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Idiopathic pulmonary fibrosis (IPF) is a fatal lung disease with no ideal drugs. Our previous research demonstrated that phosphodiesterase 1 (PDE1) could be a promising target for the treatment of IPF. However, only a few selective PDE1 inhibitors are available, and the mechanism of recognition between inhibitors and the PDE1 protein is not fully understood. This study carried out a step-by-step optimization of a dihydropyrimidine hit Z94555858 . By targeting the metal pocket of PDE1, a lead compound 3f was obtained, exhibiting an IC 50 value of 11 nM against PDE1, moderate selectivity over other PDEs, and significant anti-fibrotic effects in bleomycin-induced pulmonary fibrosis rats. The structure-activity relationship study aided by molecular docking revealed that forming halogen bonds with water in the metal pocket greatly enhanced the PDE1 inhibition, providing a novel strategy for further rational design of PDE1 inhibitors.

Long-term synaptic plasticity at glutamatergic synapses on striatal spiny projection neurons (SPNs) is central to learning goal-directed behaviors and habits. Our studies reveal that SPNs manifest a heterosynaptic, nitric oxide (NO)-dependent form of long-term postsynaptic depression of glutamatergic SPN synapses (NO-LTD) that is preferentially engaged at quiescent synapses. Plasticity is gated by Ca 2+ entry through Ca V 1.3 Ca 2+ channels and phosphodiesterase 1 (PDE1) activation, which blunts intracellular cyclic guanosine monophosphate (cGMP) and NO signaling. Both experimental and simulation studies suggest that this Ca 2+ -dependent regulation of PDE1 activity allows for local regulation of dendritic cGMP signaling. In a mouse model of Parkinson disease (PD), NO-LTD is absent because of impaired interneuronal NO release; re-balancing intrastriatal neuromodulatory signaling restores NO release and NO-LTD. Taken together, these studies provide important insights into the mechanisms governing NO-LTD in SPNs and its role in psychomotor disorders such as PD., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

PDE1B had been found to be involved in various diseases, including tumors and non-tumors. However, little was known about the definite role of PDE1B in osteosarcoma. Therefore, we mined public data on osteosarcoma to reveal the prognostic values and immunological roles of the PDE1B gene. Three osteosarcoma-related datasets from online websites were utilized for further data analysis. R 4.3.2 software was utilized to conduct difference analysis, prognostic analysis, gene set enrichment analysis (GSEA), nomogram construction, and immunological evaluations, respectively. Experimental verification of the PDE1B gene in osteosarcoma was conducted by qRT-PCR and western blot, based on the manufacturer's instructions. The PDE1B gene was discovered to be lowly expressed in osteosarcoma, and its low expression was associated with poor OS (all P < 0.05). Experimental verifications by qRT-PCR and western blot results remained consistent (all P < 0.05). Univariate and multivariate Cox regression analyses indicated that the PDE1B gene had independent abilities in predicting OS in the TARGET osteosarcoma dataset (both P < 0.05). GSEA revealed that PDE1B was markedly linked to the calcium, cell cycle, chemokine, JAK STAT, and VEGF pathways. Moreover, PDE1B was found to be markedly associated with immunity (all P < 0.05), and the TIDE algorithm further shed light on that patients with high-PDE1B expression would have a better immune response to immunotherapies than those with low-PDE1B expression, suggesting that the PDE1B gene could prevent immune escape from osteosarcoma. The PDE1B gene was found to be a tumor suppressor gene in osteosarcoma, and its high expression was related to a better OS prognosis, suppressing immune escape from osteosarcoma., (© 2024. The Author(s).)

Cocaine use disorder is a significant public health issue without an effective pharmacological treatment. Successful treatments are hindered in part by an incomplete understanding of the molecular mechanisms that underlie long-lasting maladaptive plasticity and addiction-like behaviors. Here, we leverage a large RNA sequencing dataset to generate gene coexpression networks across six interconnected regions of the brain's reward circuitry from mice that underwent saline or cocaine self-administration. We identify phosphodiesterase 1b ( Pde1b ), a Ca 2+ /calmodulin-dependent enzyme that increases cAMP and cGMP hydrolysis, as a central hub gene within a nucleus accumbens (NAc) gene module that was bioinformatically associated with addiction-like behavior. Chronic cocaine exposure increases Pde1b expression in NAc D2 medium spiny neurons (MSNs) in male but not female mice. Viral-mediated Pde1b overexpression in NAc reduces cocaine self-administration in female rats but increases seeking in both sexes. In female mice, overexpressing Pde1b in D1 MSNs attenuates the locomotor response to cocaine, with the opposite effect in D2 MSNs. Overexpressing Pde1b in D1/D2 MSNs had no effect on the locomotor response to cocaine in male mice. At the electrophysiological level, Pde1b overexpression reduces sEPSC frequency in D1 MSNs and regulates the excitability of NAc MSNs. Lastly, Pde1b overexpression significantly reduced the number of differentially expressed genes (DEGs) in NAc following chronic cocaine, with discordant effects on gene transcription between sexes. Together, we identify novel gene modules across the brain's reward circuitry associated with addiction-like behavior and explore the role of Pde1b in regulating the molecular, cellular, and behavioral responses to cocaine., Competing Interests: The authors declare no competing financial interests., (Copyright © 2024 the authors.)

Liver fibrosis is a common pathological feature of most chronic liver diseases with no effective drugs available. Phosphodiesterase 1 (PDE1), a subfamily of the PDE super enzyme, might work as a potent target for liver fibrosis by regulating the concentration of cAMP and cGMP. However, there are few PDE1 selective inhibitors, and none has been investigated for liver fibrosis treatment yet. Herein, compound AG-205/1186117 with the dihydropyrimidine scaffold was selected as the hit by virtual screening. A hit-to-lead structural modification led to a series of dihydropyrimidine derivatives. Lead 13h exhibited the IC 50 of 10 nM against PDE1, high selectivity over other PDEs, as well as good safety properties. Administration of 13h exerted significant anti-liver fibrotic effects in bile duct ligation-induced fibrosis rats, which also prevented TGF-β-induced myofibroblast differentiation in vitro, confirming that PDE1 could work as a potential target for liver fibrosis.

Phosphodiesterase 1 (PDE1) is an enzyme entrusted with the hydrolysis of the second messengers cAMP and cGMP, thereby governing a plethora of metabolic processes, encompassing ion channel modulation and cellular apoptosis. Recent advancements in the realm of small molecule structural variations have greatly facilitated the exploration of innovative applications for PDE1. Remarkably, a recent series of PDE1 inhibitors (PDE1i) have been meticulously formulated and devised, showcasing enhanced selectivity and potency. Among them, ITI-214 has entered Phase II clinical trials, holding promise for the treatment of Parkinson's disease and heart failure. Nevertheless, the majority of current PDE1 inhibitors have encountered substantial side effects in clinical trials attributable to their limited selectivity, this predicament presents a formidable obstacle in the development of specific small molecule inhibitors targeting PDE1. This Perspective endeavors to illuminate the potential design approaches, structure-activity relationships, and biological activities of current PDE1i, aiming to offer support and insights for clinical practice and the development of novel PDE1i., 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 © 2023 Elsevier Masson SAS. All rights reserved.)

Aim: Synthesis of novel bis -Schiff bases having potent inhibitory activity against phosphodiesterase (PDE-1 and -3) enzymes, potentially offering therapeutic implications for various conditions. Methods: Bis -Schiff bases were synthesized by refluxing 2,4-dihydroxyacetophenone with hydrazine hydrate, followed by treatment of substituted aldehydes with the resulting hydrazone to obtain the product compounds. After structural confirmation, the compounds were screened for their in vitro PDE-1 and -3 inhibitory activities. Results: The prepared compounds exhibited noteworthy inhibitory efficacy against PDE-1 and -3 enzymes by comparing with suramin standard. To clarify the binding interactions between the drugs, PDE-1 and -3 active sites, molecular docking studies were carried out. Conclusion: The potent compounds discovered in this study may be good candidates for drug development.

Although the association of elevated homocysteine level with cardiac hypertrophy has been reported, the molecular mechanisms by which homocysteine induces cardiac hypertrophy remain inadequately understood. In this study we aim to uncover the roles of cyclic nucleotide phosphodiesterase 1 (PDE1) and endoplasmic reticulum (ER) stress and their relationship to advance the mechanistic understanding of homocysteine-induced cardiac cell hypertrophy. H9c2 cells and primary neonatal rat cardiomyocytes are exposed to homocysteine with or without ER stress inhibitor TUDCA or PDE1-specific inhibitor Lu AF58027, or transfected with siRNAs targeting PDE1 isoforms prior to homocysteine-exposure. Cell surface area is measured and ultrastructure is examined by transmission electron microscopy. Hypertrophic markers, PDE1 isoforms, and ER stress molecules are detected by q-PCR and western blot analysis. Intracellular cGMP and cAMP are measured by ELISA. The results show that homocysteine causes the enlargement of H9c2 cells, increases the expressions of hypertrophic markers β-MHC and ANP, upregulates PDE1A and PDE1C, promotes the expressions of ER stress molecules, and causes ER dilatation and degranulation. TUDCA and Lu AF58027 downregulate β-MHC and ANP, and alleviate cell enlargement. TUDCA decreases PDE1A and PDE1C levels. Silencing of PDE1C inhibits homocysteine-induced hypertrophy, whereas PDE1A knockdown has minor effect. Both cAMP and cGMP are decreased after homocysteine-exposure, while only cAMP is restored by Lu AF58027 and TUDCA. TUDCA and Lu AF58027 also inhibit cell enlargement, downregulate ANP, β-MHC and PDE1C, and enhance cAMP level in homocysteine-exposed primary cardiomyocytes. ER stress mediates homocysteine-induced hypertrophy of cardiac cells via upregulating PDE1C expression Cyclic nucleotide, especially cAMP, is the downstream mediator of the ER stress-PDE1C signaling axis in homocysteine-induced cell hypertrophy.

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Phosphodiesterase-1 (PDE1) is a promising drug target closely related to central and peripheral diseases. With the assistance of molecular docking and dynamics simulations, we designed and synthesized a novel series of pyrazolopyrimidone derivatives as effective and metabolically stable inhibitors against PDE1. Most compounds have good inhibitory activities against PDE1 at the concentration of 20 nM. Compound 2j with the IC 50 of 21 nM against PDE1B, shows good metabolic stability in the rat liver microsomes (RLM) (t 1/2 of 28.5 min), indicating that compound 2j can be used as a tool to explore the molecular recognition mechanism between inhibitors and the target protein PDE1., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Abdominal aortic aneurysm (AAA) is characterized by aorta dilation due to wall degeneration, which mostly occurs in elderly males. Vascular aging is implicated in degenerative vascular pathologies, including AAA. Cyclic nucleotide phosphodiesterases, by hydrolyzing cyclic nucleotides, play critical roles in regulating vascular structure remodeling and function. Cyclic nucleotide phosphodiesterase 1C (PDE1C) expression is induced in dedifferentiated and aging vascular smooth muscle cells (SMCs), while little is known about the role of PDE1C in aneurysm. We observed that PDE1C was not expressed in normal aorta but highly induced in SMC-like cells in human and murine AAA. In mouse AAA models induced by Angiotensin II or periaortic elastase, PDE1C deficiency significantly decreased AAA incidence, aortic dilation, and elastin degradation, which supported a causative role of PDE1C in AAA development in vivo. Pharmacological inhibition of PDE1C also significantly suppressed preestablished AAA. We showed that PDE1C depletion antagonized SMC senescence in vitro and/or in vivo, as assessed by multiple senescence biomarkers, including senescence-associated β-galactosidase activity, γ-H2AX foci number, and p21 protein level. Interestingly, the role of PDE1C in SMC senescence in vitro and in vivo was dependent on Sirtuin 1 (SIRT1). Mechanistic studies further showed that cAMP derived from PDE1C inhibition stimulated SIRT1 activation, likely through a direct interaction between cAMP and SIRT1, which leads to subsequent up-regulation of SIRT1 expression. Our findings provide evidence that PDE1C elevation links SMC senescence to AAA development in both experimental animal models and human AAA, suggesting therapeutical significance of PDE1C as a potential target against aortic aneurysms., Competing Interests: The authors declare no competing interest.

Streptococcus mitis is a commensal bacterial species of the oral cavity, with the potential for opportunistic pathogenesis. For successful colonization, S. mitis must be able to adhere to surfaces of the oral cavity and survive and adapt to frequently changing environmental conditions. Cyclic-di-AMP (c-di-AMP) is a nucleotide second messenger, involved in the regulation of stress responses and biofilm formation in several bacterial species. Cyclic-di-AMP is produced by diadenylate cyclases and degraded by phosphodiesterases. We have previously shown that in S. mitis, one diadenylate cyclase (CdaA) and at least two phosphodiesterases (Pde1 and Pde2) regulate the intracellular concentration of c-di-AMP. In this study, we utilized S. mitis deletion mutants of cdaA, pde1, and pde2 to analyze the role of c-di-AMP signaling in various stress responses, biofilm formation, and adhesion to eukaryotic cells. Here, we demonstrate that the Δpde1 mutant displayed a tendency toward increased susceptibility to acetic acid at pH 4.0. Deletion of cdaA increases auto-aggregation of S. mitis but reduces biofilm formation on an abiotic surface. These phenotypes are more pronounced under acidic extracellular conditions. Inactivation of pde1 or pde2 reduced the tolerance to ciprofloxacin, and UV radiation and the Δpde1 mutant was more susceptible to Triton X-100, indicating a role for c-di-AMP signaling in responses to DNA damage and cell membrane perturbation. Finally, the Δpde2 mutant displayed a tendency toward a reduced ability to adhere to oral keratinocytes. Taken together, our results indicate an important role for c-di-AMP signaling in cellular processes important for colonization of the mouth., (© 2021 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.)

Diminished nitric oxide-cGMP-mediated relaxation plays a crucial role in cardiovascular aging, leading to decreased vasodilation, vascular hypertrophy and stiffening, and ultimately, cardiovascular dysfunction. Aging is the time-related worsening of physiologic function due to complex cellular and molecular interactions, and it is at least partly driven by DNA damage. Genetic deletion of the DNA repair enzyme ERCC1 endonuclease in Ercc1 Δ/- mice provides us an efficient tool to accelerate vascular aging, explore mechanisms, and test potential treatments. Previously, we identified the cGMP-degrading enzyme phosphodiesterase 1 as a potential treatment target in vascular aging. In the present study, we studied the effect of acute and chronic treatment with ITI-214, a selective phosphodiesterase 1 inhibitor on vascular aging features in Ercc1 Δ/- mice. Compared with wild-type mice, Ercc1 Δ/- mice at the age of 14 weeks showed decreased reactive hyperemia, diminished endothelium-dependent and -independent responses of arteries in organ baths, carotid wall hypertrophy, and elevated circulating levels of inflammatory cytokines. Acute ITI-214 treatment in organ baths restored the arterial endothelium-independent vasodilation in Ercc1 Δ/- mice. An 8-week treatment with 100 mg/kg per day ITI-214 improved endothelium-independent relaxation in both aorta and coronary arteries, at least partly restored the diminished reactive hyperemia, lowered the systolic and diastolic blood pressure, normalized the carotid hypertrophy, and ameliorated inflammatory responses exclusively in Ercc1 Δ/- mice. These findings suggest phosphodiesterase 1 inhibition would provide a powerful tool for nitric oxide-cGMP augmentation and have significant therapeutic potential to battle arteriopathy related to aging. SIGNIFICANCE STATEMENT: The findings implicate the key role of phosphodiesterase 1 in vascular function and might be of clinical importance for the prevention of mortalities and morbidities related to vascular complications during aging, as well as for patients with progeria that show a high risk of cardiovascular disease., (Copyright © 2021 by The American Society for Pharmacology and Experimental Therapeutics.)

Positive modulation of cAMP signalling by phosphodiesterase (PDE) inhibitors has recently been explored as a potential target for the reversal of cognitive and behavioural deficits implicating the corticoaccumbal circuit. Previous studies show that PDE type 1 isoform B (PDE1B) inhibition may improve memory function in rodent models; however, the contribution of PDE1B inhibition to impulsivity, attentional and motivational functions as well as its neurophysiological effects have not been investigated. To address this, we recorded single unit activity in medial prefrontal cortex (mPFC) and nucleus accumbens (NAc) in Lister Hooded rats treated with the PDE1B inhibitor Lu AF64386 and tested in the 5-choice serial reaction time task (5-CSRTT). We also asked whether PDE1B inhibition modulates neurophysiological deficits produced by subchronic phencyclidine (PCP) treatment, a rat pharmacological model of schizophrenia. Lu AF64386 significantly affected behavioural parameters consistent with a reduction in goal-directed behaviour, however without affecting accuracy. Additionally, it reduced mPFC neuronal activity. Pre-treatment with PCP did not affect behavioural parameters, however it significantly disrupted overall neuronal firing while increasing phasic responses to reward-predicting cues and disrupting mPFC-NAc cross-talk. The latter two effects were reversed by Lu AF64386. These findings suggest PDE1B inhibition may be beneficial in disorders implicating a dysfunction of the mPFC-NAc network., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

In addition to maintaining cellular ER Ca 2+ stores, store-operated Ca 2+ entry (SOCE) regulates several Ca 2+ -sensitive cellular enzymes, including certain adenylyl cyclases (ADCYs), enzymes that synthesize the secondary messenger cyclic AMP (cAMP). Ca 2+ , acting with calmodulin, can also increase the activity of PDE1-family phosphodiesterases (PDEs), which cleave the phosphodiester bond of cAMP. Surprisingly, SOCE-regulated cAMP signaling has not been studied in cells expressing both Ca 2+ -sensitive enzymes. Here, we report that depletion of ER Ca 2+ activates PDE1C in human arterial smooth muscle cells (HASMCs). Inhibiting the activation of PDE1C reduced the magnitude of both SOCE and subsequent Ca 2+ /calmodulin-mediated activation of ADCY8 in these cells. Because inhibiting or silencing Ca 2+ -insensitive PDEs had no such effects, these data identify PDE1C-mediated hydrolysis of cAMP as a novel and important link between SOCE and its activation of ADCY8. Functionally, we showed that PDE1C regulated the formation of leading-edge protrusions in HASMCs, a critical early event in cell migration. Indeed, we found that PDE1C populated the tips of newly forming leading-edge protrusions in polarized HASMCs, and co-localized with ADCY8, the Ca 2+ release activated Ca 2+ channel subunit, Orai1, the cAMP-effector, protein kinase A, and an A-kinase anchoring protein, AKAP79. Because this polarization could allow PDE1C to control cAMP signaling in a hyper-localized manner, we suggest that PDE1C-selective therapeutic agents could offer increased spatial specificity in HASMCs over agents that regulate cAMP globally in cells. Similarly, such agents could also prove useful in regulating crosstalk between Ca 2+ /cAMP signaling in other cells in which dysregulated migration contributes to human pathology, including certain cancers., Competing Interests: Conflict of interest None declared., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Hypoxic coronary vasospasm may lead to myocardial ischaemia and cardiac dysfunction. Inosine 3',5'-cyclic monophosphate (cIMP) is a putative second messenger to mediate this pathological process. Nevertheless, it remains unclear as to whether levels of cIMP can be regulated in living tissue such as coronary artery and if so, what is the consequence of this regulation on hypoxia-induced vasoconstriction. In the present study, we found that cIMP was a key determinant of hypoxia-induced constriction but not that of the subsequent relaxation response in porcine coronary arteries. Subsequently, coronary arteries were treated with various phosphodiesterase (PDE) inhibitors to identify PDE types that are capable of regulating cIMP levels. We found that inhibition of PDE1 and PDE5 substantially elevated cIMP content in endothelium-denuded coronary artery supplemented with exogenous purified cIMP. However, cGMP levels were far lower than their levels in intact coronary arteries and lower than cIMP levels measured in endothelium-denuded coronary arteries supplemented with exogenous cIMP. The increased cIMP levels induced by PDE1 or PDE5 inhibition further led to augmented hypoxic constriction without apparently affecting the relaxation response. In intact coronary artery, PDE1 or PDE5 inhibition up-regulated cIMP levels under hypoxic condition. Concomitantly, cGMP level increased to a comparable level. Nevertheless, the hypoxia-mediated constriction was enhanced in this situation that was largely compromised by an even stronger inhibition of PDEs. Taken together, these data suggest that cIMP levels in coronary arteries are regulated by PDE1 and PDE5, whose inhibition at a certain level leads to increased cIMP content and enhanced hypoxic constriction., (© 2020 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

N 6 -Methyladenosine (m 6 A) is the most abundant modification of mammalian mRNAs. RNA methylation fine tunes RNA stability and translation, altering cell fate. The fat mass- and obesity-associated protein (FTO) is an m 6 A demethylase with oncogenic properties in leukemia. Here, we show that FTO expression is suppressed in ovarian tumors and cancer stem cells (CSC). FTO inhibited the self-renewal of ovarian CSC and suppressed tumorigenesis in vivo , both of which required FTO demethylase activity. Integrative RNA sequencing and m 6 A mapping analysis revealed significant transcriptomic changes associated with FTO overexpression and m 6 A loss involving stem cell signaling, RNA transcription, and mRNA splicing pathways. By reducing m 6 A levels at the 3'UTR and the mRNA stability of two phosphodiesterase genes ( PDE1C and PDE4B ), FTO augmented second messenger 3', 5'-cyclic adenosine monophosphate (cAMP) signaling and suppressed stemness features of ovarian cancer cells. Our results reveal a previously unappreciated tumor suppressor function of FTO in ovarian CSC mediated through inhibition of cAMP signaling. SIGNIFICANCE: A new tumor suppressor function of the RNA demethylase FTO implicates m 6 A RNA modifications in the regulation of cyclic AMP signaling involved in stemness and tumor initiation., (©2020 American Association for Cancer Research.)

Among ageing-related illnesses, cardiovascular disease (CVD) remains the leading cause of morbidity and mortality causing one-third of all deaths worldwide. Ageing evokes a number of functional, pharmacological and morphological changes in the vasculature, accompanied by a progressive failure of protective and homeostatic mechanisms, resulting in target organ damage. Impaired vasomotor, proliferation, migration, antithrombotic and anti-inflammatory function in both the endothelial and vascular smooth muscle cells are parts of the vascular ageing phenotype. The endothelium regulates these functions by the release of a wide variety of active molecules including endothelium-derived relaxing factors such as nitric oxide, prostacyclin (PGI 2 ) and endothelium-derived hyperpolarization (EDH). During ageing, a functional decay of the nitric oxide pathway takes place. Nitric oxide signals to VSMC and other important cell types for vascular homeostasis through the second messenger cyclic guanosine monophosphate (cGMP). Maintenance of proper cGMP levels is an important goal in sustainment of proper vascular function during ageing. For this purpose, different components can be targeted in this signalling system, and among them, phosphodiesterase-1 (PDE1) and soluble guanylate cyclase (sGC) are crucial. This review focuses on the role of PDE1 and sGC in conditions that are relevant for vascular ageing., (© 2019 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).)

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, and devastating lung disease lacking effective therapy. To identify whether phosphodiesterase-1 (PDE1) inhibition could act as a novel target for the treatment of IPF, hit-to-lead structural optimizations were performed on the PDE9/PDE1 dual inhibitor ( R )- C33 , leading to compound 3m with an IC 50 of 2.9 nM against PDE1C, excellent selectivity across PDE subfamilies, reasonable drug-like properties, and remarkable pharmacodynamic effects as an anti-IPF agent. Oral administration of compound 3m (10 mg/kg) exerted more significant anti-pulmonary fibrosis effects than pirfenidone (150 mg/kg) in a bleomycin-induced IPF rat model and prevented transforming growth factor-β-induced fibroblast-to-myofibroblast conversion in vitro, indicating that PDE1 inhibition could serve as a novel target for the efficient treatment of IPF.

A new series of Vinpocetine derivatives were synthesized and evaluated for their inhibitory activity on PDE1A in vitro. Seven compounds with higher inhibitory activity were selected for surface plasmon resonance (SPR) binding experiments. Compared with Vinpocetine, these high potency compounds presented a higher binding affinity with PDE1A, which was consistent with inhibitory activity. After further screening, compounds 5, 7, 21, 34 and Vinpocetine were selected to examine the vasorelaxant effects on endothelium-intact rat thoracic aortic rings. The study suggested that the effects of compounds 7 and 21 were the most significant with the maximum value of 93.46 ± 0.77% and 92.90 ± 0.78% (n = 5) at a concentration of 100 μM respectively. Based on these studies, compounds 7 and 21 were considered for further development as hit compounds., (Copyright © 2019. Published by Elsevier Ltd.)

A novel, potent, and highly specific inhibitor of calcium-calmodulin-dependent phosphodiesterases (PDE) of the PDE1 family, ITI-214, was used to investigate the role of PDE1 in inflammatory responses. ITI-214 dose-dependently suppressed lipopolysaccharide (LPS)-induced gene expression of pro-inflammatory cytokines in an immortalized murine microglial cell line, BV2 cells. RNA profiling (RNA-Seq) was used to analyze the impact of ITI-214 on the BV2 cell transcriptome in the absence and the presence of LPS. ITI-214 was found to regulate classes of genes that are involved in inflammation and cell migration responses to LPS exposure. The gene expression changes seen with ITI-214 treatment were distinct from those elicited by inhibitors of other PDEs with anti-inflammatory activity (e.g., a PDE4 inhibitor), indicating a distinct mechanism of action for PDE1. Functionally, ITI-214 inhibited ADP-induced migration of BV2 cells through a P2Y12-receptor-dependent pathway, possibly due to increases in the extent of cAMP and VASP phosphorylation downstream of receptor activation. Importantly, this effect was recapitulated in P2 rat microglial cells in vitro, indicating that these pathways are active in native microglial cells. These studies are the first to demonstrate that inhibition of PDE1 exerts anti-inflammatory effects through effects on microglia signaling pathways. The ability of PDE1 inhibitors to prevent or dampen excessive inflammatory responses of BV2 cells and microglia provides a basis for exploring their therapeutic utility in the treatment of neurodegenerative diseases associated with increased inflammation and microglia proliferation such as Parkinson's disease and Alzheimer's disease., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

The calcium-regulated phosphodiesterase 1 (PDE1) family is highly expressed in the brain, but its functional role in neurones is poorly understood. Using the selective PDE1 inhibitor Lu AF64196 and biosensors for cyclic nucleotides including a novel biosensor for cGMP, we analyzed the effect of PDE1 on cAMP and cGMP in individual neurones in brain slices from male newborn mice. Release of caged NMDA triggered a transient increase of intracellular calcium, which was associated with a decrease in cAMP and cGMP in medium spiny neurones in the striatum. Lu AF64196 alone did not increase neuronal cyclic nucleotide levels, but blocked the NMDA-induced reduction in cyclic nucleotides indicating that this was mediated by calcium-activated PDE1. Similar effects were observed in the prefrontal cortex and the hippocampus. Upon corelease of dopamine and NMDA, PDE1 was shown to down-regulate the D1-receptor mediated increase in cAMP. PDE1 inhibition increased long-term potentiation in rat ventral striatum, showing that PDE1 is implicated in the regulation of synaptic plasticity. Overall, our results show that PDE1 reduces cyclic nucleotide signaling in the context of glutamate and dopamine coincidence. This effect could have a therapeutic value for treating brain disorders related to dysfunctions in dopamine neuromodulation., (© The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Pharmacological activation of protein kinase A (PKA) reduces migration of arterial smooth muscle cells (ASMCs), including those isolated from human arteries (HASMCs). However, when individual migration-associated cellular events, including the polarization of cells in the direction of movement or rearrangements of the actin cytoskeleton, are studied in isolation, these individual events can be either promoted or inhibited in response to PKA activation. While pharmacological inhibition or deficiency of exchange protein activated by cAMP-1 (EPAC1) reduces the overall migration of ASMCs, the impact of EPAC1 inhibition or deficiency, or of its activation, on individual migration-related events has not been investigated. Herein, we report that EPAC1 facilitates the formation of leading-edge protrusions (LEPs) in HASMCs, a critical early event in the cell polarization that underpins their migration. Thus, RNAi-mediated silencing, or the selective pharmacological inhibition, of EPAC1 decreased the formation of LEPs by these cells. Furthermore, we show that the ability of EPAC1 to promote LEP formation by migrating HASMCs is regulated by a phosphodiesterase 1C (PDE1C)-regulated "pool" of intracellular HASMC cAMP but not by those regulated by the more abundant PDE3 or PDE4 activities. Overall, our data are consistent with a role for EPAC1 in regulating the formation of LEPs by polarized HASMCs and show that PDE1C-mediated cAMP hydrolysis controls this localized event.

Phosphodiesterase (PDE) inhibition has been broadly investigated as a target for a wide variety of indications including central nervous system (CNS) disorders. Cyclic nucleotide (cNT) changes within associated tissues may serve as a biomarker of PDE inhibition. We recently developed robust sample harvesting and bioanalytical methods to quantify cNT levels in rodent brain and cerebrospinal fluid (CSF). Herein, we report on the application of those methods to study rodent species-specific and rodent brain region-specific cNT changes following individual or concomitant PDE inhibitor administration. Male Sprague Dawley (Crl:CD ® [SD]) rats were dosed subcutaneously (sc) with a PDE1B inhibitor (DNS-0056), a PDE2A inhibitor (PF-05180999), a PDE9A inhibitor (PF-4447943), and a PDE10A inhibitor (MP10), each at a single dose of 10 or 30 mg/kg, or concomitantly with all 4 inhibitors at 10 mg/kg each. Male Carworth Farms (Crl:CF1 ® [CF-1]) mice were dosed intraperitoneally (ip) with the four individual inhibitors at a single dose of 10 mg/kg or concomitantly with all 4 inhibitors at 10 mg/kg each. The doses studied are generally adequate for affecting measurable cNT levels in the tissues of interest and were thereby chosen for this investigation. Measured 3',5'-cyclic adenosine monophosphate (cAMP) changes were generally statistically insignificant in the brain, striatum and CSF after administration of the aforementioned PDE inhibitors. However, the levels of 3',5'-cyclic guanosine monophosphate (cGMP) increased in both rat and mouse striatum (2.2-, 2.1- and 1.7-fold and 6.4-, 2.8- and 1.7-fold, respectively) after PDE2A, 9A, and 10A inhibitor dosing. In all cases, the cNT changes followed the same trend in the brain, striatum and CSF after PDE inhibitor dosing and dose response was observed in rats. Concomitant treatment with PDE1B, PDE2A, PDE9A and PDE10A inhibitors resulted in a 4.4- and 36.7-fold increase of cGMP in rat and mouse striatum. The drug exposures after concomitant treatment were also higher than in the individual inhibitor-treated animals. cGMP enhancement observed could be due to synergistic effects, though an additive effect of the combined inhibitor concentrations may also contribute., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Although UVA radiation (315-400 nm) represents 95% of the UV radiation reaching the earth's surface, surprisingly little is known about its effects on plants [1]. We show that in Arabidopsis, short-term exposure to UVA inhibits the opening of stomata, and this requires a reduction in the cytosolic level of cGMP. This process is independent of UVR8, the UVB receptor. A cGMP-activated phosphodiesterase (AtCN-PDE1) was responsible for the UVA-induced decrease in cGMP in Arabidopsis. AtCN-PDE1-like proteins form a clade within the large HD-domain/PDEase-like protein superfamily, but no eukaryotic members of this subfamily have been functionally characterized. These genes have been lost from the genomes of metazoans but are otherwise conserved as single-copy genes across the tree of life. In longer-term experiments, UVA radiation increased growth and decreased water-use efficiency. These experiments revealed that PDE1 is also a negative regulator of growth. As the PDE1 gene is ancient and not represented in animal lineages, it is likely that at least one element of cGMP signaling in plants has evolved differently to the system present in metazoans., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

No current treatment targets cardiac proteotoxicity or can reduce mortality of heart failure (HF) with preserved ejection fraction (HFpEF). Selective degradation of misfolded proteins by the ubiquitin-proteasome system (UPS) is vital to the cell. Proteasome impairment contributes to HF. Activation of cAMP-dependent protein kinase (PKA) or cGMP-dependent protein kinase (PKG) facilitates proteasome functioning. Phosphodiesterase 1 (PDE1) hydrolyzes both cyclic nucleotides and accounts for most PDE activities in human myocardium. We report that PDE1 inhibition (IC86430) increases myocardial 26 S proteasome activities and UPS proteolytic function in mice. Mice with CryAB R120G -based proteinopathy develop HFpEF and show increased myocardial PDE1A expression. PDE1 inhibition markedly attenuates HFpEF, improves mouse survival, increases PKA-mediated proteasome phosphorylation, and reduces myocardial misfolded CryAB. Therefore, PDE1 inhibition induces PKA- and PKG-mediated promotion of proteasomal degradation of misfolded proteins and treats HFpEF caused by CryAB R120G , representing a potentially new therapeutic strategy for HFpEF and heart disease with increased proteotoxic stress.

Background: Phosphodiestrase (PDE) enzymes are suggested to play a leading role in fibrogenesis of liver where studies showed the possible implication of PDE 1 & 4 in liver injury proposing them as possible targets for treating liver fibrosis., Aim: The present study was designed to investigate, for the first time, the possible therapeutic effects of selective inhibitors of PDE-1 (vinpocetine) and PDE-4 (roflumilast) in liver fibrosis induced by diethylnitrosamine (DEN) in rats., Main Methods: Rats were given DEN (100 mg/kg, i.p.) once weekly for 6 weeks to induce liver fibrosis. Vinpocetine (10 mg/kg/day) or roflumilast (0.5 mg/kg/day) was then orally administered for 2 weeks., Key Findings: Vinpocetine significantly suppressed the contents of hydroxyproline, transforming growth factor-beta 1 (TGF-β1), nuclear factor-kappa B (NF-κB) whereas roflumilast normalized them. Moreover, tumor necrosis factor-alpha (TNF-α) content and protein expressions of toll-like receptor 4 (TLR4) and tissue inhibitor of metalloproteinase-1 (TIMP-1) were markedly decreased whereas cAMP response element binding (CREB) protein expression was significantly elevated by both treatments. Additionally, vinpocetine and roflumilast up-regulated the gene expression of bone morphogenetic protein and activin membrane-bound inhibitor (BAMBI) receptor where roflumilast showed better results. PDE1 and 4 activities were inhibited by vinpocetine and roflumilast, respectively. The superior results offered by roflumilast could be related to the higher cAMP level obtained relative to vinpocetine., Significance: Our study manifested the up-regulation of PDE enzymes (1 & 4) in liver fibrosis and addressed the therapeutic role of vinpocetine and roflumilast as PDEIs through a cAMP-mediated TLR4 inflammatory and fibrogenic signaling pathways., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The aim of this work was to determine bioavailability and in vivo calmodulin-dependent-phosphodiesterase (CaMPDE)-inhibitory activity of six flaxseed-protein-derived peptides (AGA, AKLMS, QIAK, RWIQ, QQAKQ, and KQLSTGC) after oral administration to Wistar rats. Initial experiments tested the cytotoxicity and cellular-transport potentials of the peptides using Caco-2 cells. The cytotoxicity assay indicated that none of the six peptides had an adverse effect on the proliferation and viability of the Caco-2 cells, whereas the transport assay confirmed peptide translocation across the cell membrane. However, only two of the peptides (AGA and RWIQ) were detected in the rat serum up to 90 min postgavage, with traces of RWIQ persisting in serum 1 week after oral gavage. The six peptides inhibited plasma activity of CaMPDE with AGA (34.63%), QIAK (36.66%), and KQLSTGC (34.21%) being the most effective 30 min after gavage. In contrast, only AGA maintained significant plasma-CaMPDE-activity inhibition (44.35%) after 60 min.

Vinpocetine, a phosphodiesterase (PDE) type-1 inhibitor, increases cAMP and cGMP levels and is currently used for the management of cerebrovascular disorders, such as stroke, cerebral hemorrhage, and cognitive dysfunctions. In this study, we first determined that vinpocetine effectively suppressed adipogenesis and lipid accumulation. However, we questioned which molecular mechanism is involved because the role of PDE in adipogenesis is still controversial. Vinpocetine decreased adipogenic cell signaling, including the phosphorylation of ERK, AKT, JAK2, and STAT3, and adipokine secretion, including IL-6, IL-10, and IFN-α. Interestingly, vinpocetine increased the phosphorylation of HSL, suggesting the induction of the lipolysis pathway. Moreover, vinpocetine increased UCP1 expression via increasing cAMP and PKA phosphorylation. The administration of vinpocetine with a normal-chow diet (NFD) or a high-fat diet (HFD) in mice attenuated body weight gain in mice fed both the NFD and HFD. These effects were larger in the HFD-fed mice, without a difference in food intake. Vinpocetine drastically decreased fat weight and adipocyte cell sizes in gonadal and inguinal white adipose tissues and in the liver in both diet groups. Serum triacylglycerol levels and fasting blood glucose levels were reduced by vinpocetine treatment. This study suggested that vinpocetine prevents adipocyte differentiation through the inhibition of adipogenesis-associated cell signaling in the early stages of adipogenesis. Moreover, upregulating cAMP levels leads to an increase in lipolysis and UCP1 expression and then inhibits lipid accumulation. Therefore, we suggest that vinpocetine could be an effective agent for treating obesity, as well as improving cognition and cardiovascular function in older individuals.

Oligodendrocyte gene expression is downregulated in stress-related neuropsychiatric disorders, including depression. In mice, chronic social stress (CSS) leads to depression-relevant changes in brain and emotional behavior, and the present study shows the involvement of oligodendrocytes in this model. In C57BL/6 (BL/6) mice, RNA-sequencing (RNA-Seq) was conducted with prefrontal cortex, amygdala and hippocampus from CSS and controls; a gene enrichment database for neurons, astrocytes and oligodendrocytes was used to identify cell origin of deregulated genes, and cell deconvolution was applied. To assess the potential causal contribution of reduced oligodendrocyte gene expression to CSS effects, mice heterozygous for the oligodendrocyte gene cyclic nucleotide phosphodiesterase (Cnp1) on a BL/6 background were studied; a 2 genotype (wildtype, Cnp1 +/- ) × 2 environment (control, CSS) design was used to investigate effects on emotional behavior and amygdala microglia. In BL/6 mice, in prefrontal cortex and amygdala tissue comprising gray and white matter, CSS downregulated expression of multiple oligodendroycte genes encoding myelin and myelin-axon-integrity proteins, and cell deconvolution identified a lower proportion of oligodendrocytes in amygdala. Quantification of oligodendrocyte proteins in amygdala gray matter did not yield evidence for reduced translation, suggesting that CSS impacts primarily on white matter oligodendrocytes or the myelin transcriptome. In Cnp1 mice, social interaction was reduced by CSS in Cnp1 +/- mice specifically; using ionized calcium-binding adaptor molecule 1 (IBA1) expression, microglia activity was increased additively by Cnp1 +/- and CSS in amygdala gray and white matter. This study provides back-translational evidence that oligodendrocyte changes are relevant to the pathophysiology and potentially the treatment of stress-related neuropsychiatric disorders., (© 2018 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.)

Adrenal cortex steroids are essential for body homeostasis, and adrenal insufficiency is a life-threatening condition. Adrenal endocrine activity is maintained through recruitment of subcapsular progenitor cells that follow a unidirectional differentiation path from zona glomerulosa to zona fasciculata (zF). Here, we show that this unidirectionality is ensured by the histone methyltransferase EZH2. Indeed, we demonstrate that EZH2 maintains adrenal steroidogenic cell differentiation by preventing expression of GATA4 and WT1 that cause abnormal dedifferentiation to a progenitor-like state in Ezh2 KO adrenals. EZH2 further ensures normal cortical differentiation by programming cells for optimal response to adrenocorticotrophic hormone (ACTH)/PKA signaling. This is achieved by repression of phosphodiesterases PDE1B, 3A, and 7A and of PRKAR1B. Consequently, EZH2 ablation results in blunted zF differentiation and primary glucocorticoid insufficiency. These data demonstrate an all-encompassing role for EZH2 in programming steroidogenic cells for optimal response to differentiation signals and in maintaining their differentiated state., Competing Interests: The authors declare no conflict of interest.

Phosphodiesterase 1C (PDE1C) is expressed in mammalian heart and regulates cardiac functions by controlling levels of second messenger cyclic AMP and cyclic GMP (cAMP and cGMP, respectively). However, molecular mechanisms of cardiac Pde1c regulation are currently unknown. In this study, we demonstrate that treatment of wild type mice and H9c2 myoblasts with Wy-14,643, a potent ligand of nuclear receptor peroxisome-proliferator activated receptor alpha (PPARα), leads to elevated cardiac Pde1C mRNA and cardiac PDE1C protein, which correlate with reduced levels of cAMP. Furthermore, using mice lacking either Pparα or cardiomyocyte-specific Med1 , the major subunit of Mediator complex, we show that Wy-14,643-mediated Pde1C induction fails to occur in the absence of Pparα and Med1 in the heart. Finally, using chromatin immunoprecipitation assays we demonstrate that PPARα binds to the upstream Pde1C promoter sequence on two sites, one of which is a palindrome sequence (agcTAGGttatcttaacctagc) that shows a robust binding. Based on these observations, we conclude that cardiac Pde1C is a direct transcriptional target of PPARα and that Med1 may be required for the PPARα mediated transcriptional activation of cardiac Pde1C .

Background: cAMP plays a critical role in regulating cardiomyocyte survival. Various cAMP signaling pathways behave distinctly or in opposition. We have previously reported that activation of cAMP hydrolysis by cyclic nucleotide phosphodiesterase 1C (PDE1C) promotes cardiomyocytes death/apoptosis, yet the underlying molecular mechanism remains unknown. In this study, we aimed to identify the specific cAMP signaling pathway modulated by PDE1C and determine the mechanism by which Ca 2+ /calmodulin-stimulated PDE1C is activated., Methods: To study cardiomyocyte death/apoptosis, we used both isolated mouse adult cardiomyocytes in vitro and doxorubicin-induced cardiotoxicity in vivo. We used a variety of pharmacological activators and inhibitors as well as genetically engineered molecular tools to manipulate the expression and activity of proteins of interest., Results: We found that the protective effect of PDE1C inhibition/deficiency on Ang II or doxorubicin-induced cardiomyocyte death/apoptosis is dependent on cAMP-generating adenosine A 2 receptors (A 2 Rs), suggesting that PDE1C's cAMP-hydrolyzing activity selectively modulates A 2 R-cAMP signaling in cardiomyocytes. In addition, we found that the effects of PDE1C activation on Ang II-mediated cAMP reduction and cardiomyocyte death are dependent on transient receptor potential-canonical (TRPC) channels, in particular TRPC3. We also observed synergistic protective effects on cardiomyocyte survival from the combination of A 2 R stimulation together with PDE1 or TRPC inhibition. Coimmunostaining and coimmunoprecipitation studies showed that PDE1C is localized in proximity with A 2 R and TRPC3 in the plasma membrane and perhaps T tubules. It is important to note that we found that doxorubicin-induced cardiac toxicity and dysfunction in mice are attenuated by the PDE1 inhibitor IC86340 or in PDE1C knockout mice, and this protective effect is significantly diminished by A 2 R antagonism., Conclusions: We have characterized a novel multiprotein complex comprised of A 2 R, PDE1C, and TRPC3, in which PDE1C is activated by TRPC3-derived Ca 2+ , thereby antagonizing A 2 R-cAMP signaling and promoting cardiomyocyte death/apoptosis. Targeting these molecules individually or in combination may represent a compelling therapeutic strategy for potentiating cardiomyocyte survival.

Background: Phosphodiesterase type-1 (PDE1) hydrolyzes cAMP and cGMP and is constitutively expressed in the heart, although cardiac effects from its acute inhibition in vivo are largely unknown. Existing data are limited to rodents expressing mostly the cGMP-favoring PDE1A isoform. Human heart predominantly expresses PDE1C with balanced selectivity for cAMP and cGMP. Here, we determined the acute effects of PDE1 inhibition in PDE1C-expressing mammals, dogs, and rabbits, in normal and failing hearts, and explored its regulatory pathways., Methods: Conscious dogs chronically instrumented for pressure-volume relations were studied before and after tachypacing-induced heart failure (HF). A selective PDE1 inhibitor (ITI-214) was administered orally or intravenously±dobutamine. Pressure-volume analysis in anesthetized rabbits tested the role of β-adrenergic and adenosine receptor signaling on ITI-214 effects. Sarcomere and calcium dynamics were studied in rabbit left ventricular myocytes., Results: In normal and HF dogs, ITI-214 increased load-independent contractility, improved relaxation, and reduced systemic arterial resistance, raising cardiac output without altering systolic blood pressure. Heart rate increased, but less so in HF dogs. ITI-214 effects were additive to β-adrenergic receptor agonism (dobutamine). Dobutamine but not ITI-214 increased plasma cAMP. ITI-214 induced similar cardiovascular effects in rabbits, whereas mice displayed only mild vasodilation and no contractility effects. In rabbits, β-adrenergic receptor blockade (esmolol) prevented ITI-214-mediated chronotropy, but inotropy and vasodilation remained unchanged. By contrast, adenosine A 2B -receptor blockade (MRS-1754) suppressed ITI-214 cardiovascular effects. Adding fixed-rate atrial pacing did not alter the findings. ITI-214 alone did not affect sarcomere or whole-cell calcium dynamics, whereas β-adrenergic receptor agonism (isoproterenol) or PDE3 inhibition (cilostamide) increased both. Unlike cilostamide, which further enhanced shortening and peak calcium when combined with isoproterenol, ITI-214 had no impact on these responses. Both PDE1 and PDE3 inhibitors increased shortening and accelerated calcium decay when combined with forskolin, yet only cilostamide increased calcium transients., Conclusions: PDE1 inhibition by ITI-214 in vivo confers acute inotropic, lusitropic, and arterial vasodilatory effects in PDE1C-expressing mammals with and without HF. The effects appear related to cAMP signaling that is different from that provided via β-adrenergic receptors or PDE3 modulation. ITI-214, which has completed phase I trials, may provide a novel therapy for HF.

Midbrain dopamine neurons are well known for their role in reward-based reinforcement learning. We found that the activity of dopamine axons in the posterior tail of the striatum (TS) scaled with the novelty and intensity of external stimuli, but did not encode reward value. We demonstrated that the ablation of TS-projecting dopamine neurons specifically inhibited avoidance of novel or high-intensity stimuli without affecting animals' initial avoidance responses, suggesting a role in reinforcement rather than simply in avoidance itself. Furthermore, we found that animals avoided optogenetic activation of dopamine axons in TS during a choice task and that this stimulation could partially reinstate avoidance of a familiar object. These results suggest that TS-projecting dopamine neurons reinforce avoidance of threatening stimuli. More generally, our results indicate that there are at least two axes of reinforcement learning using dopamine in the striatum: one based on value and one based on external threat.

New Findings: What is the central question of this study? The ciliary beat frequency (CBF) of the airway is controlled by [Ca 2+ ] i . However, the effects of a reduction in [Ca 2+ ] i on CBF are still controversial (an increase, a decrease or no change). What is the main finding and its importance? This study demonstrated that [Ca 2+ ] i directly regulates CBF (direct action) and also indirectly regulates CBF via cAMP accumulation controlled by Ca 2+ -dependent PDE1 activity (indirect action). The final CBF is determined by the balance of direct and indirect actions. PDE1 plays crucial roles in the regulation of airway CBF. ABSTRACT: [Ca 2+ ] i plays crucial roles in the regulation of ciliary beat frequency (CBF) and ciliary bend angle (CBA) of airway cilia. Moreover, Ca 2+ -dependent PDE1A existing in the CBF-regulating metabolon of cilia modifies the CBF by regulating the cAMP accumulation. This study demonstrated that the CBF is regulated by a direct and an indirect action of [Ca 2+ ] i ; the direct action changes CBF mediated via [Ca 2+ ] i , and the indirect action changes CBF mediated via cAMP, the accumulation of which is controlled by PDE1 activity. Upon reducing [Ca 2+ ] i to various levels, the direct action decreases CBF and the indirect action increases CBF. The final CBF is determined by the extent of cAMP accumulation, which is determined by the amount of inhibition of PDE1 activity, dependent on a reduction in [Ca 2+ ] i ; a slight decrease induced by a nominally Ca 2+ -free solution (no cAMP accumulation via PDE1) decreases CBF, and an extreme decrease induced by 50 μm BAPTA-AM increases CBF via cAMP accumulation by inhibiting PDE1 in a similar manner to a PDE1 inhibitor (8MmIBMX). The increase in CBA in response to a reduction in [Ca 2+ ] i is smaller than the increase in CBF, because no PDE1A exists in the CBA-regulating metabolon. On the contrary, an increase in [Ca 2+ ] i induced by ionomycin, which decreases cAMP accumulation by PDE1A activation, caused a slower procaterol-stimulated increase in CBF than that decreased by a Ca 2+ -free solution. A decrease in [Ca 2+ ] i stimulates cAMP accumulation, whereas an increase in [Ca 2+ ] i inhibits cAMP accumulation in airway ciliary cells. Thus, changes in [Ca 2+ ] i modulate CBF and CBA via cAMP accumulation by controlling the activity of PDE1., (© 2017 The Authors. Experimental Physiology © 2017 The Physiological Society.)

Sei-hai-to (TJ-90, Qing Fei Tang), a Chinese traditional medicine, increases ciliary beat frequency (CBF) and ciliary bend angle (CBA) mediated via cAMP (3',5'-cyclic adenosine monophosphate) accumulation modulated by Ca 2+ -activated phosphodiesterase 1 (PDE1A). A high concentration of TJ-90 (≥40 μg/mL) induced two types of CBF increases, a transient increase (an initial increase, followed by a decrease) and a sustained increase without any decline, while it only sustained the CBA increase. Upon inhibiting increases in intracellular Ca 2+ concentration ([Ca 2+ ] i ) by 10 μM BAPTA-AM (Ca 2+ -chelator, 1,2-Bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis(acetoxymethyl ester) or Ca 2+ /calmodulin-dependent PDE1 by 8MmIBMX (a selective PDE1 inhibitor), TJ-90 (400 μg/mL) induced only the sustained CBF increase without any transient CBF increase. The two types of the CBF increase (the transient increase and the sustained increase) induced by TJ-90 (≥40 μg/mL) were mimicked by the stimulation with both procaterol (100 pM) and ionomycin (500 nM). Thus, TJ-90 stimulates small increases in the intracellular cAMP concentration ([cAMP] i ) and [Ca 2+ ] i in airway ciliary cells of mice. These small increases in [cAMP] i and [Ca 2+ ] i cause inducing a transient CBF increase or a sustained CBF increase in an airway ciliary cells, depending on the dominant signal, Ca 2+ -signal, or cAMP-signal., Competing Interests: The authors declare no conflict of interest.

Background: Previous studies have confirmed a family risk of nephrolithiasis (NL), but only 15% of all cases are associated with an identified monogenic factor. In clinical practice, our group encountered a patient with NL combined with cystic kidney disease that had 3 affected family members. No known mutations association with NL was detected in this family, and thus further investigation of the molecular cause of NL was deemed to be necessary., Results: Quality analysis from the sequencing stage showed a more than 80-fold average depth and 95% coverage for each sample, and six mutations within six genes were chosen as candidate variants for further validation. Genotyping of rs182089527in the phosphodiesterase 1A (PDE1A) gene in the validation cohort indicated that the alternative allele was present in 15 patients with heterozygosity and in 1 patient with homozygosity, and exhibited significant enrichment in NL patients (Fisher's exact test, adjusted p = 0.0042) and kidney cystic patients (Fisher's exact test, adjusted p = 0.067) compared to controls. In addition, function analysis displayed a significant decrease in the protein and mRNA expression levels resulting from the rs182089527 mutant sequence compared with the wild-type sequence. Moreover, patients with this mutation displayed a high level of creatinine and urea in urinalysis., Conclusions: Our study provides genetic evidence that the rs182089527 mutation in PDE1A is involved in the development of NL and kidney cysts, which should help to improve personalized medicine for diagnosis and treatment.

Phosphodiesterase-1b (Pde1b) is highly expressed in striatum, dentate gyrus, CA3 and substantia nigra. In a new Floxed Pde1b × Cre CMV global knockout (KO) mouse model, we show an immobility-resistance phenotype that recapitulates that found in constitutive Pde1b KO mice. We use this new mouse model to show that the resistance to acute stress-induced depression-like phenotype is not the product of changes in locomotor activity or reactivity to other stressors (learned helplessness, novelty suppressed feeding or dexamethasone suppression), and is not associated with anhedonia using the sucrose preference test. Using tamoxifen inducible Cre, we show that the immobility-resistant phenotype depends on the age of induction. The effect is present when Pde1b is Reduced from conception, P0 or P32, but not if reduced as adults (P60). We also mapped regional brain expression of PDE1B protein and of the Cre driver. These data add to the suggestion that PDE1B may be a target for drug development with therapeutic potential in depression alone or in combination with existing antidepressants., (© 2017 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.)

Aim of the Study: Idiopathic pulmonary fibrosis (IPF) is a lethal human disease with short survival time and few treatment options. In this study, we aim to demonstrate that cyclic nucleotide phosphodiesterase 1A (PDE1A), a Ca2+/calmodulin-stimulating PDE family member, plays a critical role in the induction of fibrosis and angiogenesis in the lung., Materials and Methods: To induce pulmonary damage, adult male SD rats were treated with bleomycin in a dose of 6 mg/kg body weight by a single intratracheal instillation. For in vivo silencing of PDE1A in rat lung, a nonspecific control siRNA or PDE1A-specific siRNA was used to treat rat through nasal instillation. Human normal pulmonary fibroblasts MRC-5 and hFL1 and rat lung fibroblasts were used as in vitro model. Immunohistochemistry and immunoflurescence staining were performed to detect PDE1A and α-SMA expression. Reverse transcription-qPCR was performed to detect microRNA and mRNA expression. In vitro wound healing assay was performed to detect pulmonary fibroblasts'mortality ability., Results: In vitro studies showed that PDE1A can stimulate lung fibroblasts to undergo myofibroblastic changes. This led to the identification of miR-541-5p as one of the miRNA candidates associated with bleomycin response. We found that miR-541-5p expression is downregulated in TGF-β-treated lung fibroblasts and the rat pulmonary fibrosis model. Overexpression of miR-541-5p in lung fibroblasts inhibited mortality of human lung fibroblasts., Conclusions: MiR-541-5p is a key effector in lung fibroblastsby by regulating PDE1A expression at protein translation level and its overexpression is protective against bleomycin-induced lung fibrosis.

Background and Purpose: The PDE enzymes (PDE1-11) hydrolyse and thus inactivate cyclic nucleotides and are important in the regulation of the cardiovascular system. Here,we have investigated the effects on the cardiovascular system, of two novel selective PDE1 inhibitors, Lu AF41228 and Lu AF58027., Experimental Approach: We used rat mesenteric small arteries (internal diameters of 200-300 μm), RT-PCR and measured isometric wall tension. Effects of Lu AF41228 and Lu AF58027 on heart rate and BP were assessed in both anaesthetized and conscious male rats., Key Results: Nanomolar concentrations of Lu AF41228 and Lu AF58027 inhibited PDE1A, PDE1B and PDE1C enzyme activity, while micromolar concentrations were required to observe inhibitory effects at other PDEs. RT-PCR revealed expression of PDE1A, PDE1B and PDE1C in rat brain, heart and aorta, but only PDE1A and PDE1B in mesenteric arteries. In rat isolated mesenteric arteries contracted with phenylephrine or U46619, Lu AF41228 and Lu AF58027 induced concentration-dependent relaxations which were markedly reduced by inhibitors of guanylate cyclase, ODQ, and adenylate cyclase, SQ22536, and in preparations without endothelium. In anaesthetized rats, Lu AF41228 and Lu AF58027 dose-dependently lowered mean BP and increased heart rate. In conscious rats with telemetric pressure transducers, repeated dosing with Lu AF41228 lowered mean arterial BP 10-15 mmHg and increased heart rate., Conclusions and Implications: These novel PDE1 inhibitors induce vasodilation and lower BP, suggesting a potential use of these vasodilators in the treatment of hypertension and vasospasm., (© 2017 The British Pharmacological Society.)

It has been proposed that a reduction in intracellular calcium causes an increase in intracellular cAMP and PKA activity through stimulation of calcium inhibitable adenylyl cyclase 6 and inhibition of phosphodiesterase 1 (PDE1), the main enzymes generating and degrading cAMP in the distal nephron and collecting duct, thus contributing to the development and progression of autosomal dominant polycystic kidney disease (ADPKD). In zebrafish pde1a depletion aggravates and overexpression ameliorates the cystic phenotype. To study the role of PDE1A in a mammalian system, we used a TALEN pair to Pde1a exon 7, targeting the histidine-aspartic acid dipeptide involved in ligating the active site Zn++ ion to generate two Pde1a null mouse lines. Pde1a mutants had a mild renal cystic disease and a urine concentrating defect (associated with upregulation of PDE4 activity and decreased protein kinase A dependent phosphorylation of aquaporin-2) on a wild-type genetic background and aggravated renal cystic disease on a Pkd2WS25/- background. Pde1a mutants additionally had lower aortic blood pressure and increased left ventricular (LV) ejection fraction, without a change in LV mass index, consistent with the high aortic and low cardiac expression of Pde1a in wild-type mice. These results support an important role of PDE1A in the renal pathogenesis of ADPKD and in the regulation of blood pressure.

The threat of a dirty bomb which could cause internal contamination has been of major concern for the past decades. Because of their high chemical toxicity and their presence in the nuclear fuel cycle, uranium and neptunium are two actinides of high interest. Calmodulin (CaM) which is a ubiquitous protein present in all eukaryotic cells and is involved in calcium-dependent signaling pathways has a known affinity for uranyl and neptunyl ions. The impact of the complexation of these actinides on the physiological response of the protein remains, however, largely unknown. An isothermal titration calorimetry (ITC) was developed to monitor in vitro the enzymatic activity of the phosphodiesterase enzyme which is known to be activated by CaM and calcium. This approach showed that addition of actinyl ions (AnO 2 n+ ), uranyl (UO 2 2+ ) and neptunyl (NpO 2 + ), resulted in a decrease of the enzymatic activity, due to the formation of CaM-actinide complexes, which inhibit the enzyme and alter its interaction with the substrate by direct interaction. Results from dynamic light scattering rationalized this result by showing that the CaM-actinyl complexes adopted a specific conformation different from that of the CaM-Ca 2+ complex. The effect of actinides could be reversed using a hydroxypyridonate actinide decorporation agent (5-LIO(Me-3,2-HOPO)) in the experimental medium demonstrating its capacity to efficiently bind the actinides and restore the calcium-dependent enzyme activation., (Copyright © 2017 Elsevier Inc. All rights reserved.)

We studied the effects of water-soluble cationic dinitrosyl iron complexes with thiocarbamide and its aliphatic derivatives, new synthetic analogs of natural NO donors, active centers of nitrosyl [1Fe-2S]proteins, on activities of Ca 2+ -ATPase of sarcoplasmic reticulum and cGMP phosphodiesterase. Nitrosyl iron complexes [Fe(C 3 N 2 H 8 S)Cl(NO) 2 ] 0 [Fe(NO) 2 (C 3 N 2 H 8 S) 2 ] + Cl - (I), [Fe(SC(N(CH 3 ) 2 ) 2 (NO) 2 ]Cl (II), [Fe(SC(NH 2 ) 2 ) 2 (NO) 2 Cl×H 2 O (III), and [Fe(SC(NH 2 ) 2 ) 2 (NO) 2 ] 2 SO 4 ×H 2 O (IV) in a concentration of 10 -4 M completely inhibited the transporting and hydrolytic functions of Ca 2+ -ATPase. In a concentration of 10 -5 M, they inhibited active Ca 2+ transport by 57±6, 75±8, 80±8, and 85±9% and ATP hydrolysis by 0, 40±4, 48±5, and 38±4%, respectively. Complex II reversibly and noncompetitively inhibited the hydrolytic function of Ca 2+ -ATPase (Ki=1.7×10 -6 M). All the studied iron-sulphur complexes in a concentration of 10 -4 M inhibited cGMP phosphodiesterase function. These data suggest that the studied complexes can exhibit antimetastatic, antiaggregation, vasodilatatory, and antihypertensive activities.

Purpose: Pathological cardiac remodeling, characterized by cardiac hypertrophy and fibrosis, is a pathological feature of many cardiac disorders that leads to heart failure and cardiac arrest. Vinpocetine, a derivative of the alkaloid vincamine, has been used for enhancing cerebral blood flow to treat cognitive impairment. However, its role in pathological cardiac remodeling remains unknown. The aim of this study is to examine the effect of vinpocetine on pathological cardiac remodeling induced by chronic stimulation with angiotensin II (Ang II)., Methods: Mice received Ang II infusion via osmotic pumps in the presence of vehicle or vinpocetine. Cardiac hypertrophy and fibrosis were assessed by morphological, histological, and biochemical analyses. Mechanistic studies were carried out in vitro with isolated mouse adult cardiac myocytes and fibroblasts., Results: We showed that chronic Ang II infusion caused cardiac hypertrophy and fibrosis, which were all significantly attenuated by systemic administration of vinpocetine. In isolated adult mouse cardiomyocytes, vinpocetine suppressed Ang II-stimulated myocyte hypertrophic growth. In cultured cardiac fibroblasts, vinpocetine suppressed TGFβ-induced fibroblast activation and matrix gene expression, consistent with its effect in attenuating cardiac fibrosis. The effects of vinpocetine on cardiac myocyte hypertrophy and fibroblast activation are likely mediated by targeting cyclic nucleotide phosphodiesterase 1 (PDE1)., Conclusions: Our results reveal a novel protective effect of vinpocetine in attenuating pathological cardiac remodeling through suppressing cardiac myocyte hypertrophic growth and fibroblast activation and fibrotic gene expression. These studies may also shed light on developing novel therapeutic agents for antagonizing pathological cardiac remodeling.

The stress response has been largely modified in all domesticated animals, offering a strong tool for genetic mapping. In chickens, ancestral Red Junglefowl react stronger both in terms of physiology and behavior to a brief restraint stress than domesticated White Leghorn, demonstrating modified functions of the hypothalamic-pituitary-adrenal (HPA) axis. We mapped quantitative trait loci (QTL) underlying variations in stress-induced hormone levels using 232 birds from the 12th generation of an advanced intercross between White Leghorn and Red Junglefowl, genotyped for 739 genetic markers. Plasma levels of corticosterone, dehydroepiandrosterone (DHEA), and pregnenolone (PREG) were measured using LC-MS/MS in all genotyped birds. Transcription levels of the candidate genes were measured in the adrenal glands or hypothalamus of 88 out of the 232 birds used for hormone assessment. Genes were targeted for expression analysis when they were located in a hormone QTL region and were differentially expressed in the pure breed birds. One genome-wide significant QTL on chromosome 5 and two suggestive QTL together explained 20% of the variance in corticosterone response. Two significant QTL for aldosterone on chromosome 2 and 5 (explaining 19% of the variance), and one QTL for DHEA on chromosome 4 (explaining 5% of the variance), were detected. Orthologous DNA regions to the significant corticosterone QTL have been previously associated with the physiological stress response in other species but, to our knowledge, the underlying gene(s) have not been identified. SERPINA10 had an expression QTL (eQTL) colocalized with the corticosterone QTL on chromosome 5 and PDE1C had an eQTL colocalized with the aldosterone QTL on chromosome 2. Furthermore, in both cases, the expression levels of the genes were correlated with the plasma levels of the hormones. Hence, both these genes are strong putative candidates for the domestication-induced modifications of the stress response in chickens. Improved understanding of the genes associated with HPA-axis reactivity can provide insights into the pathways and mechanisms causing stress-related pathologies., (Copyright © 2017 Fallahsharoudi et al.)

Monitoring of neuronal activity within circuits facilitates integrated responses and rapid changes in behavior. We have identified a system in Caenorhabditis elegans where neuropeptide expression is dependent on the ability of the BAG neurons to sense carbon dioxide. In C. elegans, CO 2 sensing is predominantly coordinated by the BAG-expressed receptor-type guanylate cyclase GCY-9. GCY-9 binding to CO 2 causes accumulation of cyclic GMP and opening of the cGMP-gated TAX-2/TAX-4 cation channels; provoking an integrated downstream cascade that enables C. elegans to avoid high CO 2 . Here we show that cGMP regulation by GCY-9 and the PDE-1 phosphodiesterase controls BAG expression of a FMRFamide-related neuropeptide FLP-19 reporter (flp-19::GFP). This regulation is specific for CO 2 -sensing function of the BAG neurons, as loss of oxygen sensing function does not affect flp-19::GFP expression. We also found that expression of flp-19::GFP is controlled in parallel to GCY-9 by the activity-dependent transcription factor CREB (CRH-1) and the cAMP-dependent protein kinase (KIN-2) signaling pathway. We therefore show that two parallel pathways regulate neuropeptide gene expression in the BAG sensory neurons: the ability to sense changes in carbon dioxide and CREB transcription factor. Such regulation may be required in particular environmental conditions to enable sophisticated behavioral decisions to be performed., Competing Interests: The authors declare no competing financial interests.

Increasing evidence indicates that the white (w) gene in Drosophila possesses extra-retinal functions in addition to its classical role in eye pigmentation. We have previously shown that w+ promotes fast and consistent locomotor recovery from anoxia, but how w+ modulates locomotor recovery is largely unknown. Here we show that in the absence of w+, several PDE mutants, especially cyclic guanosine monophosphate (cGMP)-specific PDE mutants, display wildtype-like fast locomotor recovery from anoxia, and that during the night time, locomotor recovery was light-sensitive in white-eyed mutant w1118, and light-insensitive in PDE mutants under w1118 background. Data indicate the involvement of cGMP in the modulation of recovery timing and presumably, light-evoked cGMP fluctuation is associated with light sensitivity of locomotor recovery. This was further supported by the observations that w-RNAi-induced delay of locomotor recovery was completely eliminated by upregulation of cGMP through multiple approaches, including PDE mutation, simultaneous overexpression of an atypical soluble guanylyl cyclase Gyc88E, or sildenafil feeding. Lastly, prolonged sildenafil feeding promoted fast locomotor recovery from anoxia in w1118. Taken together, these data suggest that a White-cGMP interaction modulates the timing of locomotor recovery from anoxia., Competing Interests: The authors have declared that no competing interests exist.

An important hallmark of cardiac failure is abnormal second messenger signaling due to impaired synthesis and catabolism of cyclic adenosine 3',5'- monophosphate (cAMP) and cyclic guanosine 3',5'- monophosphate (cGMP). Their dysregulation, altered intracellular targeting, and blunted responsiveness to stimulating pathways all contribute to pathological remodeling, muscle dysfunction, reduced cell survival and metabolism, and other abnormalities. Therapeutic enhancement of either cyclic nucleotides can be achieved by stimulating their synthesis and/or by suppressing members of the family of cyclic nucleotide phosphodiesterases (PDEs). The heart expresses seven of the eleven major PDE subtypes - PDE1, 2, 3, 4, 5, 8, and 9. Their differential control over cAMP and cGMP signaling in various cell types, including cardiomyocytes, provides intriguing therapeutic opportunities to counter heart disease. This review examines the roles of these PDEs in the failing and hypertrophied heart and summarizes experimental and clinical data that have explored the utility of targeted PDE inhibition.

The focus of this chapter is on the cyclic nucleotide phosphodiesterase 1 (PDE1) family. PDE1 is one member of the 11 PDE families (PDE 1-11). It is the only phosphodiesterase family that is calcium/calmodulin activated. As a result, whereas other families of PDEs 2-11 play a dominant role controlling basal levels of cyclic nucleotides, PDE1 is involved when intra-cellular calcium levels are elevated and, thus, has an "on demand" or activity-dependent involvement in the control of cyclic nucleotides in excitatory cells including neurons, cardiomyocytes and smooth muscle. As a Class 1 phosphodiesterase, PDE1 hydrolyzes the 3' bond of 3'-5'-cyclic nucleotides, cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP). Here, we review evidence for this family of enzymes as drug targets for development of therapies aimed to address disorders of the central nervous system (CNS) and of degenerative diseases. The chapter includes sections on the potential for cognitive enhancement in mental disorders, as well as a review of PDE1 enzyme structure, enzymology, tissue distribution, genomics, inhibitors, pharmacology, clinical trials, and therapeutic indications. Information is taken from public databases. A number of excellent reviews of the phosphodiesterase family have been written as well as reviews of the PDE1 family. References cited here are not comprehensive, rather pointing to major reviews and key publications.