{"doi":"10.1016/j.jneumeth.2025.110500","title":"Development of high-content screening assay for gene silencing in adult sensory neurons","abstract":"BACKGROUND: High-content screening in post-mitotic neurons faces challenges due to low transfection efficiency and expensive viral or electroporation methods. To accelerate discovery specifically in adult sensory neurons, we sought a scalable, low-cost platform that preserves neuronal health. METHOD: We developed a 384-well lipid-based siRNA screening assay using adult EGFP-expressing Fischer 344 DRG neurons. Key optimizations included a systematic comparison of plate plastics and lipid/siRNA ratios and reagents, yielding maximal knock-down with minimal toxicity. RESULTS: Optimal conditions (0.12 μL reagent, 2.5 pmol siRNA/well) reduced EGFP fluorescence by ≥ 50 % in 45 % of neurons, with mean knockdown efficiencies up to 60 % and minimal impact on neurite length. PTEN-targeting siRNAs increased neurite outgrowth by 40 % (p < 0.001), while death siRNA reduced length by 30 % (p < 0.001), demonstrating sensitivity to both stimulatory and inhibitory gene perturbations. COMPARISON: Our approach offers substantially lower cost and higher throughput than alternatives. Relative to electroporation protocols for adult DRG neurons, reagent cost is reduced ∼12-fold and hands-on time drops from ∼2 days to ∼3 h, while eliminating specialized equipment. Notably, the assay was optimized for cost-efficiency and scalability; for instance, our 384-well format protocol can screen hundreds of genes in triplicate for under $10,000, making high-content screening feasible in smaller laboratories. CONCLUSIONS: This platform enables rapid, cost-effective evaluation of hundreds to thousands of candidate genes in adult sensory neurons, facilitating identification of neurite growth regulators.","journal":"Journal of Neuroscience Methods","year":2025,"id":567641,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":0,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9578,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2025-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1471950,"name":"David Do","orcid":"0000-0003-3604-5609","position":1,"is_corresponding":false},{"id":1471951,"name":"Richard Lie","orcid":"0000-0003-4389-7383","position":2,"is_corresponding":false},{"id":1471952,"name":"Grace Woodruff","orcid":"0000-0002-7052-1050","position":3,"is_corresponding":false},{"id":1472223,"name":"Rodrigo López González","orcid":null,"position":4,"is_corresponding":false},{"id":1471953,"name":"Gunnar Poplawski","orcid":"0000-0002-1917-3441","position":5,"is_corresponding":false},{"id":1472222,"name":"Phillip Canete","orcid":null,"position":0,"is_corresponding":true}],"reference_count":46,"raw_metadata":null,"created_at":"2026-07-19T02:56:48.183032Z","pmid":"40484286","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}