Mapping Protein Distribution in the Canine Photoreceptor Sensory Cilium and Calyceal Processes by Ultrastructure Expansion Microscopy.

IF 5 2区 医学 Q1 OPHTHALMOLOGY
Kei Takahashi, Raghavi Sudharsan, William A Beltran
{"title":"Mapping Protein Distribution in the Canine Photoreceptor Sensory Cilium and Calyceal Processes by Ultrastructure Expansion Microscopy.","authors":"Kei Takahashi, Raghavi Sudharsan, William A Beltran","doi":"10.1167/iovs.66.2.1","DOIUrl":null,"url":null,"abstract":"<p><strong>Purpose: </strong>Photoreceptors are highly polarized sensory neurons, possessing a unique ciliary structure known as the photoreceptor sensory cilium (PSC). Vertebrates have two subtypes of photoreceptors: rods, which are responsible for night vision, and cones, which enable daylight vision and color perception. Despite the identification of functional and morphological differences between these subtypes, ultrastructural analysis of the PSC molecular architecture between rods and cones is still lacking. This study employed ultrastructure expansion microscopy (U-ExM) to characterize the PSC molecular architecture in canine retina.</p><p><strong>Methods: </strong>Canine neuroretinas (5-mm punches) were fixed in paraformaldehyde solution for either short or long durations. Additionally, 20-µm-thick cryosections from frozen archival retinal tissues fixed using the longer protocol were analyzed. A U-ExM protocol previously developed for mouse retina was adapted to these canine tissues with a battery of specific antibodies that label the various compartments of the PSC.</p><p><strong>Results: </strong>We demonstrated that U-ExM is applicable to both non-frozen and cryopreserved retinal tissues processed with standard paraformaldehyde fixation. Using this validated U-ExM protocol, we revealed the molecular localization of numerous ciliopathy-related proteins in canine photoreceptors. Furthermore, we identified significant architectural differences in the PSC, ciliary rootlet, and calyceal processes between canine rods and cones.</p><p><strong>Conclusions: </strong>U-ExM is a powerful tool for studying the PSC molecular architecture using frozen archival retinas that are processed following standard paraformaldehyde fixation and embedding protocols. The findings gained from this study pave the way for a better understanding of alterations in the molecular architecture of the PSC in canine models of retinal ciliopathies.</p>","PeriodicalId":14620,"journal":{"name":"Investigative ophthalmology & visual science","volume":"66 2","pages":"1"},"PeriodicalIF":5.0000,"publicationDate":"2025-02-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11798334/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Investigative ophthalmology & visual science","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1167/iovs.66.2.1","RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"OPHTHALMOLOGY","Score":null,"Total":0}
引用次数: 0

Abstract

Purpose: Photoreceptors are highly polarized sensory neurons, possessing a unique ciliary structure known as the photoreceptor sensory cilium (PSC). Vertebrates have two subtypes of photoreceptors: rods, which are responsible for night vision, and cones, which enable daylight vision and color perception. Despite the identification of functional and morphological differences between these subtypes, ultrastructural analysis of the PSC molecular architecture between rods and cones is still lacking. This study employed ultrastructure expansion microscopy (U-ExM) to characterize the PSC molecular architecture in canine retina.

Methods: Canine neuroretinas (5-mm punches) were fixed in paraformaldehyde solution for either short or long durations. Additionally, 20-µm-thick cryosections from frozen archival retinal tissues fixed using the longer protocol were analyzed. A U-ExM protocol previously developed for mouse retina was adapted to these canine tissues with a battery of specific antibodies that label the various compartments of the PSC.

Results: We demonstrated that U-ExM is applicable to both non-frozen and cryopreserved retinal tissues processed with standard paraformaldehyde fixation. Using this validated U-ExM protocol, we revealed the molecular localization of numerous ciliopathy-related proteins in canine photoreceptors. Furthermore, we identified significant architectural differences in the PSC, ciliary rootlet, and calyceal processes between canine rods and cones.

Conclusions: U-ExM is a powerful tool for studying the PSC molecular architecture using frozen archival retinas that are processed following standard paraformaldehyde fixation and embedding protocols. The findings gained from this study pave the way for a better understanding of alterations in the molecular architecture of the PSC in canine models of retinal ciliopathies.

求助全文
约1分钟内获得全文 求助全文
来源期刊
CiteScore
6.90
自引率
4.50%
发文量
339
审稿时长
1 months
期刊介绍: Investigative Ophthalmology & Visual Science (IOVS), published as ready online, is a peer-reviewed academic journal of the Association for Research in Vision and Ophthalmology (ARVO). IOVS features original research, mostly pertaining to clinical and laboratory ophthalmology and vision research in general.
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信