acsl1, Acsl4, and Dgat1 mRNA levels, confirming a role of the cAMP/PKA pathway in the regulation of lipid storage in SCs. Altogether, these results suggest that FSH, via the cAMP/PKA pathway, regulates lipid storage in SCs ensuring the availability of substrates to satisfy their energy requirements., Competing Interests: Declaration of competing interest The authors declare that no conflict of interest could be perceived as prejudicing the impartiality of the research reported., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Acute hypoxic exercise will cause insufficient oxygen supply in brain tissue, and a succession of variations such as central dysfunction will occur. For example, the muscles don't have an adequate supply of oxygen, which leads to decrease in exercise capacity (Imray et al., 2005) [1]. The prefrontal cortex in the brain is primarily responsible for regulating the executive functions of the brain. TRPV4 channel is a cation channel with permeability to Ca 2+ . Signals such as hypotonic solution stimulation, cell swelling, temperature stimulation, mechanical stimulation, arachidonic acid and its metabolites can activate TRPV4 channel., Purpose: In conditions of ischemia and hypoxia, the central nervous system of the brain is damaged. Therefore, studying the biological mechanism of TRPV4 pathway can help prevent the damage caused by cerebral ischemia and hypoxia to the human., Results: Studies have found that PKA-mediated phosphorylation at Ser-824 affects AA. This is an important signaling pathway for coronary dilatation. This signaling pathway can activate TRPV4 channels. Therefore, we studied the effect of acute hypoxic exercise on the PKA/AA/TRPV4 pathway in the prefrontal cortex of rats. Furthermore, we concluded that the hypoxic environment can shorten the time of increasing load exercise in rats. In this way, the rat entered a state of exhaustion in advance, and the exercise ability was significantly reduced. After further study, blocking TRPV4 channel can prolong the time of incremental load exercise in hypoxic environment, and the exercise ability is improved. Acute hypoxic exercise led to an increase in the concentration of 14,15-EET, which was speculated to be one of the reasons for the increased expression of TRPV4 channels. Acute hypoxic exercise can activate the PKA/AA/TRPV4 signaling pathway in the prefrontal cortex of rats. Further research on blocking the TRPV4 channel can alleviate the activation of the PKA/AA/TRPV4 signaling pathway in the prefrontal cortex of rats by acute hypoxic exercise., Conclusion: These results suggest that blocking the TRPV4 channel may be one of the ways to reduce the damage and apoptosis of prefrontal cortex cells in rats due to acute hypoxic exercise. In future studies, multiple time points will be selected for collection. Alternatively, TRPV4 agonists, PKA agonists or blockers and 14,15-EET agonists or blockers can be added for further pathway validation. To provide a biological mechanism for the study of nutrient targets on the problem of reduced exercise capacity when military personnel and travel enthusiasts first went to the plateau. Better medical reference for athletes training at high altitudes or patients with respiratory issues., (Copyright © 2024. Published by Elsevier Inc.)

Liver fibrosis is a well-established risk factor for liver cancer development. Despite extensive mechanistic studies on liver fibrosis, the role of the immune cell network in fibrotic disease remains poorly understood. In this study, we demonstrate that regulatory T cells (Tregs) are involved in preventing liver fibrosis by regulating the mevalonate pathway. Blocking the mevalonate pathway increased the granzyme B secretion from Tregs, while restoring the pathway reduced it. Statin treatment, which inhibits the mevalonate pathway, alleviated liver fibrosis progression and enhanced the immunosuppressive function of Tregs in vivo. Mechanistically, mevalonate products, including geranylgeranyl pyrophosphate, inhibited the phosphorylation and activation of LKB1, that is a key regulator of Treg homeostasis. Furthermore, these products disrupted the interaction between LKB1 and cAMP-dependent protein kinase (PKA), leading to further reduction of LKB1 phosphorylation. These findings suggest that targeting LKB1 in Tregs through statin treatment prevents the progression of liver fibrosis, offering a promising and safe therapeutic strategy for liver disease and liver cancer., Competing Interests: Declaration of competing interest All of authors in this study declare no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

CACNA1C encodes the α1c subunit of the L-type Ca 2+ channel, Cav1.2. Ventricular myocytes from haploinsufficient Cacna1c ( Cacna1c +/- ) rats exhibited reduced expression of Cav1.2 but an apparently normal sarcolemmal Ca 2+ influx with an impaired response to sympathetic stress. We tested the hypothesis that the altered phosphorylation of Cav1.2 might underlie the sarcolemmal Ca 2+ influx phenotype in Cacna1c +/- myocytes using immunoblotting of the left ventricular (LV) tissue from Cacna1c +/- versus wildtype (WT) hearts. Activation of cAMP-dependent protein kinase A (PKA) increases L-type Ca 2+ current and phosphorylates Cav1.2 at serine-1928. Using an antibody directed against this phosphorylation site, we observed elevated phosphorylation of Cav1.2 at serine-1928 in LV myocardium from Cacna1c +/- rats under basal conditions (+110% versus WT). Sympathetic stress was simulated by isoprenaline (100 nM) in Langendorff-perfused hearts. Isoprenaline increased the phosphorylation of serine-1928 in Cacna1c +/- LV myocardium by ≈410%, but the increase was significantly smaller than in WT myocardium (≈650%). In conclusion, our study reveals altered PKA-dependent phosphorylation of Cav1.2 with elevated phosphorylation of serine-1928 under basal conditions and a diminished phosphorylation reserve during β-adrenergic stimulation. These alterations in the phosphorylation of Cav1.2 may explain the apparently normal sarcolemmal Ca 2+ influx in Cacna1c +/- myocytes under basal conditions as well as the impaired response to sympathetic stimulation.

Hepatitis E virus (HEV) exists in two distinct forms: a non-enveloped form (neHEV), which is present in feces and bile, and a quasi-enveloped form (eHEV), found in circulating blood and culture supernatants. This study aimed to elucidate the roles of Ras-associated binding 13 (Rab13) and protein kinase A (PKA) in the entry mechanisms of both eHEV and neHEV, utilizing small interfering RNA (siRNA) and chemical inhibitors. The results demonstrated that the entry of both viral forms is dependent on Rab13 and PKA. Further investigation into the involvement of tight junction (TJ) proteins revealed that the targeted knockdown of zonula occludens-1 (ZO-1) significantly impaired the entry of both eHEV and neHEV. In addition, in ZO-1 knockout (KO) cells inoculated with either viral form, HEV RNA levels in culture supernatants did not increase, even up to 16 days post-inoculation. Notably, the absence of ZO-1 did not affect the adsorption efficiency of eHEV or neHEV, nor did it influence HEV RNA replication. In cell-to-cell spread assays, ZO-1 KO cells inoculated with eHEV showed a lack of expression of HEV ORF2 and ORF3 proteins. In contrast, neHEV-infected ZO-1 KO cells showed markedly reduced ORF2 and ORF3 protein expression within virus-infected foci, compared to non-targeting knockout (NC KO) cells. These findings underscore the crucial role of ZO-1 in facilitating eHEV entry and mediating the cell-to-cell spread of neHEV in infected cells.

Background: The heart expresses 2 main subtypes of cAMP-dependent protein kinase (PKA; type I and II) that differ in their regulatory subunits, RIα and RIIα. Embryonic lethality of RIα knockout mice limits the current understanding of type I PKA function in the myocardium. The objective of this study was to test the role of RIα in adult heart contractility and pathological remodeling., Methods: We measured PKA subunit expression in human heart and developed a conditional mouse model with cardiomyocyte-specific knockout of RIα (RIα-icKO). Myocardial structure and function were evaluated by echocardiography, histology, and ECG and in Langendorff-perfused hearts. PKA activity and cAMP levels were determined by immunoassay, and phosphorylation of PKA targets was assessed by Western blot. L-type Ca 2+ current ( I Ca,L ), sarcomere shortening, Ca 2+ transients, Ca 2+ sparks and waves, and subcellular cAMP were recorded in isolated ventricular myocytes (VMs)., Results: RIα protein was decreased by 50% in failing human heart with ischemic cardiomyopathy and by 75% in the ventricles and in VMs from RIα-icKO mice but not in atria or sinoatrial node. Basal PKA activity was increased ≈3-fold in RIα-icKO VMs. In young RIα-icKO mice, left ventricular ejection fraction was increased and the negative inotropic effect of propranolol was prevented, whereas heart rate and the negative chronotropic effect of propranolol were not modified. Phosphorylation of phospholamban, ryanodine receptor, troponin I, and cardiac myosin-binding protein C at PKA sites was increased in propranolol-treated RIα-icKO mice. Hearts from RIα-icKO mice were hypercontractile, associated with increased I Ca,L, and [Ca 2+ ] i transients and sarcomere shortening in VMs. These effects were suppressed by the PKA inhibitor, H89. Global cAMP content was decreased in RIα-icKO hearts, whereas local cAMP at the phospholamban/sarcoplasmic reticulum Ca 2+ ATPase complex was unchanged in RIα-icKO VMs. RIα-icKO VMs had an increased frequency of Ca 2+ sparks and proarrhythmic Ca 2+ waves, and RIα-icKO mice had an increased susceptibility to ventricular tachycardia. On aging, RIα-icKO mice showed progressive contractile dysfunction, cardiac hypertrophy, and fibrosis, culminating in congestive heart failure with reduced ejection fraction that caused 50% mortality at 1 year., Conclusions: These results identify RIα as a key negative regulator of cardiac contractile function, arrhythmia, and pathological remodeling., Competing Interests: Dr Stratakis holds patents on the PRKAR1A, phosphodiesterase 11A, and GPR101 molecules and their function. He is a recipient of funding from Pfizer, Inc for research on growth hormone and growth hormone–producing tumors. Dr Stratakis has working relationships with the ELPEN and H. Lundbeck A/S pharmaceutical companies. The other authors report no conflicts.

The mechanisms of ductus arteriosus (DA) closure involve vasoconstriction and vascular remodeling. Previous findings indicate that the glucagon-like peptide-1 receptor agonist (GLP-1RA) exhibits antihypertensive and anti-remodeling effects in the pulmonary circulation. However, its role in the DA remains unknown. This study aimed to investigate whether exendin-4 (Ex-4), a GLP-1RA, can regulate DA patency and elucidate its mechanisms. After confirming the presence of GLP-1R in neonatal rat DA tissue in vivo, the effects of Ex-4 on DA patency in neonatal rats were sequentially examined. Two hours after birth, we observed spontaneous closure of the DA in control rats. In contrast, Ex-4 prevented the closure of DA, accompanied by reduced intimal thickening. Ex-4 attenuated oxygen-induced vasoconstriction in isolated DA rings ex vivo. This effect was diminished in the presence of H89, a PKA inhibitor. In vitro, Ex-4 inhibited platelet-derived growth factor (PDGF)-BB-induced proliferation and migration of DA smooth muscle cells. Additionally, Ex-4 inhibited PDGF-BB-induced reactive oxygen species (ROS) production, calcium mobilization, and signal transduction of MAPK and Akt pathways. Furthermore, Ex-4 preserved the nuclear expression of Nrf2 attenuated by PDGF-BB. Similarly, all these in vitro effects of Ex-4 were blunted by H89. In conclusion, Ex-4 maintains postnatal DA patency through vasodilatation and anti-remodeling via the PKA pathway. The GLP-1R/PKA pathway emerges as a promising target of DA patency in clinical management., Competing Interests: Declaration of competing interest The authors have declared that no competing interest exists., (Copyright © 2024 Elsevier B.V. All rights reserved.)

GABAergic synapses critically modulate neuronal excitability, and plastic changes in inhibitory synaptic strength require reversible interactions between GABA A receptors (GABA A Rs) and their postsynaptic anchor gephyrin. Inhibitory long-term potentiation (LTP) depends on the postsynaptic recruitment of gephyrin and GABA A Rs, whereas the neurotransmitter GABA can induce synaptic removal of GABA A Rs. However, the mechanisms and players underlying plastic adaptation of synaptic strength are incompletely understood. Here we show that neurobeachin (Nbea), a receptor trafficking protein, is a component of inhibitory synapses, interacts with gephyrin and regulates the downscaling of inhibitory synaptic transmission. We found that the recruitment of Nbea to GABAergic synapses is activity-dependent and that Nbea regulates GABA A R internalization in a protein kinase A (PKA)-dependent manner. In heterozygous neurons lacking one Nbea allele, re-expression of Nbea but not expression of a PKA binding-deficient Nbea mutant rescued the internalization of GABA A Rs. Our data suggest a mechanism by which Nbea mediates PKA anchoring at inhibitory postsynaptic sites to downregulate GABAergic transmission. They emphasize the importance of kinase positioning in the regulation of synaptic strength., Competing Interests: Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Cyclic nucleotide-dependent phosphodiesterases (PDEs) play essential roles in regulating the malaria parasite life cycle, suggesting that they may be promising antimalarial drug targets. PDE inhibitors are used safely to treat a range of noninfectious human disorders. Here, we report three subseries of fast-acting and potent Plasmodium falciparum PDEβ inhibitors that block asexual blood-stage parasite development and that are also active against human clinical isolates. Two of the inhibitor subseries also have potent transmission-blocking activity by targeting PDEs expressed during sexual parasite development. In vitro drug selection experiments generated parasites with moderately reduced susceptibility to the inhibitors. Whole-genome sequencing of these parasites detected no mutations in PDEβ but rather mutations in downstream effectors: either the catalytic or regulatory subunits of cyclic adenosine monophosphate-dependent protein kinase (PKA) or in the 3-phosphoinositide-dependent protein kinase that is required for PKA activation. Several properties of these P. falciparum PDE inhibitor series make them attractive for further progression through the antimalarial drug discovery pipeline.

Ethnopharmacological Relevance: Ischemic stroke is an acute central nervous system disease that poses a threat to human health. It induces a series of severe pathological mechanisms, ultimately leading to neuronal cell death in the brain due to local ischemia and hypoxia. Buyang Huanwu decoction (BYHWD), as a representative formula for treating ischemic stroke, has shown good therapeutic effects in stroke patients., Aim of the Study: This study aimed to explore the mechanism of BYHWD in promoting neural remodeling after ischemic stroke from the perspective of neuronal synaptic plasticity, based on the cAMP/PKA/CREB signaling pathway., Materials and Methods: A modified suture technique was employed to establish a rat model of MCAO. The rats were divided into sham, model, and BYHWD (20 g/kg) groups. After the corresponding intervention, rat brains from each group were collected. TMT quantitative proteomics technology was employed for the research. Following proteomics studies, we investigated the mechanism of BYHWD in the intervention of ischemic stroke through animal experiments and cell experiments. The experimental animals were divided into sham, model, and BYHWD (5 g/kg, 10 g/kg, and 20 g/kg) groups. Infarct volume and severity of brain injury were measured by TTC staining. HE staining was utilized to evaluate alterations in tissue morphology. The Golgi staining was used to observe changes in cell body, dendrites, and dendritic spines. Transmission electron microscopy was used to observe the ultrastructure of synapses in the cortex and hippocampus. TUNEL staining was conducted to identify apoptotic neurons. Meanwhile, a stable and reliable (OGD/R) SH-SY5Y cell model was established. The effect of BYHWD-containing serum on SH-SY5Y cell viability was measured by CCK-8 kit. The apoptosis situation of SH-SY5Y cells was determined by Annexin V-FITC/PI. Immunofluorescence was employed to measure the fluorescence intensity of synaptic-related factors Syt1, Psd95, and Syn1. Synaptic plasticity pathways were assessed by using RT-qPCR and Western blot to determine the expression levels of cAMP, Psd95, Prkacb, Creb1/p-Creb1, BDNF, Shank2, Syn1, Syt1, Bcl-2, Bcl-2/Bax mRNA and proteins., Results: After treatment with BYHWD, notable alterations were detected in the signaling pathways linked to synaptic plasticity and the cAMP signaling pathway-related targets among the intervention targets. This trend of change was also reflected in other bioinformatics analyses, indicating the important role of synaptic plasticity changes before and after modeling and drug intervention. The results of vivo and vitro experiments showed that BYHWD improved local pathological changes, and reduced cerebral infarct volume, and neurological function scores in MCAO rats. It increased dendritic spine density, improved synaptic structural plasticity, and had a certain neuroprotective effect. BYHWD increased the postsynaptic membrane thickness, synaptic interface curvature, and synaptic quantity. 10% BYHWD-containing serum was determined as the optimal concentration for treatment. 10% BYHWD-containing serum significantly reduced the overall apoptotic rate of (OGD/R) SH-SY5Y cells. Immunofluorescence experiments demonstrated that 10% BYHWD-containing serum could improve synaptic plasticity and increase the relative expression levels of synaptic-related proteins Syt1, Psd95, and Syn1. BYHWD and decoction-containing serum upregulated the mRNA and protein expression levels in (OGD/R) SH-SY5Y cells and MCAO rats, suggesting its ability to improve damaged neuronal synaptic plasticity and enhance transmission efficiency, which might be achieved through the regulation of the cAMP/PKA/CREB pathway., Conclusions: This study may provide a basis for clinical medication by elucidating the underlying experimental evidence for the promotion of neural plasticity after ischemic stroke by BYHWD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Synaptic loss and dendritic degeneration are common pathologies in several neurodegenerative diseases characterized by progressive cognitive and/or motor decline, such as Alzheimer's disease (AD) and frontotemporal dementia/amyotrophic lateral sclerosis (FTD/ALS). An essential regulator of neuronal health, the cAMP-dependent transcription factor CREB positively regulates synaptic growth, learning, and memory. Phosphorylation of CREB by protein kinase A (PKA) and other cellular kinases promotes neuronal survival and maturation via transcriptional activation of a wide range of downstream target genes. CREB pathway dysfunction has been strongly implicated in AD pathogenesis, and recent data suggest that impaired CREB activation may contribute to disease phenotypes in FTD/ALS as well. However, the mechanisms behind reduced CREB activity in FTD/ALS pathology are not clear. In this study, we found that cortical-like neurons derived from iPSC lines carrying the hexanucleotide repeat expansion in the C9ORF72 gene, a common genetic cause of FTD/ALS, displayed a diminished activation of CREB, resulting in decreased dendritic and synaptic health. Importantly, we determined such impairments to be mechanistically linked to an imbalance in the ratio of regulatory and catalytic subunits of the CREB activator PKA and to be conserved in C9-ALS patient's postmortem tissue. Modulation of cAMP upstream of this impairment allowed for a rescue of CREB activity and an amelioration of dendritic morphology and synaptic protein levels. Our data elucidate the mechanism behind early CREB pathway dysfunction and discern a feasible therapeutic target for the treatment of FTD/ALS and possibly other neurodegenerative diseases., Competing Interests: Competing interests statement:The authors declare no competing interest.

Background: Tong-Qiao-Huo-Xue Decoction (TQHXD) is a traditional Chinese medicinal formula widely used in the treatment of vascular dementia (VD). Although it has demonstrated good clinical efficacy, the specific molecular mechanisms underlying its therapeutic effects on VD remain unclear., Objective: This study aimed to elucidate the neuroprotective mechanisms of TQHXD to provide a scientific basis for the clinical treatment of VD., Methods: The chemical components of TQHXD were qualitatively analyzed using ultra-performance liquid chromatography (UPLC) and gas chromatography (GC). Network pharmacology predicted the potential protective mechanisms of TQHXD against VD. A rat model of VD was established through bilateral vessel occlusion (2-VO), and an oxygen-glucose deprivation/reperfusion (OGD/R) model was used to induce damage to neuronal cells of the hippocampus. In vivo experiments assessed changes in cerebral blood flow, learning and memory capabilities, hippocampal neuronal morphology, dendritic length, dendritic spine density, and synapse number in rats. We examined the expression of synaptic remodeling-related proteins and pathway proteins in the hippocampal region. In vitro assays evaluated cell viability, apoptosis, reactive oxygen species (ROS) levels, and expression of synaptic remodeling-related proteins and signaling pathway., Results: Multiple active components were identified in TQHXD. KEGG enrichment analysis suggested that the therapeutic effects of TQHXD on VD may be related to the cAMP signaling pathway. Treatment with TQHXD significantly improved learning and memory performance in VD rats, improved hippocampus morphology, and increased dendritic length, dendritic spine density, and number of synapses. Furthermore, TQHXD improved cell viability, reduced apoptosis, and decreased intracellular ROS levels in vitro. Western blotting, immunofluorescence, and enzyme-linked immunosorbent assay results collectively demonstrated that TQHXD upregulated the expression of synaptic remodeling-related proteins and pathway-related proteins both in vivo and in vitro., Conclusions: TQHXD treated VD by promoting synaptic remodeling in hippocampal neurons, likely through activation of the cAMP/PKA/CREB pathway., Competing Interests: Declaration of competing interest All authors of this research paper have directly participated in the planning, execution, or analysis of the study. The contents of this manuscript are not under consideration for publication elsewhere, and will not be copyrighted, submitted, or published elsewhere. There are no directly related manuscripts or abstracts, published or unpublished, by any author(s) of this paper. The authors also claim that there is no competitive financial interest., (Copyright © 2024 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Background: Connexin 43 (Cx43) plays a crucial role in mediating intracellular communication and facitating the interaction between exosomes and recipient cells. This study investigates whether the activation of cAMP/protein kinase A (PKA) can regulate exosomal Cx43 expression and contribute to the functional recovery following ischemia-reperfusion (I/R) injury., Methods: An intraluminal vascular occlusion was performed on Lewis rats to simulate I/R injury. Concurrently, a PKA activator (8-Bromo-cAMP, 5 mg kg -1 ) or PKA inhibitor (H 89 2HCl, 20 mg kg -1 ) was administered intravenously via the tail vein (n = 10). Exosomes were isolated from cerebrospinal fluid, and the expression of exosomal markers (CD63 and CD81) and Cx43 was analyzed using Western blot. The expression of CD63 and CD81 in astrocytes was measured to assess exosome uptake. Spatial learning and memory capability were evaluated using the Morris water maze test., Results: 8-Bromo-cAMP significantly increased exosome release in cerebrospinal fluid, accompanied by elevated Cx43 expression. Additionally, 8-Bromo-cAMP enhanced exosome uptake by astrocytes, alleviated blood-brain barrier damage and edema, and improved cognitive function., Conclsions: PKA activation enhances exosome production, promotes cognitive function recovery, and attenuates cerebral I/R injury by up-regulating exosomal Cx43 expression., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The fields of allostery and amyloid-related pathologies, such as Parkinson's disease (PD), have been extensively explored individually, but less is known about how amyloids control allostery. Recent advancements have revealed that amyloids can drive allosteric effects in both intrinsically disordered proteins, such as alpha-synuclein (αS), and multi-domain signaling proteins, such as protein kinase A (PKA). Amyloid-driven allostery plays a central role in explaining the mechanisms of gain-of-pathological-function mutations in αS (e.g. E46K, which causes early PD onset) and loss-of-physiological-function mutations in PKA (e.g. A211D, which predisposes to tumors). This review highlights allosteric effects of disease-related mutations and how they can cause exposure of amyloidogenic regions, leading to amyloids that are either toxic or cause aberrant signaling. We also discuss multiple potential modulators of these allosteric effects, such as MgATP and kinase substrates, opening future opportunities to improve current pharmacological interventions against αS and PKA-related pathologies. Overall, we show that amyloid-driven allosteric models are useful to explain the mechanisms underlying disease-related mutations., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Brain astrocyte glycogenolysis is regulated in part by the second messenger adenosine 3'5'-cyclic monophosphate (cAMP). Hypothalamic astrocyte glycogen metabolism shapes glucose counterregulation, under the control of glucose transporter-2 (GLUT2), a plasma membrane glucose carrier and sensor. Hypothalamic astrocyte cAMP is subject to neurotransmitter control, but effects of nutrient cues on this messenger are unclear. Here, an established hypothalamic primary astrocyte culture model and gene knockdown tools were used to investigate the premise that GLUT2 exerts sex-dimorphic regulation of cAMP-protein kinase A (PKA) signalling in these glia. Data show that basal cAMP was elevated in female versus male; GLUT2 gene silencing up-regulated or down-regulated this profile in male versus female. Glucoprivation increased cAMP content in astrocytes of each sex, yet GLUT2 siRNA pretreatment exacerbated (male) or blunted (female) this stimulatory effect. PKA and phosphoPKA levels in glucose-supplied astrocytes were increased (male) or decreased (female) by GLUT2 knockdown. PKA protein was amplified, yet phosphoPKA was refractory to glucose withdrawal in male, while females showed sustained PKA expression alongside diminished phosphoPKA. GLUT2 siRNA pretreatment exacerbated glucoprivic augmentation of PKA content in male but down-regulated both PKA and phosphoPKA proteins in female. Evidence for parallel GLUT2 siRNA-associated changes in cAMP and PKA, albeit in opposing directions in the two sexes, infers that GLUT2 control of hypothalamic astrocyte cAMP-dependent PKA signalling is sex-specific. Data also disclose that in the female, GLUT2 curbs the baseline phosphoPKA/PKA expression ratio but is not involved in glucoprivic suppression of this ratio., (© 2024 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Adrenergic receptors (ARs) are preferentially expressed by innate lymphocytes such as natural killer (NK) cells. Here, we study the effect of epinephrine-mediated stimulation of the β2-adrenergic receptor (β2AR) on the function of human NK cells. Epinephrine stimulation inhibited early NK cell signaling events and blocked the function of the integrin LFA-1. This reduced the adhesion of NK cells to ICAM-1, explaining how NK cells are mobilized into the peripheral blood upon epinephrine release during acute stress or exercise. Additionally, epinephrine stimulation transiently reduced NK cell degranulation, serial killing, and cytokine production and affected metabolic changes upon NK cell activation via the cAMP-protein kinase A (PKA) pathway. Repeated exposure to β2AR agonists resulted in the desensitization of the β2AR via a PKA feedback loop-initiated G-protein switch. Therefore, acute epinephrine stimulation of chronically β2AR stimulated NK cells no longer resulted in inhibited signaling and reduced LFA-1 activity. Sustained stimulation by long-acting β2-agonists (LABA) not only inhibited NK cell functions but also resulted in desensitization of the β2AR. However, peripheral NK cells from LABA-treated asthma patients still reacted unchanged to epinephrine stimulation, demonstrating that local LABA administration does not result in detectable systemic effects on NK cells., (© 2024 The Author(s). European Journal of Immunology published by Wiley‐VCH GmbH.)

Leydig cells are the main testosterone-producing cells in males. During androgen synthesis, cholesterol enters the mitochondria via the STAR protein and is converted into pregnenolone by the CYP11A1 enzyme. This steroid is then exported from the mitochondria to be metabolized to progesterone by the HSD3B1 enzyme in the endoplasmic reticulum. In this study, we used 3'Tag-RNA-Seq to identify progesterone-regulated genes in MA-10 Leydig cells. Our results indicate that high concentrations of progesterone (30 μM) are involved in a negative feedback loop that inhibits cAMP/PKA-dependent activation of Star and Cyp11a1 expression and participate in cAMP/PKA-dependent down-regulation of genes related to the metabolism of steroid hormones. Linked to activation of the MAPK signaling pathway, endoplasmic reticulum stress and apoptosis, most of the genes encoding bZIP transcription factors are upregulated by progesterone in MA-10 Leydig cells. However, only DDIT3 protein levels are increased in response to progesterone in MA-10 Leydig cells. Like normal Leydig cells, MA-10 cells very weakly express the classical nuclear receptor for progesterone, suggesting that gene regulation by progesterone is rather mediated by one of the non-classical membrane receptors for progesterone However, current findings suggest that the inhibitory effect of progesterone on STAR protein increase in response to forskolin is not dependent on PGRMC1/2 or PAQR9. Furthermore, the increase in progesterone synthesis in response to activation of the cAMP/PKA pathway is rather inhibited by siRNA-mediated knockdown of PAQR9. Overall, this study shows that progesterone produced by Leydig cells participates in the regulation of steroidogenesis through autocrine action involving negative feedback upon activation of the cAMP/PKA pathway., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: The polysaccharide (PSP) from purple sweet potato has great potential for regulating apoptosis, but its regulatory role in acute lung injury (ALI) is unknown., Methods: The objective of this study was to investigate the protective effect of PSP on lipopolysaccharide (LPS)-induced ALI in mice and lung epithelial A549 cells and its mechanism. Moreover, subacute toxicity evaluation of PSP was carried out on ICR mice., Results: The results showed that compared with the ALI group, PSP significantly reduced the total protein content, wet-to-dry (W/D) ratio, the number of neutrophils, lymphocytes, and monocytes. Moreover, PSP was able to reduce cell apoptosis, the levels of macrophage inflammatory protein-2 (MIP-2), intercellular adhesion molecule-1 (ICAM-1), tumor necrosis factor-α (TNF-α), malondialdehyde (MDA) and myeloperoxidase (MPO) and increase the level of superoxide (SOD). In addition, PSP could up-regulate the levels of VIP, cAMP, p-PKA/PKA and AQP1 in mice and A549 cells. And PSP exhibited no apparent adverse effects on the mice., Conclusions: PSP had a protective effect on LPS-induced ALI in mice and lung epithelial A549 cells, which may be related to the inflammatory response and via VIP/cAMP/PKA/AQPs signaling pathway. Thus, PSP may be a promising pharmacologic agent for ALI therapy., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

The phosphorylation reaction, catalyzed by the enzyme protein kinase A (PKA), plays one of the key roles in the work of the glutamatergic system, primarily involved in memory functioning. The analysis of the dynamic behavior of the enzyme-substrate complex allows one to learn the mechanism of the enzymatic reaction. According to the results of classical molecular dynamics calculations followed by hierarchical clustering, the most preferred proton acceptor during the phosphorylation reaction catalyzed by PKA is the carboxyl group of the amino acid residue Asp166; however, the γ-phosphate group of ATP can also act as an acceptor.

The secondary metabolites of several plant species, particularly sesquiterpenic lactones (SLs) have been studied by different research groups for over 30 years. This group of metabolites presents numerous biological activities such as antibacterial, antiviral, antiulcer, cell proliferation inhibitor, and oocyte activator with participation in exocytosis processes. This study aims to assess some sperm parameters in epididymal gametes of Chichilla lanigera exposed to increasing concentrations (0 to 2 mM) of DhL for various incubation times from 10 to 40 minutes. We determined the participation of different cell signalling pathways in the induced acrosome reaction. Our results showed an alteration in the progressive motility pattern and cell viability depending on DhL concentration and exposure time of gametes. When analyzing acrosomal status, higher percentages than the negative control were obtained in all tested doses. Both isolated and joint inhibition tests of PKA and phospholipases (PLC and PLA 2 ) showed a greater participation of PI-PLC. This is the first report concerning the effects of this lactone on the medium of sperm incubation. Consequently, further studies will be necessary to determine the molecular implications of this lactone on the fertilizing potential of the sperm.

The phosphorylation of synaptic proteins is a significant biochemical reaction that controls the sleep-wake cycle in mammals 1-3 . Protein phosphorylation in vivo is reversibly regulated by kinases and phosphatases. In this study, we investigate a pair of kinases and phosphatases that reciprocally regulate sleep duration. First, we perform a comprehensive screen of protein kinase A (PKA) and phosphoprotein phosphatase (PPP) family genes by generating 40 gene knockout mouse lines using prenatal and postnatal CRISPR targeting. We identify a regulatory subunit of PKA (Prkar2b), a regulatory subunit of protein phosphatase 1 (PP1; Pppr1r9b) and catalytic and regulatory subunits of calcineurin (also known as PP2B) (Ppp3ca and Ppp3r1) as sleep control genes. Using adeno-associated virus (AAV)-mediated stimulation of PKA and PP1-calcineurin activities, we show that PKA is a wake-promoting kinase, whereas PP1 and calcineurin function as sleep-promoting phosphatases. The importance of these phosphatases in sleep regulation is supported by the marked changes in sleep duration associated with their increased and decreased activities, ranging from approximately 17.3 h per day (PP1 expression) to 4.3 h per day (postnatal CRISPR targeting of calcineurin). Localization signals to the excitatory post-synapse are necessary for these phosphatases to exert their sleep-promoting effects. Furthermore, the wake-promoting effect of PKA localized to the excitatory post-synapse negated the sleep-promoting effect of PP1-calcineurin. These findings indicate that PKA and PP1-calcineurin have competing functions in sleep regulation at excitatory post-synapses., Competing Interests: Competing interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Autophagy is a crucial intracellular degradation process that provides energy and supports nutrient deprivation adaptation. However, the mechanisms by which these cells detect lipid scarcity and regulate autophagy are poorly understood. In this study, we demonstrate that protein kinase A (PKA)-dependent lipolysis delays autophagy initiation during short-term nutrient deprivation by inhibiting AMP-activated protein kinase (AMPK). Using coherent anti-Stokes Raman spectroscopy, we visualized free fatty acids (FFAs) in vivo and observed that lipolysis-derived FFAs were used before the onset of autophagy. Our data suggest that autophagy is triggered when the supply of FFAs is insufficient to meet energy demands. Furthermore, PKA activation promotes lipolysis and suppresses AMPK-driven autophagy during early fasting. Disruption of this regulatory axis impairs motility and reduces the lifespan of Caenorhabditis elegans during fasting. These findings establish PKA as a critical regulator of catabolic pathways, prioritizing lipolysis over autophagy by modulating AMPK activity to prevent premature autophagic degradation during transient nutrient deprivation., Competing Interests: Declaration of Competing Interests The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Alzheimer's disease (AD) is a degenerative brain disorder characterized by progressive cognitive decline and neuronal death due to extracellular deposition of amyloid β (Aβ) and intracellular deposition of tau proteins. Recently approved antibody drugs targeting Aβ have been shown to slow the progression of the disease, but they have minimal effects on cognitive improvement. Therefore, there is a need to develop drugs with cognitive-enhancing effects that can be used in conjunction with these antibody treatments. In this study, we investigated whether Polygala tenuifolia (PT), traditionally known for its cognitive-enhancing effects, can improve synaptic plasticity and identified its active components and mechanisms. PT demonstrated a dose-dependent effect in enhancing long-term potentiation (LTP), and among its components, 3,4,5-trimethoxycinnamic acid methyl ester (TMCA) showed a similar LTP-enhancing effect. TMCA increased the phosphorylation of the GluA1 subunit of the α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors and increased the amount of GluA1 on the synapse without affecting the amount of GluA2. Additionally, the increase in GluA1 induced by TMCA was inhibited by a PKA inhibitor. Consistent with these results, the enhancement of LTP by TMCA was inhibited by a GluA1 antagonist and a PKA inhibitor. In silico molecular docking experiments confirmed that TMCA binds to PKA. Finally, we confirmed the LTP-enhancing effect of TMCA in hippocampal slices from 5XFAD mice. These results suggest that PT and its active component, TMCA, can interact with PKA to enhance LTP, indicating the potential for improving cognitive function in AD patients., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

cAMP-PKA signaling initiates the crucial process of oocyte meiotic maturation in many animals, but inhibits it in vertebrates. To address this 'cAMP paradox', we exchanged the key PKA substrate ARPP19 between representative species, the vertebrate Xenopus and the cnidarian Clytia, comparing its phosphorylation and function. We found that, as in Xenopus, Clytia maturing oocytes undergo ARPP19 phosphorylation on a highly conserved Gwl site, which inhibits PP2A and promotes M-phase entry. In contrast, despite a PKA phosphorylation signature motif recognizable across most animals, Clytia ARPP19 was only poorly phosphorylated by PKA in vitro and in vivo. Furthermore, unlike Xenopus ARPP19, exogenous Clytia ARPP19 did not delay Xenopus oocyte maturation. We conclude that, in Clytia, ARPP19 does not intervene in oocyte maturation initiation because of both poor recognition by PKA and the absence of effectors that mediate vertebrate oocyte prophase arrest. We propose that ancestral ARPP19 phosphorylated by Gwl has retained a key role in M-phase across eukaryotes and has acquired new functions during animal evolution mediated by enhanced PKA phosphorylation, allowing co-option into oocyte maturation regulation in the vertebrate lineage., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

Objective: To determine the effect of electroacupuncture (EA) on β 1 -adrenergic receptor (β 1 -AR) and post-receptor protein kinase A (PKA) signaling pathway after acute myocardial ischemia (MI)., Methods: An MI model was established by ligating the left anterior descending coronary artery of wild-type (WT) C57/BL and β 1 -AR +/- mice (heterozygous for β 1 -AR gene deletion). EA treatment was administered at HT5-HT7 or LU9-LU8. We evaluated cardiac function by measuring ST segment displacement, ischemic area and serum levels of creatine kinase (CK)-MB and lactate dehydrogenase (LDH). Pathological morphology/apoptosis of myocardial tissue were examined using hematoxylin-eosin and terminal deoxynucleotidyl transferase dUTP nick end labeling staining. Norepinephrine (NE) levels in myocardial tissue were detected by ELISA. Levels of β 1 and post-receptor PKA signaling components were evaluated by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and Western blotting., Results: EA stimulation at HT7-HT5 could better regulate the level of β 1 -AR in myocardial tissue than that at LU9-LU8. Following EA, the ST segment, serum CK-MB/ LDH and area of myocardial infarction were decreased in WT mice, and the degree of myocardial pathology/apoptosis and expression of cleaved caspase-3 were decreased. Myocardial levels of Gs protein (Gs), adenylate cyclase (AC), cyclic adenosine monophosphate (cAMP), phosphorylated protein kinase A (p-PKA), L-type voltage-gated calcium channel α1C (Cav1.2), serine phosphate 16-phospholamban (p-PLB s16 ) and sarcoplasmic reticulum Ca 2+ -adenosine triphosphate (ATP)ase 2a (SERCA2a) increased after EA. However, these effects of EA were not replicated in β 1 -AR +/- mice. Interestingly, myocardial NE content decreased after EA in WT and β 1 -AR +/- mice., Conclusion: EA may enhance cardiac function and reduced MI area/apoptosis by restoring the activity of β 1 -AR and post-receptor PKA signaling., Competing Interests: Declaration of conflicting interestsThe authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Objective: The study aimed to investigate the effects of glucagon on metabolic pathways in mouse models of obesity, fatty liver disease, and type 2 diabetes (T2D) to determine the extent and variability of hepatic glucagon resistance in these conditions., Methods: We investigated glucagon's effects in mouse models of fatty liver disease, obesity, and type 2 diabetes (T2D), including male BKS-db/db, high-fat diet-fed, and western diet-fed C57Bl/6 mice. Glucagon tolerance tests were performed using the selective glucagon receptor agonist acyl-glucagon (IUB288). Blood glucose, serum and liver metabolites include lipids and amino acids were measured. Additionally, liver protein expression related to glucagon signalling and a comprehensive liver metabolomics were performed., Results: Western diet-fed mice displayed impaired glucagon response, with reduced blood glucose and PKA activation. In contrast, high-fat diet-fed and db/db mice maintained normal glucagon sensitivity, showing significant elevations in blood glucose and phospho-PKA motif protein expression. Acyl-glucagon treatment also lowered liver alanine and histidine levels in high-fat diet-fed mice, but not in western diet-fed mice. Additionally, some amino acids, such as methionine, were increased by acyl-glucagon only in chow diet control mice. Despite normal glucagon sensitivity in PKA signalling, db/db mice had a distinct metabolomic response, with acyl-glucagon significantly altering 90 metabolites in db/+ mice but only 42 in db/db mice, and classic glucagon-regulated metabolites, such as cyclic adenosine monophosphate (cAMP), being less responsive in db/db mice., Conclusions: The study reveals that hepatic glucagon resistance in obesity and T2D is complex and not uniform across metabolic pathways, underscoring the complexity of glucagon action in these conditions., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Adam Rose reports financial support was provided by Diabetes Australia. Adam Rose reports financial support was provided by National Health and Medical Research Council. Yuqin Wu reports financial support was provided by Australian Physiological Society. Adam Rose reports a relationship with Boehringer Ingelheim Pharma GmbH & Co KG that includes: funding grants. Patricia Rusu reports a relationship with Boehringer Ingelheim Pharma GmbH & Co KG that includes: funding grants. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Background: The β-adrenergic augmentation of cardiac contraction, by increasing the conductivity of L-type voltage-gated Ca V 1.2 channels, is of great physiological and pathophysiological importance. Stimulation of β-adrenergic receptors (βAR) activates protein kinase A (PKA) through separation of regulatory (PKAR) from catalytic (PKAC) subunits. Free PKAC phosphorylates the inhibitory protein Rad, leading to increased Ca 2+ influx. In cardiomyocytes, the core subunit of Ca V 1.2, Ca V 1.2α 1 , exists in two forms: full-length or truncated (lacking the distal C-terminus (dCT)). Signaling efficiency is believed to emanate from protein interactions within multimolecular complexes, such as anchoring PKA (via PKAR) to Ca V 1.2α 1 by A-kinase anchoring proteins (AKAPs). However, AKAPs are inessential for βAR regulation of Ca V 1.2 in heterologous models, and their role in cardiomyocytes also remains unclear., Results: We show that PKAC interacts with Ca V 1.2α 1 in heart and a heterologous model, independently of Rad, PKAR, or AKAPs. Studies with peptide array assays and purified recombinant proteins demonstrate direct binding of PKAC to two domains in Ca V 1.2α 1 -CT: the proximal and distal C-terminal regulatory domains (PCRD and DCRD), which also interact with each other. Data indicate both partial competition and possible simultaneous interaction of PCRD and DCRD with PKAC. The βAR regulation of Ca V 1.2α 1 lacking dCT (which harbors DCRD) was preserved, but subtly altered, in a heterologous model, the Xenopus oocyte., Conclusions: We discover direct interactions between PKAC and two domains in Ca V 1.2α 1 . We propose that these tripartite interactions, if present in vivo, may participate in organizing the multimolecular signaling complex and fine-tuning the βAR effect in cardiomyocytes., Competing Interests: Declarations. Ethics approval and consent to participate: Experiments were approved by Tel Aviv University Institutional Animal Care and Use Committee (IACUC permit # 01–20-083). Consent for publication: Not applicable. Competing interests: The authors declare that they have no competing interests., (© 2024. The Author(s).)

Chronic stress disrupts dopamine (DA) transmission, adversely affecting mood and contribution to neuropsychiatric disorders like ADHD, autism, schizophrenia, anxiety, depression, and drug addiction. The neuropeptide oxytocin (OXT) plays a key role in social cognition, bonding, attachment, and parenting behaviors. In addition, OXT can modulate the activity of the HPA axis, counteracting the effects of stress, and alleviating fear and anxiety. However, whether OXT can mitigate stress-induced DA dysfunction and the underlying mechanisms remains unclear. This study investigated the neuroprotective effects of OXT on corticosterone (CORT) induced DA dysfunction in the neuroblastoma cell line SH-SY5Y. The results revealed that CORT decreases the levels of intracellular signaling molecules associated with DA function, including phosphorylated tyrosine hydroxylase (pTH), phosphorylated cAMP response element-binding protein (pCREB), and protein kinase A (PKA). Interestingly, pretreatment with OXT mitigated CORT-induced DA dysfunction through its potent PKA activator properties. In addition, the neuroprotective effect of OXT was abolished by atosiban (an OXT receptor antagonist) or H89 (a PKA inhibitor). Our results suggest that OXT protects dopaminergic neuroblastoma cells from CORT-induced DA dysfunction, potentially through the involvement of oxytocin receptors and the PKA/CREB signaling pathway. These findings contribute to the understanding of the neurobiological mechanisms underlying stress resilience and highlight potential pathways for developing targeted treatments that leverage the neuroprotective properties of OXT to address disorders characterized by DA dysregulation and impaired stress responses., Competing Interests: Declarations. Competing Interests: The authors declare no competing interests., (© 2024. The Author(s).)

Purpose: Ulcerative colitis (UC) is a chronic, non-specific inflammatory condition of the colon, characterized by recurrent episodes and a notable lack of effective pharmacological treatments. Scutellarin, a natural component, exhibits appreciable pharmacological effects and therapeutic potential for various diseases. However, its effects on UC are not fully understood, and the precise mechanisms remain to be deciphered. This study aimed to assess the therapeutic efficacy of scutellarin and elucidate its underlying mechanisms in treating UC., Methods: This study utilized dextran sulfate sodium (DSS)-induced mice to evaluate the therapeutic potential of scutellarin against UC and to elucidate the mechanisms involving the gut microbiota. An antibiotics cocktail (ABX) and fecal microbiota transplantation (FMT) were also used to determine the mechanistic role of the gut microbiota. An integrative approach combining fecal metabolomics and network pharmacology analysis was used to explore the gut microbiota-directed molecular mechanism., Results: The results showed that scutellarin provided various therapeutic benefits in UC management, including alleviating weight loss, slowing disease progression, and reducing inflammatory damage in colon structures. The improved gut microbiota after scutellarin administration contributed to these effects. Fecal metabolome revealed that scutellarin selectively mitigated DSS-induced dysregulation of gut microbiota-derived metabolites, including glycolic acid, γ-aminobutyric acid, glutamate, tryptophan, xanthine, and β-hydroxypyruvate. Network pharmacology analysis, along with in vivo experimental verification, implicated the cAMP/PKA/NF-κB pathway in the action of these metabolites in treating UC, which may be the mechanism responsible for scutellarin's curative effects on UC., Conclusion: This study demonstrates the potential of scutellarin in alleviating UC by activating the cAMP/PKA/NF-κB pathway through gut microbiota-derived metabolites, highlighting scutellarin as a promising therapeutic agent for UC., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Genetic abnormalities of the parathyroid hormone 1 receptor (PTH1R) lead to profound craniomaxillofacial bone and dentition defects on account of inappropriate tissue metabolism and cellular differentiation. The coordinated activity of differentiation and viability in bone cells is indispensable for bone metabolism. Recent research demonstrates mesenchymal progenitors are responsive to PTH1R signaling for osteogenic differentiation, whereas the effect of PTH1R on cellular survival remains incompletely understood. Here, we report that mice with deletion of PTH1R in Prx1-positive mesenchymal cells ( Prx1Cre;PTH1R fl/fl ) exhibit decreased alveolar bone mass due in part to apoptotic response activation. The exploration of oral bone-derived mesenchymal stem cells (OMSCs) with PTH1R deficiency suggests PTH1R signaling modulates OMSCs' apoptosis by interfering mitochondrial function and morphology. The underlying molecular mechanisms are studied by transcriptome sequencing analysis, finding that inositol trisphosphate receptor-3 (IP3R-3), an endoplasmic reticulum calcium channel protein, serves as a modulator of pro-apoptosis in OMSCs. Furthermore, we find PTH1R and its downstream protein kinase A (PKA) pathway dampen IP3R-3's expression. Of note, OMSCs with IP3R-3 overexpression recapitulate the PTH1R-deletion phenotypes, while IP3R-3 silence rescues mitochondrial dysfunction. Altogether, our study uncovers the anti-apoptotic function of PTH1R signaling in OMSCs and proves that excess apoptosis partly contributes to a weakening potential of osteogenic differentiation and aberrant mandibular development.