The video demonstrates a cryosectioning technique for obtaining discrete dorsal root ganglion (DRG) tissue sections. It details the process of attaching DRG sections to glass slides without overlap, ensuring optimal placement for subsequent immunohistochemistry studies. The effectiveness of the method is confirmed through confocal fluorescence microscopy imaging.

cryosectioning of the dorsal root ganglion (drg)

Criosezionazione del ganglio della radice dorsale del topo

The dorsal root ganglion (DRG) contains the primary sensory neurons, the tissue macrophages, and the satellite cells that surround the primary sensory neurons1,2,3,4. It is a key anatomic structure in processing innocuous and noxious signals, and plays critical roles in pain, itch, and various peripheral nerve disorders5,6,7,8,9,10,11,12,13. Although several methods have been developed to dissect DRG tissue from the mouse spinal cord14,15,16, cryosectioning of the DRG tissue remains challenging as the DRG tissue is quite small, and cryostat sections of DRG samples tend to curve into rolls, making it difficult to properly transfer the cryostat sections onto glass slides. However, proper cryosectioning of the DRG tissue is crucial for immunohistochemistry studies and the structure of DRG sensory neurons17,18,19,20,21,22,23. Moreover, as single-cell RNA sequencing results have revealed the remarkable heterogeneity of DRG sensory neurons in both humans24 and mice25, proper cryosectioning of DRG tissue is critical for investigating the functional role of different DRG cells in various physiological and pathological conditions.
Although the tissue-clearing technique has been applied to investigate the 3D reconstruction of the DRG26 as an alternative technique of cryosectioning the DRG, the tissue-clearing technique is time- and labor-consuming. In comparison, cryosectioning of the DRG is quick and relatively easy to perform, and thus it remains to be a key technique for immunohistochemistry and structure studies of the DRG and other regions of the central nervous system. However, obtaining high-quality, intact, and flat cryostat sections onto glass slides remains to be a challenge in neuroscience research because of the tiny sample size of tissues, like the DRG and certain brain regions, and there is no article describing the optimal protocol at this point for cryosectioning small-sized tissue samples, such as mouse DRGs.
This protocol provides an easy, step-by-step technique for cryostat sectioning of the mouse DRG to reliably obtain as many high-quality DRG sections on the slides for subsequent DRG studies. While specifically designed for cryosectioning DRG samples, this technique can potentially be used for cryosectioning various other tissues with a small sample size.

Autore in primo piano: Una procedura per la criosezione del ganglio della radice dorsale del topo  

Una procedura per la criosezione del ganglio della radice dorsale del topo

Watch this Scientific Journal Video about Cryosectioning of the Dorsal Root Ganglion DRG at JoVE.com

Cryosectioning of the Dorsal Root Ganglion (DRG)  

Criosezione del ganglio della radice dorsale (DRG)

Iniziare la criosezione tenendo la sezione DRG a temperatura di taglio ottimale nella parte inferiore, utilizzando un piccolo pennello, pre-raffreddato a 20 gradi Celsius. Toccare delicatamente la parte inferiore della sezione utilizzando l'estremità di una pinzetta a temperatura ambiente in modo che aderisca alla superficie della piattaforma. Quindi, posiziona lentamente un vetrino caricato sulla sezione.Non appena la sezione si attacca alla slitta, tirare delicatamente la slitta all'indietro. Seguire la stessa procedura per ottenere più sezioni DRG sulla diapositiva senza sovrapporre le sezioni. Il corretto posizionamento delle sezioni DRG sui vetrini si vede dove i tessuti non si sovrappongono alla sezione DRG e all'OCT di altre sezioni DRG.L'immagine al microscopio a fluorescenza confocale ha dimostrato la qualità delle sezioni DRG ottenute con questo protocollo. Con questa tecnica, gli studi immunoistochimici del DRG possono essere eseguiti utilizzando diversi anticorpi.

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Research

JoVE Journal

Neuroscience

Cryosectioning of the Dorsal Root Ganglion (DRG)

544 Views. Iniziare la criosezione tenendo la sezione DRG a temperatura di taglio ottimale nella parte inferiore, utilizzando un piccolo pennello, pre-raffreddato a 20 gradi Celsius. Toccare delicatamente la parte inferiore della sezione utilizzando l'estremità di una pinzetta a temperatura ambiente in modo che aderisca alla superficie della piattaforma. Quindi, posiziona lentamente un vetrino caricato sulla sezione.Non appena la sezione si attacca alla slitta, tirare delicatamente la slitta all...

Video: Cryosectioning of the Dorsal Root Ganglion (DRG)  

A Procedure for Mouse Dorsal Root Ganglion Cryosectioning

High-quality mouse dorsal root ganglion (DRG) cryostat sections are crucial for proper immunochemistry staining and RNAscope studies in the research of inflammatory and neuropathic pain, itch, as well as other peripheral neurological conditions. However, it remains a challenge to consistently obtain high-quality, intact, and flat cryostat sections onto glass slides because of the tiny sample size of the DRG tissue. So far, there is no article describing an optimal protocol for DRG cryosectioning. This protocol presents a step-by-step method to resolve the frequently encountered difficulties associated with DRG cryosectioning. The presented article explains how to remove the surrounding liquid from the DRG tissue samples, place the DRG sections on the slide facing the same orientation, and flatten the sections on the glass slide without curving up. Although this protocol has been developed for cryosectioning the DRG samples, it can be applied for the cryosectioning of many other tissues with a small sample size.

Presented here is the development for consistently acquiring high-quality dorsal root ganglion cryostat sections.

High-quality mouse dorsal root ganglion (DRG) cryostat sections are crucial for proper immunochemistry staining and RNAscope studies in the research of ...