Biology Reference
In-Depth Information
2.2. Materials
and Instruments
Both rats and mice could be used before and after the animal models
of neurological disorders are produced. For beginners, early
postnatal animals (<2 weeks) are better for recording because in
comparison to the older animals the brain slices are easier to
prepare and the electrode has easier access to the cell membrane in
younger ones due to the fewer nerve fi bers in the tissue. However,
the pathology of animal models produced in early postnatal animals
is quite different from that of adults. It is recommended to use the
young adult animals (>4 weeks) for studies on animal models of
neurological disorders.
2.2.1. Animals
The following chemicals (Sigma-Aldrich, St. Louis, MO) are used
for preparing solutions: sucrose, glucose, NaCl, KCl, CaCl 2 , MgCl 2 ,
MgSO 4 , NaHCO 3 , NaH 2 PO 4 , KMeSO 4 , CsMeSO 4 , CsCl, QX314,
EGTA, and HEPES.
2.2.2. Chemicals
and Drugs
General Chemicals
The following drugs (Sigma-Aldrich) are applied in some experi-
ments to isolate currents mediated by different ion channels and
neurotransmitter receptors.
Drugs
1. Ion channel blockers: Tetrodotoxin (0.5-1.0 mM) for blocking
voltage-dependent Na + channels, CdCl 2 (0.1-0.3 mM) for
blocking voltage-dependent Ca 2+ channels, 4-aminopyridine
(2-5 mM) and tetraethylammonium (5 mM) for blocking
voltage-dependent K + channels.
2. Neurotransmitter receptor antagonists: (−)-2-amino-5-phospho-
nopentanoic acid (APV, 50 mM) for NMDA receptors, 6-cyano-
7-nitroquinoxaline-2,3-dione (CNQX, 10 mM) for AMPA receptors,
(−)-bicuculline methiodide (30 mM) for GABA A receptors.
1. Surgical tools: Including bone rongeur, forceps, scissors, blade,
and syringe (60 ml, plus 18-20-G needle).
2. Vibratome (VT1000S, Leica, Nussloch, Germany).
3. Water bath (Isotemp 105, Fisher Scientifi c).
4. Brain slice incubator (Fig. 6 ): A 250-ml beaker is used as a
container. The bottom of a petri dish (50 mm) is cut and
replaced with a piece of nylon mesh (with 0.5-mm-sized holes,
tightly stretched and glued). The dish is elevated by three plastic
legs. The brain slices are incubated on the nylon mesh. The
artifi cial cerebral spinal fl uid (ACSF) is continuously oxygenated.
Brain slices can be kept healthy up to 5 h after cutting.
2.2.3. Instruments
Brain Slice Preparation
1. Patch-clamp recording setup: Amplifi er (Axopatch 200B,
Molecular Devices), stimulator (Master 8, A.M.P.I., Jerusalem,
Israel), isolator (Iso-Flex, A.M.P.I.), computer A/D interface
(ITC-16, Instrutech, Long Island, NY), oscilloscope (V-552,
Hitachi), vibration isolated workstation (LW3048B-OPT,
Newport), micromanipulator (MC1000e-R, SD Instruments,
Whole-Cell Patch-Clamp
Recording
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