Culturing Murine Spiral Ganglion Neuron Explants on Multielectrode Arrays

Abstract

Source: Hahnewald, S., et al Spiral Ganglion Neuron Explant Culture and Electrophysiology on Multi Electrode Arrays. J. Vis. Exp. (2016)

This video demonstrates the process for isolating spiral ganglion explants from the inner ear and co-culturing them with the organ of Corti on a multielectrode array. Spiral ganglia are carefully extracted and sectioned into smaller tissue sections or explants. These explants are cultured on the multielectrode array's active area to promote neuronal process outgrowth.

Protocol

All procedures involving sample collection have been performed in accordance with the institute's IRB guidelines.

1. Prepare Solutions for Experiments

1. Prepare the extracellular matrix (ECM) coating solution (see Material Table): Thaw ECM mix on ice. Dilute the ECM mix 1:10 in basic culture medium (without neurotrophic factors or Fetal Bovine Serum (FBS)) and store on ice.
2. Prepare the culture medium (see Material Table): Prepare a stock of Neurobasal medium not containing FBS or Brain-Derived Neurotrophic Factor (BDNF). Supplement neurobasal media with fresh FBS (10%) a....

Results

Figure 1. Cell culture preparation. (A) Freshly dissected mouse inner ear. White dashed line indicates the location of the cochlea, black dashed line indicates the vestibular region. (B) Mouse inner ear after removal of the cochlear bony wall. .......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Neurobasal medium	Invitrogen	21103-049	24 ml (for 25 ml)
HEPES	Invitrogen	15630-080	250 μl (for 25 ml)
Glutamax	Invitrogen	35050-061	250 μl (for 25 ml)
B27	Invitrogen	17504-044	500 μl (for 25 ml)
FBS	GIBCO	10099-141	10% (for 25 ml)
BDNF	R&D Systems	248-BD-025/CF	Final 5 ng/ml (for 25 ml)
NaCl			145mM
KCl			4mM
MgCl2			1mM
CaCl2			2mM
HEPES			5mM
Na-pyruvate			2mM
Glucose			5mM
PBS	Invitrogen	10010023
BSA	Sigma	A4503-50G	2%
Triton X-100	Sigma	X100	0.01%
Petri dish 35 mm	Huberlab	7.627 102
Petri dish 94 mm	Huberlab	7.633 180
Dumont #5 tweezer	WPI	14098
Dumont #55 tweezer	WPI	14099
Enzymatic solution: Terg-a-Zyme	Sigma	Z273287-11KG
Extracellular Matrix (ECM) mix: Matrigel TM	Corning	356230
MEA electrodes	Qwane Biosciences		(Lausanne, Switzerland)