Glutamate Gradually Elevates [Zn2+]i via the CaM−CaMKII−NOS Cascade in Primary Cultured Rat Embryonic Cortical Neurons

Glutamate Gradually Elevates [Zn2+]i via the CaM−CaMKII−NOS Cascade in Primary Cultured Rat Embryonic Cortical Neurons | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Glutamate Gradually Elevates [Zn2+]i via the CaM−CaMKII−NOS Cascade in Primary Cultured Rat Embryonic Cortical Neurons Hui-Chiun Tseng, Yong-Sheng Wang, Chien-Yuan Pan This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4559368/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 30 Apr, 2025 Read the published version in Scientific Reports → Version 1 posted 10 You are reading this latest preprint version Abstract Zn2+ is essential for neuronal signaling, but imbalance cause cell death and neurodegenerative disorders. While the buffering system maintains low cytosolic Zn2+ concentration ([Zn2+]i), the details on physiological stimuli elevating [Zn2+]i for neuronal processes remain limited. Our previous reports have demonstrated that dopamine elevates [Zn2+]i through the cAMP-NO pathway, activating autophagy and inflammation in neurons. In this study, we adopted the Zn2+ imaging technique to verify how glutamate elevated [Zn2+]i in cultured cortical neurons and examined the inflammatory response. Our results showed that glutamate elevates the [Zn2+]i, by activating ionotropic glutamate receptors. Inhibitors of calmodulin (CaM), CaM-dependent protein kinase II (CaMKII), and NO synthase (NOS) blocked the glutamate-induced Zn2+ response. High-K+ buffer induced-membrane depolarization significantly elevated the intracellular Ca2+ concentration ([Ca2+]i) but only slightly increased [Zn2+]i and NO production. Glutamate also transiently increased NOS phosphorylation at Ser1417 within 15 min. The Zn2+ chelator, TPEN suppressed glutamate-induced inflammasome formation. These results indicate that glutamate-induced local increment in [Ca2+]i via the ionotropic glutamate receptors activates the CaM-CaMKII-NOS complex to produce NO and elevate [Zn2+]i. which trigger inflammation in cultured neurons. Henceforth, this novel glutamate-Zn2+ signaling pathway after glutamate depolarization elevates [Ca2+]i indicates the involvement of Zn2+ in modulating long-term neuronal activities. calmodulin calmodulin-dependent protein kinase II inflammation ionotropic glutamate receptor nitric oxide synthase Zn2+ Full Text Additional Declarations No competing interests reported. Supplementary Files ScientificreportsSupplementarymaterial20240610.pdf Cite Share Download PDF Status: Published Journal Publication published 30 Apr, 2025 Read the published version in Scientific Reports → Version 1 posted Editorial decision: Revision requested 15 Oct, 2024 Reviews received at journal 12 Oct, 2024 Reviewers agreed at journal 12 Sep, 2024 Reviews received at journal 11 Jul, 2024 Reviewers agreed at journal 17 Jun, 2024 Reviewers invited by journal 17 Jun, 2024 Editor assigned by journal 17 Jun, 2024 Editor invited by journal 17 Jun, 2024 Submission checks completed at journal 13 Jun, 2024 First submitted to journal 10 Jun, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4559368","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":319928733,"identity":"a0992654-e254-4c48-9de5-470a68295110","order_by":0,"name":"Hui-Chiun Tseng","email":"","orcid":"","institution":"Department of Life Science, National Taiwan University","correspondingAuthor":false,"prefix":"","firstName":"Hui-Chiun","middleName":"","lastName":"Tseng","suffix":""},{"id":319928735,"identity":"9113e2d9-c629-4818-866f-6118d39901cd","order_by":1,"name":"Yong-Sheng Wang","email":"","orcid":"","institution":"Department of Life Science, National Taiwan University","correspondingAuthor":false,"prefix":"","firstName":"Yong-Sheng","middleName":"","lastName":"Wang","suffix":""},{"id":319928736,"identity":"1302c8b4-d052-4252-96f1-fec172ccb326","order_by":2,"name":"Chien-Yuan Pan","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAt0lEQVRIiWNgGAWjYNCCCgglQYKWMxDVQMKASB2MbaRo4Z+Re/Bz4bw7dQYHmA/e5mH4k9hASIvEjbxk6ZnbnkkYHGBLtuZhMCCsxUAix0Cad9thoBYeM2mgllxitBj/5p0D0sL/jWgtZtK8DWBb2IjTInHmXZo1z7HDkjMPsxlbzjEwrieohb899/BtnprD/HzHmx/eeFMhZ0xIBwODQA6UwQx2J2ENQGvOEKNqFIyCUTAKRjQAAElRNcQ60hDRAAAAAElFTkSuQmCC","orcid":"","institution":"Department of Life Science, National Taiwan University","correspondingAuthor":true,"prefix":"","firstName":"Chien-Yuan","middleName":"","lastName":"Pan","suffix":""}],"badges":[],"createdAt":"2024-06-10 16:51:07","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4559368/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4559368/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1038/s41598-025-99142-1","type":"published","date":"2025-04-30T15:57:12+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":81987679,"identity":"d4a9edec-725e-4f13-8b05-47fb924bf280","added_by":"auto","created_at":"2025-05-05 16:04:38","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":728442,"visible":true,"origin":"","legend":"","description":"","filename":"ScientificreportsGlutamateZinc20240610submit.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4559368/v1_covered_91cafc7e-18be-425f-8766-d87c76cffd66.pdf"},{"id":59289523,"identity":"f6b07cf2-c945-422b-b20e-33fbf1fc5417","added_by":"auto","created_at":"2024-06-28 17:44:28","extension":"pdf","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":430870,"visible":true,"origin":"","legend":"","description":"","filename":"ScientificreportsSupplementarymaterial20240610.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4559368/v1/738953b40a0014f44f5e2b6e.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Glutamate Gradually Elevates [Zn2+]i via the CaM−CaMKII−NOS Cascade in Primary Cultured Rat Embryonic Cortical Neurons","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"calmodulin, calmodulin-dependent protein kinase II, inflammation, ionotropic glutamate receptor, nitric oxide synthase, Zn2+","lastPublishedDoi":"10.21203/rs.3.rs-4559368/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4559368/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":" Zn2+ is essential for neuronal signaling, but imbalance cause cell death and neurodegenerative disorders. 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The Zn2+ chelator, TPEN suppressed glutamate-induced inflammasome formation. These results indicate that glutamate-induced local increment in [Ca2+]i via the ionotropic glutamate receptors activates the CaM-CaMKII-NOS complex to produce NO and elevate [Zn2+]i. which trigger inflammation in cultured neurons. 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