{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,12]],"date-time":"2025-10-12T02:14:31Z","timestamp":1760235271762,"version":"build-2065373602"},"reference-count":55,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2021,8,13]],"date-time":"2021-08-13T00:00:00Z","timestamp":1628812800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100011531","name":"University of Alabama","doi-asserted-by":"publisher","award":["RG14811"],"award-info":[{"award-number":["RG14811"]}],"id":[{"id":"10.13039\/100011531","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Crenshaw Research Fund","award":["A-52248-217101-100"],"award-info":[{"award-number":["A-52248-217101-100"]}]},{"name":"Sigma Xi Grants in Aid of Research","award":["NA"],"award-info":[{"award-number":["NA"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Biology"],"abstract":"<jats:p>B-cell lymphoma-extra large (Bcl-xL) is a mitochondrial protein known to inhibit mitochondria-dependent intrinsic apoptotic pathways. An increasing number of studies have demonstrated that Bcl-xL is critical in regulating neuronal energy metabolism and has a protective role in pathologies associated with an energy deficit. However, it is less known how Bcl-xL regulates physiological processes of the brain. In this study, we hypothesize that Bcl-xL is required for neurite branching and maturation during neuronal development by improving local energy metabolism. We found that the absence of Bcl-xL in rat primary hippocampal neurons resulted in mitochondrial dysfunction. Specifically, the ATP\/ADP ratio was significantly decreased in the neurites of Bcl-xL depleted neurons. We further found that neurons transduced with Bcl-xL shRNA or neurons treated with ABT-263, a pharmacological inhibitor of Bcl-xL, showed impaired mitochondrial motility. Neurons lacking Bcl-xL had significantly decreased anterograde and retrograde movement of mitochondria and an increased stationary mitochondrial population when Bcl-xL was depleted by either means. These mitochondrial defects, including loss of ATP, impaired normal neurite development. Neurons lacking Bcl-xL showed significantly decreased neurite arborization, growth and complexity. Bcl-xL depleted neurons also showed impaired synapse formation. These neurons showed increased intracellular calcium concentration and were more susceptible to excitotoxic challenge. Bcl-xL may support positioning of mitochondria at metabolically demanding regions of neurites like branching points. Our findings suggest a role for Bcl-xL in physiological regulation of neuronal growth and development.<\/jats:p>","DOI":"10.3390\/biology10080772","type":"journal-article","created":{"date-parts":[[2021,8,13]],"date-time":"2021-08-13T09:22:38Z","timestamp":1628846558000},"page":"772","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Bcl-xL Is Required by Primary Hippocampal Neurons during Development to Support Local Energy Metabolism at Neurites"],"prefix":"10.3390","volume":"10","author":[{"given":"Joseph","family":"Jansen","sequence":"first","affiliation":[{"name":"Department of Human Nutrition and Hospitality Management, College of Human Environmental Sciences, The University of Alabama, Tuscaloosa, AL 35487, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4377-4893","authenticated-orcid":false,"given":"Madison","family":"Scott","sequence":"additional","affiliation":[{"name":"Department of Human Nutrition and Hospitality Management, College of Human Environmental Sciences, The University of Alabama, Tuscaloosa, AL 35487, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Emma","family":"Amjad","sequence":"additional","affiliation":[{"name":"Department of Human Nutrition and Hospitality Management, College of Human Environmental Sciences, The University of Alabama, Tuscaloosa, AL 35487, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Allison","family":"Stumpf","sequence":"additional","affiliation":[{"name":"Department of Human Nutrition and Hospitality Management, College of Human Environmental Sciences, The University of Alabama, Tuscaloosa, AL 35487, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kimberly","family":"Lackey","sequence":"additional","affiliation":[{"name":"Department of Biological Sciences, College of Arts and Sciences, The University of Alabama, Tuscaloosa, AL 35487, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kim","family":"Caldwell","sequence":"additional","affiliation":[{"name":"Department of Biological Sciences, College of Arts and Sciences, The University of Alabama, Tuscaloosa, AL 35487, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5563-5255","authenticated-orcid":false,"given":"Han-A","family":"Park","sequence":"additional","affiliation":[{"name":"Department of Human Nutrition and Hospitality Management, College of Human Environmental Sciences, The University of Alabama, Tuscaloosa, AL 35487, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2021,8,13]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"597","DOI":"10.1016\/0092-8674(93)90508-N","article-title":"bcl-x, a bcl-2-related gene that functions as a dominant regulator of apoptotic cell death","volume":"74","author":"Boise","year":"1993","journal-title":"Cell"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1267","DOI":"10.1002\/jnr.21584","article-title":"Postnatal developmental regulation of Bcl-2 family proteins in brain mitochondria","volume":"86","author":"Soane","year":"2008","journal-title":"J. Neurosci. Res."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"983","DOI":"10.1126\/science.275.5302.983","article-title":"Structure of Bcl-xL-Bak peptide complex: Recognition between regulators of apoptosis","volume":"275","author":"Sattler","year":"1997","journal-title":"Science"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"554","DOI":"10.1038\/379554a0","article-title":"Bax-independent inhibition of apoptosis by Bcl-XL","volume":"379","author":"Cheng","year":"1996","journal-title":"Nature"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"1963","DOI":"10.1038\/cdd.2017.123","article-title":"Inhibition of Bcl-xL prevents pro-death actions of DeltaN-Bcl-xL at the mitochondrial inner membrane during glutamate excitotoxicity","volume":"24","author":"Park","year":"2017","journal-title":"Cell Death Differ."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"920","DOI":"10.1038\/cdd.2013.19","article-title":"PINK1 protects against cell death induced by mitochondrial depolarization, by phosphorylating Bcl-xL and impairing its pro-apoptotic cleavage","volume":"20","author":"Arena","year":"2013","journal-title":"Cell Death Differ."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1009","DOI":"10.1523\/JNEUROSCI.18-03-01009.1998","article-title":"Bcl-xL is an antiapoptotic regulator for postnatal CNS neurons","volume":"18","author":"Parsadanian","year":"1998","journal-title":"J. Neurosci."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"73","DOI":"10.1016\/j.neuroscience.2005.02.052","article-title":"Overexpression of Bcl-xl protects septal neurons from prolonged hypoglycemia and from acute ischemia-like stress","volume":"135","author":"Panickar","year":"2005","journal-title":"Neuroscience"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1168","DOI":"10.1016\/j.bbadis.2013.11.007","article-title":"Contributions of Bcl-xL to acute and long term changes in bioenergetics during neuronal plasticity","volume":"1842","author":"Jonas","year":"2013","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1706","DOI":"10.4103\/1673-5374.143413","article-title":"Mitochondrial membrane protein Bcl-xL, a regulator of adult neuronal growth and synaptic plasticity: Multiple functions beyond apoptosis","volume":"9","author":"Park","year":"2014","journal-title":"Neural Regen. Res."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"263","DOI":"10.1083\/jcb.201108059","article-title":"Bcl-xL regulates mitochondrial energetics by stabilizing the inner membrane potential","volume":"195","author":"Chen","year":"2011","journal-title":"J. Cell Biol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1224","DOI":"10.1038\/ncb2330","article-title":"Bcl-xL regulates metabolic efficiency of neurons through interaction with the mitochondrial F1FO ATP synthase","volume":"13","author":"Alavian","year":"2011","journal-title":"Nat. Cell Biol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"469","DOI":"10.1038\/s41419-019-1679-x","article-title":"Parkinson\u2019s disease protein DJ-1 regulates ATP synthase protein components to increase neuronal process outgrowth","volume":"10","author":"Chen","year":"2019","journal-title":"Cell Death Dis."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"2169","DOI":"10.1073\/pnas.0711647105","article-title":"Bcl-xL induces Drp1-dependent synapse formation in cultured hippocampal neurons","volume":"105","author":"Li","year":"2008","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"773","DOI":"10.1038\/ncb2791","article-title":"A Bcl-xL-Drp1 complex regulates synaptic vesicle membrane dynamics during endocytosis","volume":"15","author":"Li","year":"2013","journal-title":"Nat. Cell Biol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1668","DOI":"10.1111\/j.1600-0854.2007.00644.x","article-title":"Moving mitochondria: Establishing distribution of an essential organelle","volume":"8","author":"Frederick","year":"2007","journal-title":"Traffic"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"39","DOI":"10.4161\/auto.2229","article-title":"Tracker dyes to probe mitochondrial autophagy (mitophagy) in rat hepatocytes","volume":"2","author":"Kim","year":"2006","journal-title":"Autophagy"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"R169","DOI":"10.1093\/hmg\/ddp326","article-title":"Mitochondrial dynamics--fusion, fission, movement, and mitophagy--in neurodegenerative diseases","volume":"18","author":"Chen","year":"2009","journal-title":"Hum. Mol. Genet."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"893","DOI":"10.1016\/j.cell.2011.10.018","article-title":"PINK1 and Parkin target Miro for phosphorylation and degradation to arrest mitochondrial motility","volume":"147","author":"Wang","year":"2011","journal-title":"Cell"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"2088","DOI":"10.1242\/dev.114454","article-title":"Neuronal polarization","volume":"142","author":"Takano","year":"2015","journal-title":"Development"},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Safiulina, D., and Kaasik, A. (2013). Energetic and dynamic: How mitochondria meet neuronal energy demands. PLoS Biol., 11.","DOI":"10.1371\/journal.pbio.1001755"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"3922","DOI":"10.1016\/j.jmb.2018.07.027","article-title":"Mechanisms Orchestrating Mitochondrial Dynamics for Energy Homeostasis","volume":"430","author":"Yu","year":"2018","journal-title":"J. Mol. Biol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"502","DOI":"10.1016\/j.tcb.2007.07.008","article-title":"Mitochondria on the move","volume":"17","author":"Boldogh","year":"2007","journal-title":"Trends Cell Biol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"351","DOI":"10.1083\/jcb.201302040","article-title":"Kinesin-1-syntaphilin coupling mediates activity-dependent regulation of axonal mitochondrial transport","volume":"202","author":"Chen","year":"2013","journal-title":"J. Cell Biol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"137","DOI":"10.1016\/j.cell.2007.11.024","article-title":"Docking of axonal mitochondria by syntaphilin controls their mobility and affects short-term facilitation","volume":"132","author":"Kang","year":"2008","journal-title":"Cell"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"315","DOI":"10.1152\/physrev.2000.80.1.315","article-title":"Mitochondria and neuronal survival","volume":"80","author":"Nicholls","year":"2000","journal-title":"Physiol. Rev."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"403","DOI":"10.1016\/j.tcb.2017.01.005","article-title":"The Interplay of Axonal Energy Homeostasis and Mitochondrial Trafficking and Anchoring","volume":"27","author":"Sheng","year":"2017","journal-title":"Trends Cell Biol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"651","DOI":"10.1016\/j.neuron.2017.09.055","article-title":"Mitostasis in Neurons: Maintaining Mitochondria in an Extended Cellular Architecture","volume":"96","author":"Misgeld","year":"2017","journal-title":"Neuron"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"216","DOI":"10.1111\/jnc.13731","article-title":"Mitochondrial dysfunction in Parkinson\u2019s disease","volume":"139","author":"Bose","year":"2016","journal-title":"J. Neurochem."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"849","DOI":"10.2174\/1568026615666150827095102","article-title":"Relationships Between Mitochondria and Neuroinflammation: Implications for Alzheimer\u2019s Disease","volume":"16","author":"Wilkins","year":"2016","journal-title":"Curr. Top. Med. Chem."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"707","DOI":"10.1083\/jcb.200809060","article-title":"Bcl-x L increases mitochondrial fission, fusion, and biomass in neurons","volume":"184","author":"Berman","year":"2009","journal-title":"J. Cell Biol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"93","DOI":"10.1089\/ars.2013.5570","article-title":"Bcl-xL Is Necessary for Neurite Outgrowth in Hippocampal Neurons","volume":"22","author":"Park","year":"2015","journal-title":"Antioxid. Redox Signal."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"1741","DOI":"10.1038\/nprot.2012.099","article-title":"Culturing pyramidal neurons from the early postnatal mouse hippocampus and cortex","volume":"7","author":"Beaudoin","year":"2012","journal-title":"Nat. Protoc."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"2406","DOI":"10.1038\/nprot.2006.356","article-title":"Culturing hippocampal neurons","volume":"1","author":"Kaech","year":"2006","journal-title":"Nat. Protoc."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1111\/tra.12456","article-title":"KymoAnalyzer: A software tool for the quantitative analysis of intracellular transport in neurons","volume":"18","author":"Neumann","year":"2017","journal-title":"Traffic"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"2453","DOI":"10.1093\/bioinformatics\/btr390","article-title":"Simple Neurite Tracer: Open source software for reconstruction, visualization and analysis of neuronal processes","volume":"27","author":"Longair","year":"2011","journal-title":"Bioinformatics"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"134","DOI":"10.1016\/j.jneumeth.2007.08.029","article-title":"NeuriteTracer: A novel ImageJ plugin for automated quantification of neurite outgrowth","volume":"168","author":"Pool","year":"2008","journal-title":"J. Neurosci. Methods"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"2550","DOI":"10.1038\/ncomms3550","article-title":"Imaging energy status in live cells with a fluorescent biosensor of the intracellular ATP-to-ADP ratio","volume":"4","author":"Tantama","year":"2013","journal-title":"Nat. Commun."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"161","DOI":"10.1038\/nmeth.1288","article-title":"A genetically encoded fluorescent reporter of ATP:ADP ratio","volume":"6","author":"Berg","year":"2009","journal-title":"Nat. Methods"},{"key":"ref_40","first-page":"1981","article-title":"Mitochondrial biogenesis is required for axonal growth","volume":"143","author":"Vaarmann","year":"2016","journal-title":"Development"},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"509","DOI":"10.1124\/jpet.104.075945","article-title":"Mechanisms of acetaminophen-induced hepatotoxicity: Role of oxidative stress and mitochondrial permeability transition in freshly isolated mouse hepatocytes","volume":"312","author":"Reid","year":"2005","journal-title":"J. Pharmacol. Exp. Ther."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"1167","DOI":"10.1038\/cdd.2009.37","article-title":"Glutathione disulfide induces neural cell death via a 12-lipoxygenase pathway","volume":"16","author":"Park","year":"2009","journal-title":"Cell Death Differ."},{"key":"ref_43","doi-asserted-by":"crossref","unstructured":"Xu, B., Chen, S., Luo, Y., Chen, Z., Liu, L., Zhou, H., Chen, W., Shen, T., Han, X., and Chen, L. (2011). Calcium signaling is involved in cadmium-induced neuronal apoptosis via induction of reactive oxygen species and activation of MAPK\/mTOR network. PLoS ONE, 6.","DOI":"10.1371\/journal.pone.0019052"},{"key":"ref_44","first-page":"19","article-title":"Role of Bassoon and Piccolo in Assembly and Molecular Organization of the Active Zone","volume":"7","author":"Gundelfinger","year":"2015","journal-title":"Front. Synaptic. Neurosci."},{"key":"ref_45","doi-asserted-by":"crossref","unstructured":"Park, H.A., Mnatsakanyan, N., Broman, K., Davis, A.U., May, J., Licznerski, P., Crowe-White, K.M., Lackey, K.H., and Jonas, E.A. (2019). Alpha-Tocotrienol Prevents Oxidative Stress-Mediated Post-Translational Cleavage of Bcl-xL in Primary Hippocampal Neurons. Int. J. Mol. Sci., 21.","DOI":"10.3390\/ijms21010220"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"109","DOI":"10.1016\/j.yjmcc.2020.05.013","article-title":"ATP synthase c-subunit ring as the channel of mitochondrial permeability transition: Regulator of metabolism in development and degeneration","volume":"144","author":"Mnatsakanyan","year":"2020","journal-title":"J. Mol. Cell Cardiol."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"5823","DOI":"10.1038\/s41467-019-13766-2","article-title":"A mitochondrial megachannel resides in monomeric F1FO ATP synthase","volume":"10","author":"Mnatsakanyan","year":"2019","journal-title":"Nat. Commun."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"10580","DOI":"10.1073\/pnas.1401591111","article-title":"An uncoupling channel within the c-subunit ring of the F1FO ATP synthase is the mitochondrial permeability transition pore","volume":"111","author":"Alavian","year":"2014","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"557","DOI":"10.1002\/bip.22856","article-title":"Review: Structure and mechanism of the dynein motor ATPase","volume":"105","author":"Schmidt","year":"2016","journal-title":"Biopolymers"},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"671","DOI":"10.1038\/373671a0","article-title":"Pathway of processive ATP hydrolysis by kinesin","volume":"373","author":"Gilbert","year":"1995","journal-title":"Nature"},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"12865","DOI":"10.1073\/pnas.0403429101","article-title":"Multiple ATP-hydrolyzing sites that potentially function in cytoplasmic dynein","volume":"101","author":"Takahashi","year":"2004","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"299","DOI":"10.1016\/j.molcel.2010.01.025","article-title":"The BCL-2 family reunion","volume":"37","author":"Chipuk","year":"2010","journal-title":"Mol. Cell"},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"341","DOI":"10.1016\/S1074-7613(03)00234-6","article-title":"The structure of a Bcl-xL\/Bim fragment complex: Implications for Bim function","volume":"19","author":"Liu","year":"2003","journal-title":"Immunity"},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"287","DOI":"10.1016\/S1097-2765(00)80456-6","article-title":"The proapoptotic activity of the Bcl-2 family member Bim is regulated by interaction with the dynein motor complex","volume":"3","author":"Puthalakath","year":"1999","journal-title":"Mol. Cell"},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"647","DOI":"10.1038\/nrc883","article-title":"The Bcl2 family: Regulators of the cellular life-or-death switch","volume":"2","author":"Cory","year":"2002","journal-title":"Nat. Rev. Cancer"}],"container-title":["Biology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2079-7737\/10\/8\/772\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T06:45:42Z","timestamp":1760165142000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2079-7737\/10\/8\/772"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,8,13]]},"references-count":55,"journal-issue":{"issue":"8","published-online":{"date-parts":[[2021,8]]}},"alternative-id":["biology10080772"],"URL":"https:\/\/doi.org\/10.3390\/biology10080772","relation":{},"ISSN":["2079-7737"],"issn-type":[{"type":"electronic","value":"2079-7737"}],"subject":[],"published":{"date-parts":[[2021,8,13]]}}}