Lipid Alterations in the Amygdala and Frontal Cortex Detected by Magnetic Resonance Spectroscopy: Potential Imaging Markers for Alzheimer’s Disease

Lipid Alterations in the Amygdala and Frontal Cortex Detected by Magnetic Resonance Spectroscopy: Potential Imaging Markers for Alzheimer’s Disease | 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 Research Article Lipid Alterations in the Amygdala and Frontal Cortex Detected by Magnetic Resonance Spectroscopy: Potential Imaging Markers for Alzheimer’s Disease Luís Jesuino de Oliveira Andrade, Gabriela Correia Matos de Oliveira, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7033365/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Objective To quantitatively evaluate lipid levels in the amygdala and frontal cortex using Magnetic Resonance Spectroscopy (MRS) in individuals with AD compared to healthy controls. Methods Fifty Magnetic Resonance Imaging /MRS scans from patients with confirmed AD were compared to 50 age- and sex-matched controls. MRS data were acquired at 1.5T focusing on spectral peaks associated with lipids. A voxel-based MRS technique was used to assess lipid concentrations in defined frequency ranges. Lipid concentrations were quantified, and statistical comparisons were performed between groups using the Shapiro-Wilk test. In cases where normality was not met, nonparametric Mann-Whitney U tests were employed. Results AD patients showed elevated lipid signals in both regions: 2.5–3.5 parts per million (ppm) in the amygdala and 3.0–4.5 ppm in the frontal cortex. Statistical differences between groups were significant (p < 0.001). Conclusion These findings suggest that lipid alterations detected by MRS may reflect underlying neurodegenerative processes and could potentially serve as imaging markers for AD. Nuclear Medicine & Medical Imaging Alzheimer's disease Magnetic Resonance Spectroscopy Amygdala Frontal Cortex Lipids Figures Figure 1 Figure 2 Figure 3 Figure 4 INTRODUCTION The human brain has specific regions committed to cognition, emotion, and behavior. Among these, the amygdala and frontal cortex play fundamental roles in emotional regulation, decision-making, and executive function ( 1 ). With recent technological advancement in neuroimaging technology, most significantly magnetic resonance spectroscopy (MRS), non-invasive assessment of brain metabolite profiles, such as lipid composition, has been made possible ( 2 ). Lipids are essential components of neuronal membranes and myelin sheaths, playing key roles in signal transmission and cellular integrity ( 3 ). Alterations in lipid composition may indicate neurodegenerative processes, such as those observed in Alzheimer’s disease (AD) ( 4 ). The amygdala, located within the medial temporal lobe, is implicated in emotional processing and memory consolidation ( 5 ). Although the role of lipids in neuronal health is not fully understood, certain lipid species, especially myelin-related lipids, are fundamental for maintaining neuronal connectivity. Similarly, the frontal cortex, responsible for higher cognitive functions, contains a substantial amount of lipid-rich white matter ( 6 ). Changes in its lipid profile may correlate with cognitive decline and contribute to AD pathophysiology ( 7 ). While MRS has been increasingly used to investigate brain metabolism, few studies have focused on regional lipid changes in AD. Most research has emphasized global brain measures, leaving regional variations, such as those in the amygdala and frontal cortex, underexplored ( 8 ). Therefore, the objective of this study is to quantitatively evaluate lipid levels within the amygdala and frontal cortex using MRS in a cohort of individuals with AD compared to a healthy, age- and sex-matched control group. We hypothesize that patients with AD exhibit significantly altered lipid spectral profiles in the amygdala and frontal cortex compared to matched healthy controls, reflecting early neurodegenerative lipid dysregulation. METHODS Control Group Description and Study Population Fifty Magnetic Resonance Imaging (MRI) examinations from individuals with a confirmed diagnosis of AD were recruited for this study and contrasted with 50 normal MRI examinations. Participants were included if they met the established clinical diagnostic criteria for probable AD as per the National Institute on Aging-Alzheimer's Association guidelines ( 9 ). MRI data were acquired at the Luiz Eduardo Magalhães General Hospital (HBLEM) in Itabuna, Bahia, Brazil, over a twelve-month period from March 2024 to March 2025. Controls were screened to exclude any neurological, psychiatric, or systemic conditions that could affect brain metabolism. Subjects with AD were assessed for concurrent medication use, including cholinesterase inhibitors or memantine, and any comorbidities such as depression were documented. Subjects with significant comorbidities or unstable medical conditions were excluded to minimize confounding effects on lipid metabolism. Magnetic Resonance Imaging and Magnetic Resonance Spectroscopy All MRI data were collected using a General Electric Sigma Explorer 1.5T scanner located at the HBLEM. High-resolution T1-weighted images were acquired to provide detailed anatomical information. A voxel-based MRS technique was implemented to quantify lipid concentrations within the amygdala and frontal cortex, given the pivotal role of these regions in this investigation. The specific MRS acquisition parameters are outlined below: Voxel Placement The voxel was placed in the amygdala and frontal cortex using anatomical landmarks to ensure consistent localization. The amygdala voxel was centered at Montreal Neurological Institute coordinates (x = − 20, y = − 10, z = − 15), and the frontal cortex voxel at (x = − 30, y = 30, z = 15). MRS Parameters : Single-voxel MRS spectra were acquired using a PRESS sequence with the following parameters: echo time (TE) = 35 ms, repetition time (TR) = 2000 ms, number of acquisitions = 128. Specifically, the lipid spectral peaks were analyzed between 1.2–1.5 parts per million (ppm), a range corresponding to myelin lipids and other phospholipid components. Data Analysis MRS data underwent spectral processing and quantification of lipid concentrations using the open-source software, NMRPipe. The lipid content within the amygdala and frontal cortex was determined by calculating the ratio of total lipid signal intensity to water signal intensity, enabling inter-group comparisons. A more detailed analysis of lipid profiles was conducted to identify potential variations in the relative concentrations of key lipid species, such as myelin-associated lipids and phospholipids. These variations may signify alterations in neuronal function and myelination processes. Statistical Analysis Data normality was assessed using the Shapiro-Wilk test. In cases where normality was not met, nonparametric Mann-Whitney U tests were employed. Outliers were identified using Tukey’s method and excluded if beyond ± 3 standard deviations. Outliers were identified using the interquartile range method and were excluded from analyses if justified. Statistical significance was set at p < 0.05. All analyses were conducted using PSPP (public domain software). Ethical Considerations The study protocol was approved by the Ethics Committee of Luiz Eduardo Magalhães General Hospital, Itabuna, Bahia, Brazil. RESULTS The MRS analysis revealed significant differences in lipid content between individuals with AD and healthy controls in both the amygdala and frontal cortex. Lipid concentrations were quantified as the ratio of total lipid signal intensity to water signal intensity, allowing for standardized inter-group comparisons. Using MRS, we analyzed the lipid content of 50 healthy amygdala. The spectral data revealed lipid spectral peaks within a MRS frequency range of 0.5 to 1.5 ppm (Fig. 1 ). The analysis demonstrated a consistent lipid profile of amygdala across subjects, with minimal variability. The average lipid MRS frequency was calculated to be 0.92 ± 0.28 ppm, indicating subtle inter-individual differences. In the amygdala, AD patients exhibited a marked increase in lipid signal within the 1.5–4.0 ppm range, consistent with alterations in membrane lipids (Fig. 2 ). The mean lipid concentration in this region was significantly higher in the AD group (2.68 ± 0.49 ppm) compared to controls (0.92 ± 0.28 ppm), p < 0.001. Interindividual variability was notably greater among AD patients, suggesting heterogeneous patterns of lipid accumulation or metabolic dysregulation in this cohort. The MRS spectra obtained from the group of individuals without AD exhibited lipid spectral peaks localized within the 0.8 and 1.8 ppm frequency band of the MRS analysis (Fig. 3 ). In the frontal cortex, similar trends were observed, with AD subjects showing elevated lipid spectral peaks primarily within the 3.0–4.5 ppm range (Fig. 4 ). The mean lipid concentration in this region was 3.60 ± 0.48 ppm for the AD group versus 1.33 ± 0.26 ppm for controls, p < 0.001. Our analyses revealed a consistent lipid profile across subjects, indicating minimal inter-individual variability in the frontal cortex. The statistical analysis confirmed non-normal distribution of the data (Shapiro-Wilk test, p < 0.05); therefore, nonparametric Mann-Whitney U tests were applied. Significant differences between groups were maintained after outlier exclusion using Tukey’s method and interquartile range criteria, reinforcing the robustness of the observed lipid alterations. These results support the hypothesis that lipid metabolism is altered in key brain regions affected by AD pathology and suggest that MRS-derived lipid profiles may serve as potential biomarkers for detecting early neurodegenerative changes. DISCUSSION This study provides evidence for significant changes in lipid metabolism in important brain areas in AD. Using MRS, we have seen different lipid profiles in the amygdala and frontal cortex of AD subjects compared with healthy controls. The results show a consistent pattern of elevated lipid levels in both brain regions of patients with AD, suggesting that dysregulation of lipids might be part of the pathophysiology of this neurodegenerative disorder. These lipid alterations may serve as potential noninvasive biomarkers for AD, although direct histological confirmation via brain biopsy is beyond the scope of this work. Cerebral lipid levels can vary considerably between subjects, even among healthy ones. Among the factors that may explain these variations are age, sex, genetics, and subtle anatomic differences ( 10 ), Various techniques, particularly MRS, are employed to quantify lipids. However, the accuracy and sensitivity of MRS are influenced by several factors, including the equipment used, data acquisition parameters, and analysis methods ( 11 ). Although there is a large literature using MRS to investigate brain metabolism, defining an absolute maximum for amygdala and frontal lobe lipid content is difficult, as most consist of ranges or averages ( 12 ). The accurate identification and quantification of the lipid spectral peaks in MRS spectra can also vary across research groups because of differences in analytical methods and criteria. Lipids constitute a substantial portion of brain tissue, particularly within myelinated white matter, and their spectral signatures can be reliably detected using proton magnetic resonance spectroscopy ( 13 ). These lipids predominantly arise from the methyl and methylene moieties of fatty acids, phospholipids, and other lipid constituent’s integral to membrane architecture and cellular energy reserves ( 14 ). In neurologically healthy individuals, the amygdala demonstrates a distinct metabolic profile characterized by elevated activity, with lipid metabolism playing a prominent role, especially in white matter-rich subregions ( 15 ). Within the normal amygdala, lipid spectral peaks typically manifest as broad, low-intensity signals on MRS, reflecting the physiological presence of lipids. These signals are generally observed at chemical shifts ranging approximately from 0.9 to 1.5 ppm ( 16 ). Our findings are consistent with previous reports in the literature, demonstrating lipid spectral profiles in the amygdala that reflect the expected baseline composition of brain lipid content. The observed spectral patterns align with those described in earlier studies, supporting the notion of a stable lipid signature in this region under normal physiological conditions. The frontal lobe is involved in executive functions, motor control and complex thinking ( 17 ). Anatomically it is divided into several areas, the prefrontal cortex being the one responsible for decision making and social behavior ( 18 ). MRS have enabled the quantification of lipids in this area and the lipid signal at 1.3 ppm is an indicator of brain health. Studies show that healthy frontal lobes have lipids between 0.8 and 1.8 ppm, this is influenced by age, sex and cognitive demand ( 19 ). Furthermore, deviations from these values may be related to neurodegenerative conditions or neuropsychiatric disorders, so lipidomic is important to understand frontal lobe pathology ( 20 ). The combination of MRS with anatomical data helps us to understand the mechanisms that govern frontal lobe function. Our MRS findings in the frontal lobe of non-AD subjects align with published literature. Lipid quantification revealed a profile consistent with that reported in healthy frontal lobes. The congruence between our findings and existing knowledge underscores the potential of MRS as an important tool in lipidomics research for understanding frontal lobe pathology. Measuring lipid percentage by MRS within the frontal lobe provides important insights into neurodegenerative processes since lipid metabolism is often changed in AD ( 21 ). High lipid levels might indicate loss of membrane integrity and myelin degradation, which may be useful as markers of AD progression ( 22 ). Additionally, the role of the frontal lobe in cognitive functions further supports the need for lipid quantification in assessing the impact of AD on the health of neurons ( 23 ). Moreover, relating lipid measurements to clinical manifestations may permit the designing of targeted therapeutic interventions. Thus, lipid quantification in the frontal lobe can contribute not only to early diagnosis but also to a better understanding of the pathophysiology of AD ( 24 ). Our results align with the current literature regarding lipid changes in AD. The heightened spectral peaks identified in the frontal cortex of individuals with AD correspond with earlier research indicating an increased lipid presence in this specific brain area related to neurodegenerative mechanisms. Moreover, the uniform lipid profile observed in our control group reinforces the idea that variations in lipid composition within the frontal lobe could serve as a promising biomarker for AD. These results emphasize the important role of MRS in characterizing the biochemical alterations associated with neurodegenerative disorders. While our findings suggest lipid alterations in key brain regions of AD patients, these results should be interpreted cautiously due to study limitations, including sample size and cross-sectional design. Further validation in larger cohorts is warranted before clinical application. FINAL CONSIDERATIONS In summary, this study demonstrates significant alterations in lipid levels within the amygdala and frontal cortex of individuals with AD as detected by MRS. The distinct lipid profiles identified in these brain regions underscore a potential association between lipid metabolism dysregulation and the neurodegenerative processes characteristic of AD. In particular, the lipid spectral peaks observed in the amygdala voxel are consistent with prior findings, reinforcing the hypothesis that altered lipid composition is implicated in cognitive decline. Moreover, the relative stability of lipid levels in cognitively healthy controls highlights the deviations present in the AD cohort, suggesting that such lipid changes may serve as promising biomarkers for disease presence or progression. This study further confirms the utility and feasibility of MRS as a noninvasive tool for probing biochemical alterations in AD. These findings contribute valuable perceptions into the complex interplay between lipid metabolism in the amygdala and frontal cortex and neurodegeneration, with important implications for clinical assessment. While our results are encouraging, future research integrating lipidomic data within larger, longitudinal frameworks is essential to enhance diagnostic precision and to inform potential therapeutic avenues. Continued investigation is imperative to fully elucidate the role of cerebral lipid alterations in AD pathophysiology and their relevance. Declarations Conflict of Interest : None declared. References Pessoa L (2010) Emotion and cognition and the amygdala: from what is it? to what's to be done? Neuropsychologia 48(12):3416–3429. 10.1016/j.neuropsychologia.2010.06.038 Wilson M, Andronesi O, Barker PB, Bartha R, Bizzi A, Bolan PJ et al (2019) Methodological consensus on clinical proton MRS of the brain: Review and recommendations. Magn Reson Med 82(2):527–550. 10.1002/mrm.27742 Aureli M, Grassi S, Prioni S, Sonnino S, Prinetti A (2015) Lipid membrane domains in the brain. Biochim Biophys Acta 1851(8):1006–1016. 10.1016/j.bbalip.2015.02.001 Han X (2010) Multi-dimensional mass spectrometry-based shotgun lipidomics and the altered lipids at the mild cognitive impairment stage of Alzheimer's disease. Biochim Biophys Acta 1801(8):774–783. 10.1016/j.bbalip.2010.01.010 Zhang X, Kim J, Tonegawa S (2020) Amygdala Reward Neurons Form and Store Fear Extinction Memory. Neuron 105(6):1077–1093e7. 10.1016/j.neuron.2019.12.025 Obis E, Sol J, Andres-Benito P, Martín-Gari M, Mota-Martorell N, Galo-Licona JD et al (2023) Lipidomic Alterations in the Cerebral Cortex and White Matter in Sporadic Alzheimer's Disease. Aging Dis 14(5):1887–1916. 10.14336/AD.2023.0217 Otoki Y, Yu D, Shen Q, Sahlas DJ, Ramirez J, Gao F et al (2023) Quantitative Lipidomic Analysis of Serum Phospholipids Reveals Dissociable Markers of Alzheimer's Disease and Subcortical Cerebrovascular Disease. J Alzheimers Dis 93(2):665–682. 10.3233/JAD-220795 Murley AG, Tsvetanov KA, Rouse MA, Jones PS, Sværke K, Li W et al (2022) Proton magnetic resonance spectroscopy in frontotemporal lobar degeneration-related syndromes. Neurobiol Aging 111:64–70. 10.1016/j.neurobiolaging.2021.10.012 Hyman BT, Phelps CH, Beach TG, Bigio EH, Cairns NJ, Carrillo MC et al (2012) National Institute on Aging-Alzheimer's Association guidelines for the neuropathologic assessment of Alzheimer's disease. Alzheimers Dement 8(1):1–13. 10.1016/j.jalz.2011.10.007 Cuenca-Bermejo L, Prinetti A, Kublickiene K, Raparelli V, Kautzky-Willer A, Norris CM et al (2023) Fundamental neurochemistry review: Old brain stories - Influence of age and sex on the neurodegeneration-associated lipid changes. J Neurochem 166(3):427–452. 10.1111/jnc.15834 Currie S, Hadjivassiliou M, Craven IJ, Wilkinson ID, Griffiths PD, Hoggard N (2013) Magnetic resonance spectroscopy of the brain. Postgrad Med J 89(1048):94–106. 10.1136/postgradmedj-2011-130471 Zhu H, Barker PB (2011) MR spectroscopy and spectroscopic imaging of the brain. Methods Mol Biol 711:203–226. 10.1007/978-1-61737-992-5_9 Oz G, Alger JR, Barker PB, Bartha R, Bizzi A, Boesch C et al (2014) Clinical proton MR spectroscopy in central nervous system disorders. Radiology 270(3):658–679. 10.1148/radiol.13130531 Magnan C, Levin BE, Luquet S (2015) Brain lipid sensing and the neural control of energy balance. Mol Cell Endocrinol 418:3–8. 10.1016/j.mce.2015.09.019 Haga KK, Khor YP, Farrall A, Wardlaw JM (2009) A systematic review of brain metabolite changes, measured with 1H magnetic resonance spectroscopy, in healthy aging. Neurobiol Aging 30(3):353–363. 10.1016/j.neurobiolaging.2007.07.005 Schubert F, Kühn S, Gallinat J, Mekle R, Ittermann B (2017) Towards a neurochemical profile of the amygdala using short-TE 1 H magnetic resonance spectroscopy at 3 T. NMR Biomed 30(5). 10.1002/nbm.3685 Niedermeyer E (1998) Frontal lobe functions and dysfunctions. Clin Electroencephalogr 29(2):79–90. 10.1177/155005949802900206 Catani M (2019) The anatomy of the human frontal lobe. Handb Clin Neurol 163:95–122. 10.1016/B978-0-12-804281-6.00006-9 Landheer K, Gajdošík M, Juchem C (2020) A semi-LASER, single-voxel spectroscopic sequence with a minimal echo time of 20.1 ms in the human brain at 3 T. NMR Biomed 33(9):e4324. 10.1002/nbm.4324 Castellanos DB, Martín-Jiménez CA, Rojas-Rodríguez F, Barreto GE, González J (2021) Brain lipidomics as a rising field in neurodegenerative contexts: Perspectives with Machine Learning approaches. Front Neuroendocrinol 61:100899. 10.1016/j.yfrne.2021.100899 Su H, Rustam YH, Masters CL, Makalic E, McLean CA, Hill AF et al (2021) Characterization of brain-derived extracellular vesicle lipids in Alzheimer's disease. J Extracell Vesicles 10(7):e12089. 10.1002/jev2.12089 Yin F (2023) Lipid metabolism and Alzheimer's disease: clinical evidence, mechanistic link and therapeutic promise. FEBS J 290(6):1420–1453. 10.1111/febs.16344 Kawade N, Yamanaka K (2024) Novel insights into brain lipid metabolism in Alzheimer's disease: Oligodendrocytes and white matter abnormalities. FEBS Open Bio 14(2):194–216. 10.1002/2211-5463.13661 Moreno-Rodriguez M, Perez SE, Martinez-Gardeazabal J, Manuel I, Malek-Ahmadi M, Rodriguez-Puertas R et al (2024) Frontal Cortex Lipid Alterations During the Onset of Alzheimer's Disease. J Alzheimers Dis 98(4):1515–1532. 10.3233/JAD-231485 Additional Declarations The authors declare no competing interests. Cite Share Download PDF Status: Posted Version 1 posted 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. We do this by developing innovative software and high quality services for the global research community. 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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-7033365","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":479895732,"identity":"124a03bc-60ea-4969-969a-898f335d77ab","order_by":0,"name":"Luís Jesuino de Oliveira 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3","display":"","copyAsset":false,"role":"figure","size":67499,"visible":true,"origin":"","legend":"\u003cp\u003eLipid concentrations and tissue composition in normal frontal lobe.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSource:\u003c/strong\u003eStudy results.\u003c/p\u003e","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-7033365/v1/2b1a599e764f6aca96ab4ec1.png"},{"id":86141877,"identity":"5ced70ea-b57d-4e14-842c-ed931fbd2b83","added_by":"auto","created_at":"2025-07-07 08:30:14","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":68534,"visible":true,"origin":"","legend":"\u003cp\u003eLipid concentrations and tissue composition in AD frontal lobe.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSource:\u003c/strong\u003eStudy results.\u003c/p\u003e","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-7033365/v1/5469642bd9e28c49fbbe0669.png"},{"id":86141881,"identity":"30f41987-4c21-4e39-beef-bf419830ff0b","added_by":"auto","created_at":"2025-07-07 08:30:18","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":767377,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7033365/v1/efcae3ba-55b3-40f6-823f-0ef896b3291c.pdf"}],"financialInterests":"The authors declare no competing interests.","formattedTitle":"\u003cp\u003e\u003cstrong\u003eLipid Alterations in the Amygdala and Frontal Cortex Detected by Magnetic Resonance Spectroscopy: Potential Imaging Markers for Alzheimer’s Disease\u003c/strong\u003e\u003c/p\u003e","fulltext":[{"header":"INTRODUCTION","content":"\u003cp\u003eThe human brain has specific regions committed to cognition, emotion, and behavior. Among these, the amygdala and frontal cortex play fundamental roles in emotional regulation, decision-making, and executive function (\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e). With recent technological advancement in neuroimaging technology, most significantly magnetic resonance spectroscopy (MRS), non-invasive assessment of brain metabolite profiles, such as lipid composition, has been made possible (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eLipids are essential components of neuronal membranes and myelin sheaths, playing key roles in signal transmission and cellular integrity (\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e). Alterations in lipid composition may indicate neurodegenerative processes, such as those observed in Alzheimer’s disease (AD) (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe amygdala, located within the medial temporal lobe, is implicated in emotional processing and memory consolidation (\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e). Although the role of lipids in neuronal health is not fully understood, certain lipid species, especially myelin-related lipids, are fundamental for maintaining neuronal connectivity. Similarly, the frontal cortex, responsible for higher cognitive functions, contains a substantial amount of lipid-rich white matter (\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e). Changes in its lipid profile may correlate with cognitive decline and contribute to AD pathophysiology (\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eWhile MRS has been increasingly used to investigate brain metabolism, few studies have focused on regional lipid changes in AD. Most research has emphasized global brain measures, leaving regional variations, such as those in the amygdala and frontal cortex, underexplored (\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eTherefore, the objective of this study is to quantitatively evaluate lipid levels within the amygdala and frontal cortex using MRS in a cohort of individuals with AD compared to a healthy, age- and sex-matched control group. We hypothesize that patients with AD exhibit significantly altered lipid spectral profiles in the amygdala and frontal cortex compared to matched healthy controls, reflecting early neurodegenerative lipid dysregulation.\u003c/p\u003e "},{"header":"METHODS","content":"\u003cp\u003e \u003cb\u003eControl Group Description and Study Population\u003c/b\u003e \u003c/p\u003e\u003cp\u003eFifty Magnetic Resonance Imaging (MRI) examinations from individuals with a confirmed diagnosis of AD were recruited for this study and contrasted with 50 normal MRI examinations. Participants were included if they met the established clinical diagnostic criteria for probable AD as per the National Institute on Aging-Alzheimer's Association guidelines (\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e). MRI data were acquired at the Luiz Eduardo Magalhães General Hospital (HBLEM) in Itabuna, Bahia, Brazil, over a twelve-month period from March 2024 to March 2025. Controls were screened to exclude any neurological, psychiatric, or systemic conditions that could affect brain metabolism. Subjects with AD were assessed for concurrent medication use, including cholinesterase inhibitors or memantine, and any comorbidities such as depression were documented. Subjects with significant comorbidities or unstable medical conditions were excluded to minimize confounding effects on lipid metabolism.\u003c/p\u003e\u003cp\u003e \u003cb\u003eMagnetic Resonance Imaging and Magnetic Resonance Spectroscopy\u003c/b\u003e \u003c/p\u003e\u003cp\u003eAll MRI data were collected using a General Electric Sigma Explorer 1.5T scanner located at the HBLEM. High-resolution T1-weighted images were acquired to provide detailed anatomical information. A voxel-based MRS technique was implemented to quantify lipid concentrations within the amygdala and frontal cortex, given the pivotal role of these regions in this investigation. The specific MRS acquisition parameters are outlined below:\u003c/p\u003e\u003cp\u003e \u003cstrong\u003eVoxel Placement\u003c/strong\u003e \u003c/p\u003e\u003cp\u003eThe voxel was placed in the amygdala and frontal cortex using anatomical landmarks to ensure consistent localization. The amygdala voxel was centered at Montreal Neurological Institute coordinates (x = − 20, y = − 10, z = − 15), and the frontal cortex voxel at (x = − 30, y = 30, z = 15).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003e \u003cem\u003eMRS Parameters\u003c/em\u003e: Single-voxel MRS spectra were acquired using a PRESS sequence with the following parameters: echo time (TE) = 35 ms, repetition time (TR) = 2000 ms, number of acquisitions = 128. Specifically, the lipid spectral peaks were analyzed between 1.2–1.5 parts per million (ppm), a range corresponding to myelin lipids and other phospholipid components.\u003c/p\u003e\u003ch2\u003eData Analysis\u003c/h2\u003e\u003cp\u003eMRS data underwent spectral processing and quantification of lipid concentrations using the open-source software, NMRPipe. The lipid content within the amygdala and frontal cortex was determined by calculating the ratio of total lipid signal intensity to water signal intensity, enabling inter-group comparisons. A more detailed analysis of lipid profiles was conducted to identify potential variations in the relative concentrations of key lipid species, such as myelin-associated lipids and phospholipids. These variations may signify alterations in neuronal function and myelination processes.\u003c/p\u003e\u003ch2\u003eStatistical Analysis\u003c/h2\u003e\u003cp\u003eData normality was assessed using the Shapiro-Wilk test. In cases where normality was not met, nonparametric Mann-Whitney U tests were employed. Outliers were identified using Tukey’s method and excluded if beyond ± 3 standard deviations. Outliers were identified using the interquartile range method and were excluded from analyses if justified. Statistical significance was set at p \u0026lt; 0.05. All analyses were conducted using PSPP (public domain software).\u003c/p\u003e\u003cp\u003e \u003cb\u003eEthical Considerations\u003c/b\u003e \u003c/p\u003e\u003cp\u003e The study protocol was approved by the Ethics Committee of Luiz Eduardo Magalhães General Hospital, Itabuna, Bahia, Brazil.\u003c/p\u003e"},{"header":"RESULTS","content":"\u003cp\u003eThe MRS analysis revealed significant differences in lipid content between individuals with AD and healthy controls in both the amygdala and frontal cortex. Lipid concentrations were quantified as the ratio of total lipid signal intensity to water signal intensity, allowing for standardized inter-group comparisons.\u003c/p\u003e\n\u003cp\u003eUsing MRS, we analyzed the lipid content of 50 healthy amygdala. The spectral data revealed lipid spectral peaks within a MRS frequency range of 0.5 to 1.5 ppm (Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e\n\u003cp\u003eThe analysis demonstrated a consistent lipid profile of amygdala across subjects, with minimal variability. The average lipid MRS frequency was calculated to be 0.92\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28 ppm, indicating subtle inter-individual differences.\u003c/p\u003e\n\u003cp\u003eIn the amygdala, AD patients exhibited a marked increase in lipid signal within the 1.5\u0026ndash;4.0 ppm range, consistent with alterations in membrane lipids (Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e). The mean lipid concentration in this region was significantly higher in the AD group (2.68\u0026thinsp;\u0026plusmn;\u0026thinsp;0.49 ppm) compared to controls (0.92\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28 ppm), p\u0026thinsp;\u0026lt;\u0026thinsp;0.001. Interindividual variability was notably greater among AD patients, suggesting heterogeneous patterns of lipid accumulation or metabolic dysregulation in this cohort.\u003c/p\u003e\n\u003cp\u003eThe MRS spectra obtained from the group of individuals without AD exhibited lipid spectral peaks localized within the 0.8 and 1.8 ppm frequency band of the MRS analysis (Fig. \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e\n\u003cp\u003eIn the frontal cortex, similar trends were observed, with AD subjects showing elevated lipid spectral peaks primarily within the 3.0\u0026ndash;4.5 ppm range (Fig. \u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e). The mean lipid concentration in this region was 3.60\u0026thinsp;\u0026plusmn;\u0026thinsp;0.48 ppm for the AD group versus 1.33\u0026thinsp;\u0026plusmn;\u0026thinsp;0.26 ppm for controls, p\u0026thinsp;\u0026lt;\u0026thinsp;0.001. Our analyses revealed a consistent lipid profile across subjects, indicating minimal inter-individual variability in the frontal cortex.\u003c/p\u003e\n\u003cp\u003eThe statistical analysis confirmed non-normal distribution of the data (Shapiro-Wilk test, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05); therefore, nonparametric Mann-Whitney U tests were applied. Significant differences between groups were maintained after outlier exclusion using Tukey\u0026rsquo;s method and interquartile range criteria, reinforcing the robustness of the observed lipid alterations.\u003c/p\u003e\n\u003cp\u003eThese results support the hypothesis that lipid metabolism is altered in key brain regions affected by AD pathology and suggest that MRS-derived lipid profiles may serve as potential biomarkers for detecting early neurodegenerative changes.\u003c/p\u003e"},{"header":"DISCUSSION","content":"\u003cp\u003eThis study provides evidence for significant changes in lipid metabolism in important brain areas in AD. Using MRS, we have seen different lipid profiles in the amygdala and frontal cortex of AD subjects compared with healthy controls. The results show a consistent pattern of elevated lipid levels in both brain regions of patients with AD, suggesting that dysregulation of lipids might be part of the pathophysiology of this neurodegenerative disorder. These lipid alterations may serve as potential noninvasive biomarkers for AD, although direct histological confirmation via brain biopsy is beyond the scope of this work.\u003c/p\u003e \u003cp\u003eCerebral lipid levels can vary considerably between subjects, even among healthy ones. Among the factors that may explain these variations are age, sex, genetics, and subtle anatomic differences (\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e), Various techniques, particularly MRS, are employed to quantify lipids. However, the accuracy and sensitivity of MRS are influenced by several factors, including the equipment used, data acquisition parameters, and analysis methods (\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e). Although there is a large literature using MRS to investigate brain metabolism, defining an absolute maximum for amygdala and frontal lobe lipid content is difficult, as most consist of ranges or averages (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e). The accurate identification and quantification of the lipid spectral peaks in MRS spectra can also vary across research groups because of differences in analytical methods and criteria.\u003c/p\u003e \u003cp\u003eLipids constitute a substantial portion of brain tissue, particularly within myelinated white matter, and their spectral signatures can be reliably detected using proton magnetic resonance spectroscopy (\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e). These lipids predominantly arise from the methyl and methylene moieties of fatty acids, phospholipids, and other lipid constituent\u0026rsquo;s integral to membrane architecture and cellular energy reserves (\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e). In neurologically healthy individuals, the amygdala demonstrates a distinct metabolic profile characterized by elevated activity, with lipid metabolism playing a prominent role, especially in white matter-rich subregions (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e). Within the normal amygdala, lipid spectral peaks typically manifest as broad, low-intensity signals on MRS, reflecting the physiological presence of lipids. These signals are generally observed at chemical shifts ranging approximately from 0.9 to 1.5 ppm (\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e). Our findings are consistent with previous reports in the literature, demonstrating lipid spectral profiles in the amygdala that reflect the expected baseline composition of brain lipid content. The observed spectral patterns align with those described in earlier studies, supporting the notion of a stable lipid signature in this region under normal physiological conditions.\u003c/p\u003e \u003cp\u003eThe frontal lobe is involved in executive functions, motor control and complex thinking (\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e). Anatomically it is divided into several areas, the prefrontal cortex being the one responsible for decision making and social behavior (\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e). MRS have enabled the quantification of lipids in this area and the lipid signal at 1.3 ppm is an indicator of brain health. Studies show that healthy frontal lobes have lipids between 0.8 and 1.8 ppm, this is influenced by age, sex and cognitive demand (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e). Furthermore, deviations from these values may be related to neurodegenerative conditions or neuropsychiatric disorders, so lipidomic is important to understand frontal lobe pathology (\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e). The combination of MRS with anatomical data helps us to understand the mechanisms that govern frontal lobe function. Our MRS findings in the frontal lobe of non-AD subjects align with published literature. Lipid quantification revealed a profile consistent with that reported in healthy frontal lobes. The congruence between our findings and existing knowledge underscores the potential of MRS as an important tool in lipidomics research for understanding frontal lobe pathology.\u003c/p\u003e \u003cp\u003eMeasuring lipid percentage by MRS within the frontal lobe provides important insights into neurodegenerative processes since lipid metabolism is often changed in AD (\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e). High lipid levels might indicate loss of membrane integrity and myelin degradation, which may be useful as markers of AD progression (\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e). Additionally, the role of the frontal lobe in cognitive functions further supports the need for lipid quantification in assessing the impact of AD on the health of neurons (\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e). Moreover, relating lipid measurements to clinical manifestations may permit the designing of targeted therapeutic interventions. Thus, lipid quantification in the frontal lobe can contribute not only to early diagnosis but also to a better understanding of the pathophysiology of AD (\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e). Our results align with the current literature regarding lipid changes in AD. The heightened spectral peaks identified in the frontal cortex of individuals with AD correspond with earlier research indicating an increased lipid presence in this specific brain area related to neurodegenerative mechanisms. Moreover, the uniform lipid profile observed in our control group reinforces the idea that variations in lipid composition within the frontal lobe could serve as a promising biomarker for AD. These results emphasize the important role of MRS in characterizing the biochemical alterations associated with neurodegenerative disorders.\u003c/p\u003e \u003cp\u003eWhile our findings suggest lipid alterations in key brain regions of AD patients, these results should be interpreted cautiously due to study limitations, including sample size and cross-sectional design. Further validation in larger cohorts is warranted before clinical application.\u003c/p\u003e \u003cp\u003e \u003cb\u003eFINAL CONSIDERATIONS\u003c/b\u003e \u003c/p\u003e \u003cp\u003eIn summary, this study demonstrates significant alterations in lipid levels within the amygdala and frontal cortex of individuals with AD as detected by MRS. The distinct lipid profiles identified in these brain regions underscore a potential association between lipid metabolism dysregulation and the neurodegenerative processes characteristic of AD. In particular, the lipid spectral peaks observed in the amygdala voxel are consistent with prior findings, reinforcing the hypothesis that altered lipid composition is implicated in cognitive decline.\u003c/p\u003e \u003cp\u003eMoreover, the relative stability of lipid levels in cognitively healthy controls highlights the deviations present in the AD cohort, suggesting that such lipid changes may serve as promising biomarkers for disease presence or progression. This study further confirms the utility and feasibility of MRS as a noninvasive tool for probing biochemical alterations in AD.\u003c/p\u003e \u003cp\u003eThese findings contribute valuable perceptions into the complex interplay between lipid metabolism in the amygdala and frontal cortex and neurodegeneration, with important implications for clinical assessment. While our results are encouraging, future research integrating lipidomic data within larger, longitudinal frameworks is essential to enhance diagnostic precision and to inform potential therapeutic avenues. Continued investigation is imperative to fully elucidate the role of cerebral lipid alterations in AD pathophysiology and their relevance.\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch2\u003eConflict of Interest\u003c/b\u003e:\u003c/h2\u003e \u003cp\u003eNone declared.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003ePessoa L (2010) Emotion and cognition and the amygdala: from what is it? to what's to be done? 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J Alzheimers Dis 98(4):1515\u0026ndash;1532. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.3233/JAD-231485\u003c/span\u003e\u003cspan address=\"10.3233/JAD-231485\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Alzheimer's disease, Magnetic Resonance Spectroscopy, Amygdala, Frontal Cortex, Lipids","lastPublishedDoi":"10.21203/rs.3.rs-7033365/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7033365/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eObjective\u003c/h2\u003e \u003cp\u003eTo quantitatively evaluate lipid levels in the amygdala and frontal cortex using Magnetic Resonance Spectroscopy (MRS) in individuals with AD compared to healthy controls.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eFifty Magnetic Resonance Imaging /MRS scans from patients with confirmed AD were compared to 50 age- and sex-matched controls. MRS data were acquired at 1.5T focusing on spectral peaks associated with lipids. A voxel-based MRS technique was used to assess lipid concentrations in defined frequency ranges. Lipid concentrations were quantified, and statistical comparisons were performed between groups using the Shapiro-Wilk test. In cases where normality was not met, nonparametric Mann-Whitney U tests were employed.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eAD patients showed elevated lipid signals in both regions: 2.5\u0026ndash;3.5 parts per million (ppm) in the amygdala and 3.0\u0026ndash;4.5 ppm in the frontal cortex. Statistical differences between groups were significant (p\u0026thinsp;\u0026lt;\u0026thinsp;0.001).\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e \u003cp\u003eThese findings suggest that lipid alterations detected by MRS may reflect underlying neurodegenerative processes and could potentially serve as imaging markers for AD.\u003c/p\u003e","manuscriptTitle":"Lipid Alterations in the Amygdala and Frontal Cortex Detected by Magnetic Resonance Spectroscopy: Potential Imaging Markers for Alzheimer’s Disease","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-07-07 08:22:09","doi":"10.21203/rs.3.rs-7033365/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"ab6e42e9-7d0a-4b1a-b6b5-00f76d505028","owner":[],"postedDate":"July 7th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":50959268,"name":"Nuclear Medicine \u0026 Medical Imaging"}],"tags":[],"updatedAt":"2025-07-07T08:22:09+00:00","versionOfRecord":[],"versionCreatedAt":"2025-07-07 08:22:09","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7033365","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7033365","identity":"rs-7033365","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}