{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,18]],"date-time":"2026-06-18T23:50:13Z","timestamp":1781826613723,"version":"3.54.5"},"reference-count":37,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2023,6,26]],"date-time":"2023-06-26T00:00:00Z","timestamp":1687737600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Neurosci."],"abstract":"<jats:p>Death of dopaminergic (DAergic) neurons in the <jats:italic>substantia nigra pars compacta<\/jats:italic> of the human brain is the characteristic pathological feature of Parkinson\u2019s disease (PD). On exposure to neurotoxicants, <jats:italic>Drosophila<\/jats:italic> too exhibits mobility defects and diminished levels of brain dopamine. In the fly model of sporadic PD, our laboratory has demonstrated that there is no loss of DAergic neuronal number, however, a significant reduction in fluorescence intensity (FI) of secondary antibodies that target the primary antibody-anti-tyrosine hydroxylase (TH). Here, we present a sensitive, economical, and repeatable assay to characterize neurodegeneration based on the quantification of FI of the secondary antibody. As the intensity of fluorescence correlates with the amount of TH synthesis, its reduction under PD conditions denotes the depletion in the TH synthesis, suggesting DAergic neuronal dysfunction. Reduction in TH protein synthesis is further confirmed through Bio-Rad Stain-Free Western Blotting. Quantification of brain DA and its metabolites (DOPAC and HVA) using HPLC-ECD further demonstrated the depleted DA level and altered DA metabolism as evident from enhanced DA turnover rate. Together all these PD marker studies suggest that FI quantification is a refined and sensitive method to understand the early stages of DAergic neurodegeneration. FI quantification is performed using ZEN 2012 SP2, a licensed software from Carl Zeiss, Germany. This method will be of good use to biologists, as it with few modifications, can also be implemented to characterize the extent of degeneration of different cell types. Unlike the expensive and cumbersome confocal microscopy, the present method using fluorescence microscopy will be a feasible option for fund-constrained neurobiology laboratories in developing countries.<\/jats:p>","DOI":"10.3389\/fnins.2023.1158858","type":"journal-article","created":{"date-parts":[[2023,6,26]],"date-time":"2023-06-26T18:39:48Z","timestamp":1687804788000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":5,"title":["Fluorescence microscopy-based sensitive method to quantify dopaminergic neurodegeneration in a Drosophila model of Parkinson\u2019s disease"],"prefix":"10.3389","volume":"17","author":[{"given":"Mohamad","family":"Ayajuddin","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rahul","family":"Chaurasia","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Abhik","family":"Das","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Priyanka","family":"Modi","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Limamanen","family":"Phom","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zevelou","family":"Koza","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sarat Chandra","family":"Yenisetti","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2023,6,26]]},"reference":[{"key":"ref1","doi-asserted-by":"publisher","first-page":"865","DOI":"10.1126\/science.1067389","article-title":"Chaperone suppression of alpha-synuclein toxicity in a Drosophila model for Parkinson\u2019s disease","volume":"295","author":"Auluck","year":"2002","journal-title":"Science"},{"key":"ref2","first-page":"433","article-title":"Quantification of dopamine and its metabolites in Drosophila brain using HPLC","volume-title":"Experiments with","author":"Ayajuddin","year":"2021"},{"key":"ref3","doi-asserted-by":"publisher","first-page":"896183","DOI":"10.3389\/fnmol.2022.896183","article-title":"Adult health and transition stage-specific rotenone mediated Drosophila model of Parkinson\u2019s disease: impact on late-onset neurodegenerative disease models","volume":"15","author":"Ayajuddin","year":"2022","journal-title":"Front Mol Neurosci."},{"key":"ref4","doi-asserted-by":"publisher","first-page":"701","DOI":"10.1242\/dmm.007575","article-title":"Genetic activation of Nrf2 signaling is sufficient to ameliorate neurodegenerative phenotypes in a Drosophila model of Parkinson's disease","volume":"4","author":"Barone","year":"2011","journal-title":"Dis. 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