Preparing Rhinal Cortex-Hippocampus Slices from a Rat Pup Brain

Samenvatting

Source: Valente, C. A., et al., A Model of Epileptogenesis in Rhinal Cortex-Hippocampus Organotypic Slice Cultures. J. Vis. Exp. (2021)

The video demonstrates a method for obtaining rhinal cortex-hippocampus slices from a rat pup's brain and culturing them in membranous inserts within a multi-well plate filled with media for further analysis.

Protocol

All procedures involving animal samples have been reviewed and approved by the appropriate animal ethical review committee.

1. Preparation of rhinal cortex-hippocampus slices

NOTE: The preparation of rhinal cortex-hippocampus slices uses P6-7 Sprague-Dawley rats.

1. Culture setup and medium preparation
   1. On the day before the culture, prepare the required media and place them at 4 °C.
   2. Prepare dissection medium: 25 mM glucose in Gey's Balanced Salt Solution (GBSS).
   3. Prepare culture medium: 50% Opti-MEM, 25% HBSS, 25% Horse Serum (HS), 2....

Resultaten

Figure 1: Detailed procedure for the preparation of rhinal cortex-hippocampus organotypic slices. (A) Remove the brain from the head and place it in ice-cold GBSS with the dorsal surface faced up. (B) Insert the forceps .......

Materialen

Lijst van materialen gebruikt in dit artikel

Naam	Bedrijf	Catalogusnummer	Opmerkingen
Blades for scalpel handle	Fine Science Tools	10011-00
Brain/Tissue Slice Chamber System	Warner Instruments
Cell culture inserts, 30 mm, hydrophilic PTFE	Millipore SAS	PICM03050
Conventional incubator	Thermo Scientific Heraeus	BB15, Function Line	Set to 37 °C and 5% CO2
D(+)-Glucose monohydrate	Merck Millipore	1.08342.1000
D-(+)-Glucose solution, 45% in water	Sigma	G8769
di-Sodium hydrogen phosphate dihydrate	Merck Milipore	1.06580.1000
Dumont #5 Fine Forceps Biologie Inox	Fine Science Tools	11254-20
Dumont #5 Forceps Standard Inox	Fine Science Tools	11251-20
Dumont #7 Forceps Standard Dumoxel	Fine Science Tools	11271-30
Dumont Medical #7S Forceps Short Curve Inox	Fine Science Tools	11273-22
Gentamycin stock solution, 50 mg/mL	Thermo Fisher Scientific	15750-037
Gey's Balanced Salt Solution (GBSS)	Biological Industries	01-919-1A
Glass Electrodes	Science Products	GB150F-10	Round tips homemade
Glass Pasteur pipettes, 230 mm	VWR International	612-1702
Horse Serum, Heat Inactivated (HS)	Thermo Fisher Scientific	26050-088
Iris Spatula Curved	Fine Science Tools	10092-12
Labculture Class II Biological Safety Cabinet	HERASafe	HS 12
L-Glutamine solution 200 mM (Q)	Thermo Fisher Scientific	25030-024
Neurobasal-A Medium (NBA)	Thermo Fisher Scientific	10888-022
Opti-MEM® I Reduced-Serum Medium	Thermo Fisher Scientific	31985-047
Platinum 5 blades	Gillette
Qualitative Filter Paper, Cellulose, Grade 1, 55 mm	Whatman	1001-055	Medium retention 11µm
Qualitative Filter Paper, Cellulose, Grade 1, 90 mm	Whatman	1001-090	Medium retention 11µm
Scalpel handle	Fine Science Tools	91003-12
TC-Treated Sterile 6-Wells Plates	Corning	CORN3516
Temperatue controller	MEDICAL SYSTEMS CORP.	TC-102
Tissue Chopper	The Mickle Laboratory Engineering CO. LTD.	MTC/2	Set to 350 μm