In Vivo Imaging of Zebrafish Larval Brain for Visualizing Cerebellar Neurons

Samenvatting

Source: Schramm, P. et al. In vivo Imaging of Fully Active Brain Tissue in Awake Zebrafish Larvae and Juveniles by Skull and Skin Removal. J. Vis. Exp. (2021)

This video demonstrates the in vivo imaging of cerebellar neurons in transgenic zebrafish larvae, which express fluorescent proteins in Purkinje neurons. The process involves anesthetizing the larvae, immobilizing them in agarose, and carefully removing the pigmented skin to enhance light penetration and fluorescence intensity. Finally, the prepared specimens are imaged under a confocal microscope to visualize the labeled neurons clearly.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. Anesthesia of larvae and preparations for embedding
   1. When starting the experiment for the day, transfer the animals that are needed with a plastic Pasteur pipette to a 90 mm diameter Petri dish, which is filled with either Danieau (for larvae which are still kept in a Petri dish with Danieau) or water from the fish facility (for larvae which are older than 7 days post-fertilization [dpf] and are kept in the fish facility).

Resultaten

Figure 1: Schematic procedure for the preparation of open skull zebrafish for in.......

Materialen

Lijst van materialen gebruikt in dit artikel

Naam	Bedrijf	Catalogusnummer	Opmerkingen
Confocal Laser scanning microscope	Leica	TCS SP8
d-Tubocurare	Sigma-Aldrich	T2379-100MG
Glass Capillary type 1	WPI	1B150F-4
Glass Capillary type 2	Harvard Apparatus	GC100F-10
Glass Coverslip	deltalab	D102424
Imaging chamber	Ibidi	81156
LM-Agarose	Condalab	8050.55
Microscope Camera	Leica	DFC9000 GTC
Needle-Puller type 1	NARISHIGE	Model PC-10
Needle-Puller type 2	Sutter Instruments	Model P-2000
Pasteur-Pipettes 3ml	A.Hartenstein	20170718
Tricain	Sigma-Aldrich	E10521-50G
35 mm petri dish	Sarstedt	833900
90 mm petri dish	Sarstedt	821473001