Home › Neuroscience › Organotypic Cerebellar Cultures: Apoptotic Challenges and Detection
Neuroscience JoVE (Open Access) Citable · DOI

Organotypic Cerebellar Cultures: Apoptotic Challenges and Detection

DOI: 10.3791/2564-v
What you'll learn
  • ✓ Generate organotypic cerebellar slice cultures from neonatal tissue
  • ✓ Apply apoptotic stimuli (Fas ligand) and assess cell viability
  • ✓ Detect and quantify apoptosis in cerebellar cell types using staining
Protocol

This method describes the generation of organotypic cerebellar cultures and the effect of certain apoptotic stimuli on the viability of different cerebellar cell types.

Difficulty
advanced
Total time
~4–5 days (culture preparation 1 day, treatment and staining 1 day, imaging/analysis 1–2 days)
Model organism
Mouse (neonatal cerebellum)
Biosafety
BSL-1

Steps

1
Prepare organotypic cerebellar slice cultures

Dissect and slice neonatal mouse cerebellum, plate on culture inserts, and maintain in vitro to generate viable organotypic tissue sections.

▶ 02:09
2
Treat slices with Fas ligand and immunostain

Apply apoptotic stimulus (Fas ligand treatment) to cerebellar cultures and perform immunohistochemical staining to visualize apoptotic markers.

▶ 05:53
3
Image and analyze stained cerebellar cultures

Acquire fluorescence microscopy images of stained cerebellar slices and assess apoptosis in different cell types (granule cells, Purkinje cells, etc.).

▶ 09:49
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