Journal of Neuroscience Methods最新文献

{"title":"Adaptations to the neuronal culture for researchers at undergraduate institutions.","authors":"Briahna Galvan, Taran Singh, Prajwal Kurup, Jake Alvarez, Atta Bettagi, Henry Stewart, Anuraj Nair, Szilard Menes, Meera Patel, Jary Y Delgado","doi":"10.1016/j.jneumeth.2025.110437","DOIUrl":"https://doi.org/10.1016/j.jneumeth.2025.110437","url":null,"abstract":"<p><strong>Background: </strong>The use of rat hippocampal neurons in culture has become an essential tool in neuroscience, enabling detailed study of excitatory synapse organization, neurotransmitter release, and mechanisms of synaptic plasticity. While these cultures provide valuable insights, the physiological relevance of this simplified in vitro system remains an ongoing discussion. Research indicates that cultured hippocampal neurons undergo key maturation processes, including the development of mature dendritic spines, within weeks, mirroring aspects of in vivo development. Importantly, cultured neurons offer unique experimental flexibility, facilitating single-neuron manipulations that is technically challenging or impractical in intact brain slices or with viral vectors. Despite these advantages, establishing cultures with minimal glial support-critical for experiments involving sparse labeling of extracellular proteins for single-particle tracking-often demands substantial time, expertise, and resources, making it difficult to implement in smaller laboratories with limited personnel and funding.</p><p><strong>New method: </strong>In this study, we present modifications to the standard hippocampal culture protocol designed to improve accessibility and usability in resource-limited settings, such as undergraduate-focused institutions.</p><p><strong>Results/comparison: </strong>Our protocol reduces costs, simplifies the culturing process, and minimizes time requirements, supporting robust neuronal cultures with physiological properties comparable to those of traditional methods. These adaptations enable the execution of sophisticated experiments, including single-molecule tracking, in personnel-limited research environments.</p><p><strong>Conclusions: </strong>This approach highlights the potential for undergraduate institutions to make significant contributions to scientific advancements, rather than being viewed solely as centers for undergraduate training.</p>","PeriodicalId":16415,"journal":{"name":"Journal of Neuroscience Methods","volume":" ","pages":"110437"},"PeriodicalIF":2.7,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143788411","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Determining the Atlas Correspondence of Desikan-Killiany-Tourville and Glasser MMP1 Atlases across Magnetic Field Strengths.","authors":"Madison Lewis, Nicholas Theis, Nidhi Girish, Konasale Prasad","doi":"10.1016/j.jneumeth.2025.110445","DOIUrl":"https://doi.org/10.1016/j.jneumeth.2025.110445","url":null,"abstract":"<p><strong>Background: </strong>Over sixty-six brain atlases exist to parcellate the brain based on cytoarchitecture, function, and connectivity. Because atlas choice depends on individual study goals and hypotheses, variability in findings contributes to challenges in replication, validation, and reconciling the results across studies. Our goal was to measure the intersection of three commonly used atlases and create a tool to find regional correspondence between the atlases.</p><p><strong>New method: </strong>This study used three independent samples of anatomical MRI data acquired with different B0 magnetic field strengths: 1.5 Tesla (T), 3T, and 7T. The Desikan-Killiany- Tourville (DKT) and Glasser atlases were used to parcellate the brain. Coefficient-of- variation of regional volumes was measured to evaluate regional variability across subjects in each atlas. DKT and Glasser parcellation correspondence was calculated to answer the shared question of what Glasser regions intersect with a DKT region and vice versa and to investigate consistency of the parcellations in relation to each other across a variety of individuals and image resolutions.</p><p><strong>Results: </strong>We found that regional correspondence was consistent across field strengths for the DKT and Glasser parcellations despite showing population variability in volume, age, and sex, and was validated in the Schaefer400 atlas. Parcellation intersection data along with sample code to calculate specific regional correspondence is available.</p><p><strong>Comparison with existing methods: </strong>Prior studies have attempted to reconcile multiple atlases, but did not compare voxel- by-voxel on real data.</p><p><strong>Conclusion: </strong>This analysis created a tool for researchers and can aid in comparisons with differing atlas choice and variable field strengths.</p>","PeriodicalId":16415,"journal":{"name":"Journal of Neuroscience Methods","volume":" ","pages":"110445"},"PeriodicalIF":2.7,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143788461","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Neurocounter - a deep learning framework for high-fidelity spatial localization of neurons.","authors":"Tamal Batabyal, Aijaz Ahmad Naik, Jaideep Kapur","doi":"10.1016/j.jneumeth.2025.110444","DOIUrl":"https://doi.org/10.1016/j.jneumeth.2025.110444","url":null,"abstract":"<p><strong>Background: </strong>Many neuroscientific applications require robust and accurate localization of neurons. It is still an unsolved problem because of the enormous variation in intensity, texture, spatial overlap, morphology, and background artifacts. In addition, curating a large dataset containing complete manual annotation of neurons from high-resolution images for training a classifier requires significant time and effort. In this work, we presented Neurocounter, a deep learning network to detect and localize neurons.</p><p><strong>New method: </strong>Neurocounter contains an encoder, a decoder and an attention module. It is trained on images containing incompletely-annotated neurons having highly varied morphology, and control images containing artifacts and background structures. During training, Neurocounter progressively labels the un-annotated neurons in the training data. It detects centers of neuron soma as the output.</p><p><strong>Results: </strong>Neurocounter's self-learning ability reduces the need for time-intensive complete annotation and ensures high accuracy in the localization of neurons across various brain regions (approximately 94% F1 score). Comparison with existing methods Neurocounter shows its efficacy over the state of the arts by significantly reducing false-positive detection (by at least 3%).</p><p><strong>Conclusions: </strong>Neurocounter offers precise neuron soma detection in various scenarios, such as with background artifacts, clutter and overlapped cell soma. This tool can be potentially used to reconstruct brain-wide 3D maps of activated neurons from 2D localization of neurons.</p>","PeriodicalId":16415,"journal":{"name":"Journal of Neuroscience Methods","volume":" ","pages":"110444"},"PeriodicalIF":2.7,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143788477","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Enhancing robustness of spatial filters in motor imagery based brain-computer interface via temporal learning","authors":"Wei Liang ,&nbsp;Ren Xu ,&nbsp;Xingyu Wang ,&nbsp;Andrzej Cichocki ,&nbsp;Jing Jin","doi":"10.1016/j.jneumeth.2025.110441","DOIUrl":"10.1016/j.jneumeth.2025.110441","url":null,"abstract":"<div><h3>Background</h3><div>In motor imagery-based brain-computer interface (MI-BCI) EEG decoding, spatial filtering play a crucial role in feature extraction. Recent studies have emphasized the importance of temporal filtering for extracting discriminative features in MI tasks. While many efforts have been made to optimize feature extraction externally, stabilizing features from spatial filtering remains underexplored.</div><div>New method: To address this problem, we propose an approach to improve the robustness of temporal features by minimizing instability in the temporal domain. Specifically, we utilize Jensen-Shannon divergence to quantify temporal instability and integrate decision variables to construct an objective function that minimizes this instability. Our method enhances the stability of variance and mean values in the extracted features, improving the identification of discriminative features and reducing the effects of instability.</div></div><div><h3>Results</h3><div>The proposed method was applied to spatial filtering models, and tested on two publicly datasets as well as a self-collected dataset. Results demonstrate that the proposed method significantly boosts classification accuracy, confirming its effectiveness in enhancing temporal feature stability.</div><div>Comparison with existing methods: We compared our method with spatial filtering methods, and the-state-of-the-art models. The proposed approach achieves the highest accuracy, with 92.43 % on BCI competition III IVa dataset, 84.45 % on BCI competition IV 2a dataset, and 73.18 % on self-collected dataset.</div></div><div><h3>Conclusions</h3><div>Enhancing the instability of temporal features contributes to improved MI-BCI performance. This not only improves classification performance but also provides a stable foundation for future advancements. The proposed method shows great potential for EEG decoding.</div></div>","PeriodicalId":16415,"journal":{"name":"Journal of Neuroscience Methods","volume":"418 ","pages":"Article 110441"},"PeriodicalIF":2.7,"publicationDate":"2025-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143780245","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Spinal cord trunk preparation for analyzing cross-segmental primary afferent signal transmission and modulation","authors":"Caifeng Shao ,&nbsp;Mingwei Zhao ,&nbsp;Kun Yang","doi":"10.1016/j.jneumeth.2025.110440","DOIUrl":"10.1016/j.jneumeth.2025.110440","url":null,"abstract":"<div><h3>Background</h3><div>The spinal cord dorsal horn is pivotal for primary afferent signal transmission and modulation. Primary afferent fibers from each dorsal root arrive at the dorsal horn and travel 1–2 segments caudally and rostrally. Usually, in vitro spinal cord slices or in vivo preparations are employed for primary afferent stimulation and patch-clamp recordings to assess input signals. However, the spinal cord slices lose \"intact\" cross-segmental pathways, and in vivo studies are technically challenging.</div></div><div><h3>New method</h3><div>Here, we describe the preparation of a spinal cord trunk for analyzing afferent signal cross-segmental transmission in adult rats. By combining patch-clamp recording, Lissauer's tract stimulation, and ambient temperature manipulation, our methods enable accessing primary afferent pathways within several segments.</div></div><div><h3>Results</h3><div>Our present spinal trunk preparation can be maintained healthy for about 5 h. Lissauer’s tract stimulation induced evoked excitatory postsynaptic currents (eEPSCs) recorded in 6–10 mm rostrally in the ipsilateral dorsal horn. The eEPSCs, spontaneous EPSCs (sEPSCs), and neural excitability can be modulated by ambient temperature rise. Neuropharmacological studies can also be conducted on this spinal trunk preparation.</div></div><div><h3>Compared with existing methods</h3><div>Compared with conventional in vitro spinal cord slices, our present method maintains a relatively intact cross-segment pathway in the dorsal horn; compared with in vivo study, it avoids mechanical vibration and other technical challenges in living animals.</div></div><div><h3>Conclusion</h3><div>The rodent spinal cord trunk can be maintained for an extended period in a fully submerged chamber; combined with patch clamp recordings, our protocol facilitates the study of primary afferent transmission and modulation in the dorsal horn within adjacent segments.</div></div>","PeriodicalId":16415,"journal":{"name":"Journal of Neuroscience Methods","volume":"418 ","pages":"Article 110440"},"PeriodicalIF":2.7,"publicationDate":"2025-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143747426","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Inverse relationship between nodal strength and nodal power: Insights from separate resting fMRI and EEG datasets","authors":"Tien-Wen Lee ,&nbsp;Gerald Tramontano","doi":"10.1016/j.jneumeth.2025.110438","DOIUrl":"10.1016/j.jneumeth.2025.110438","url":null,"abstract":"<div><h3>Background</h3><div>Regional neural response and network properties have traditionally been studied separately. However, growing evidence suggests a close interplay between regional activity and inter-regional connectivity. This study aimed to examine the relationship between global functional connectivity and regional spontaneous activity, termed the global-to-local relationship.</div></div><div><h3>New method</h3><div>Resting-state fMRI data were parcellated using MOSI (modular analysis and similarity measurements), enabling multi-resolution functional partitioning. For each parcellated cluster, the mean amplitude of low-frequency fluctuations (node power) and its average functional connectivity with the remaining cortex (node strength) were computed. Correlation analyses assessed their relationship. A supplementary analysis was conducted on EEG data (1–30 Hz).</div></div><div><h3>Results</h3><div>A significant negative correlation between node strength and regional power was observed in MRI datasets. One-sample t-tests confirmed robustness across different MOSI resolutions, with individual <em>P</em> values at the level 10⁻⁴ to 10⁻⁵. The negative relationship was also found in EEG data but was restricted to delta (1–4 Hz) and theta (4–8 Hz) bands.</div></div><div><h3>Comparison with existing methods</h3><div>This study introduces two key novel aspects. First, it applies MOSI to the entire cortex, enhancing the comprehensiveness of network analysis. Second, it examines the global influence on regional neural activity, rather than limiting the focus to a specific network.</div></div><div><h3>Conclusions</h3><div>A robust negative relationship between node strength and node power was consistently observed across both MRI and EEG datasets, particularly in lower frequency bands (up to 8 Hz). These findings suggest a framework for investigating how global connectivity shapes regional neural activity, with inhibitory coupling as a potential underlying mechanism.</div></div>","PeriodicalId":16415,"journal":{"name":"Journal of Neuroscience Methods","volume":"418 ","pages":"Article 110438"},"PeriodicalIF":2.7,"publicationDate":"2025-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143780250","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Three-dimensional imaging and computational quantitation as a novel approach to assess nerve fibers, enteric glial cells, mast cells, and the proximity of mast cells to the nerve fibers in human sigmoid mucosal biopsies from healthy subjects","authors":"Pu-Qing Yuan ,&nbsp;Tao Li ,&nbsp;Swapna Mahurkar-Joshi ,&nbsp;Jessica Sohn ,&nbsp;Lin Chang ,&nbsp;Yvette Taché","doi":"10.1016/j.jneumeth.2025.110436","DOIUrl":"10.1016/j.jneumeth.2025.110436","url":null,"abstract":"<div><h3>Background</h3><div>The visualization and quantitation of nerve fibers (NFs), enteric glial cells (EGCs), mast cells (MCs), and their spatial configurations in the human colonic mucosa represent considerable challenges due to the meshed network of these components and the arborizing of NFs in a three-dimensional (3D) structure.</div></div><div><h3>New method</h3><div>We developed a novel approach combining tissue clearing, 3D imaging and computerized quantitation of NFs, EGCs and MCs in sigmoid mucosal biopsies of healthy subjects using a modified CLARITY tissue clearing protocol and adapting Imaris Surfaces Rendering Technology.</div></div><div><h3>Results</h3><div>The cleared colonic biopsies are compatible with immunostaining using 10 marker antibodies and capable of generating 3D images rendering clear spatial views and computational quantitation of NFs, MCs, EGCs, in particular the proximity of MCs to NFs with Imaris 9.7–9.9.</div></div><div><h3>Comparison with existing methods</h3><div>Our modified tissue clearing protocol shortened the membrane lipid removal time to 1 day from the original 1–2 weeks and total tissue clearing time to 3–4 days from the original 2–4 weeks. The 3D images displayed a clear spatial landscape of NFs, MCs and EGCs in the biopsies which cannot be portrayed with 2D images acquired from sections. Computerized quantitation is faster than measuring manually, allowing us to quantify a larger number of samples with less bias.</div></div><div><h3>Conclusion</h3><div>The novel approach enables faster tissue clearing/immunolabeling, high-quality 3D imaging and precise computational quantitation of NFs, cells and proximity of MCs to NFs in human sigmoid biopsies which may allow new insight to detect alterations in colonic-related diseases.</div></div>","PeriodicalId":16415,"journal":{"name":"Journal of Neuroscience Methods","volume":"418 ","pages":"Article 110436"},"PeriodicalIF":2.7,"publicationDate":"2025-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143767687","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Minimizing hearing loss: A novel surgical approach for round window exposure in miniature pigs","authors":"Zhifeng Chen ,&nbsp;Xihang Chen ,&nbsp;Haiqiao Du ,&nbsp;Hongdong Liu ,&nbsp;Yun Hu ,&nbsp;Yun Gao ,&nbsp;Xiao Yang ,&nbsp;Juanjuan Li ,&nbsp;Xianhai Zeng ,&nbsp;Xiaojun Ji ,&nbsp;Weiwei Guo ,&nbsp;Wei Chen ,&nbsp;Shiming Yang","doi":"10.1016/j.jneumeth.2025.110439","DOIUrl":"10.1016/j.jneumeth.2025.110439","url":null,"abstract":"<div><h3>Background</h3><div>A delicate operation for exposing the round window without affecting the hearing function is critical for auditory research. Despite the different surgical approaches proposed for miniature pigs, hearing protection during operation is still challenging. The efficient solution to this problem is significant for accurate auditory research.</div></div><div><h3>New method</h3><div>Eight healthy Diannan miniature pigs, aged three months and of both sexes, with normal hearing, were utilized. Three pigs were employed for anatomical studies. Based on the findings from these anatomical investigations, we propose the facial nerve medial approach designed to expose the round window while preserving hearing function in the porcine model. The remaining five pigs were used to verify the surgical approach and hearing function.</div></div><div><h3>Results</h3><div>The anatomy study confirmed that the lateral side of the round window was the genu of the facial nerve. Removing the medial wall of the genu part of the facial nerve canal allows access to the tympanic cavity and exposes the round window. The facial nerve medial approach effectively exposed the round window in vivo, with no significant change in auditory brainstem response threshold immediately post-operation.</div></div><div><h3>Comparison with existing methods</h3><div>The facial nerve medial approach to exposing the round window preserved the sound transmission structures of the ear, maintaining hearing function post-operation.</div></div><div><h3>Conclusions</h3><div>The facial nerve medial approach with a retroauricular incision allows access to the tympanic cavity and exposes the round window without impacting hearing, making it ideal for accurate auditory research in miniature pigs.</div></div>","PeriodicalId":16415,"journal":{"name":"Journal of Neuroscience Methods","volume":"418 ","pages":"Article 110439"},"PeriodicalIF":2.7,"publicationDate":"2025-03-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143767686","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Comparison of neural responses to whisker and ultrasound stimulation using a novel dual-stimulation protocol","authors":"Ye Yuan,&nbsp;Tian Liu,&nbsp;Jue Wang","doi":"10.1016/j.jneumeth.2025.110435","DOIUrl":"10.1016/j.jneumeth.2025.110435","url":null,"abstract":"<div><h3>Background</h3><div>The sensory system allows organisms to perceive and respond to environmental stimuli. This study investigates neural response differences between whisker and ultrasound stimulation in rats to evaluate cortical specificity to sensory inputs.</div></div><div><h3>New method</h3><div>A novel dual-stimulation protocol combining a step motor and ultrasound system was developed to alternately stimulate the C2 whisker and corresponding barrel column region. Experiments were conducted under varying stimulation sequences (whisker-ultrasound and ultrasound-whisker) and time intervals (10 ms, 25 ms, and 100 ms). Neural response signals were recorded, and statistical analyses (ANOVA and T-test) were performed to compare response amplitudes and peak latencies.</div></div><div><h3>Results</h3><div>Whisker stimulation consistently elicited significantly stronger neural responses than ultrasound stimulation (<em>*p &lt; 0.05</em>), regardless of sequence or interval. The efficiency of neural responses to ultrasound was closely tied to frequency, with higher frequencies producing greater amplitudes and faster latencies. Notably, at a 25 ms interval in the ultrasound-whisker sequence, whisker responses were significantly enhanced compared to whisker stimulation alone, suggesting a pre-activation effect of ultrasound.</div></div><div><h3>Comparison with existing methods</h3><div>Unlike single-modal whisker or ultrasound stimulation, the dual-stimulation protocol can enhance sensory responses, highlighting its neuromodulatory potential.</div></div><div><h3>Conclusion</h3><div>This study reveals distinct cortical activation patterns induced by whisker and ultrasound stimulation. While whisker stimulation is more sensitive, ultrasound stimulation—when optimized for frequency and timing—can effectively modulate neural responses under dual-stimulation protocol. These findings provide insights into ultrasound-based neuromodulation and sensory processing.</div></div>","PeriodicalId":16415,"journal":{"name":"Journal of Neuroscience Methods","volume":"418 ","pages":"Article 110435"},"PeriodicalIF":2.7,"publicationDate":"2025-03-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143704597","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A comprehensive protocol for simplified mouse DRG fixation, processing and F4/80 immunohistochemistry: Overcoming common challenges","authors":"Nicolette Tay ,&nbsp;Ammar Alshammari ,&nbsp;Simranpreet Kaur ,&nbsp;Allison Pettit ,&nbsp;Erica Mu ,&nbsp;Anna Reid ,&nbsp;Ingrid Winkler ,&nbsp;Irina Vetter ,&nbsp;Hana Starobova","doi":"10.1016/j.jneumeth.2025.110434","DOIUrl":"10.1016/j.jneumeth.2025.110434","url":null,"abstract":"<div><h3>Background</h3><div>Dorsal root ganglia (DRGs) contain the cell bodies of sensory neurons and non-neuronal cells that play a role in the pathophysiology of painful inflammatory conditions, such as neuropathic pain. Immunohistochemistry (IHC) is a valuable tool for visualising and quantifying immune cell markers in DRGs, providing important insights into these mechanisms. However, isolating DRGs while preserving cell morphology for IHC staining is technically challenging due to their small size and location within the spinal column.</div></div><div><h3>Objective</h3><div>Using F4/80, a pan monocyte-macrophage marker, we present an optimised protocol for the fixation, harvesting, processing, and IHC staining of formalin-fixed-paraffin-embedded (FFPE) mouse DRGs. This method is designed to maintain tissue integrity and ensure compatibility with downstream histopathological analysis.</div></div><div><h3>New Method</h3><div>The entire spinal column of mouse was fixed in 10 % neutral-buffered formalin at room temperature for 24 h before DRG isolation. DRGs were processed for 9 h, and antigen retrieval was performed using proteinase K.</div></div><div><h3>Results</h3><div>The optimised immersion-fixation approach preserved cellular morphology and antigenicity, ensuring high-quality histological outcomes.</div></div><div><h3>Comparison with Existing Methods</h3><div>While transcardial perfusion remains the gold standard for tissue fixation, it is time-intensive, requires training and raises ethical concerns. Our optimised method of whole spinal column fixation with subsequent tissue isolation is non-invasive and reduces the time between death and fixation in comparison to post-isolation fixation. Additionally, it delivers histological quality likely comparable to that of perfusion-based techniques.</div></div><div><h3>Conclusion</h3><div>This protocol is supported by a grading system to help evaluate variables and select conditions best suited to their experimental goals.</div></div>","PeriodicalId":16415,"journal":{"name":"Journal of Neuroscience Methods","volume":"418 ","pages":"Article 110434"},"PeriodicalIF":2.7,"publicationDate":"2025-03-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143704596","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}