Funding provided by the National Institute on Alcohol Abuse and Alcoholism (NIAAA; AA-016967 and AA016967-01S1; TAM), Wayne State University start up funds, and Wayne State University Research Grant Program. The content is solely the responsibility of the authors and does not represent the official views of NIAAA or the National Institutes of Health.

1. Experimental essentials
Electrode Fabrication
<ul type="circle">
 <li>There are numerous carbon fiber microelectrodes fabrication methods since most are made in-house. Typically what dictates the electrode fabrication details is the electrochemical technique that is applied to the electrode (e.g. amperometry vs. FSCV). For FSCV, microelectrodes can be made in-house using the following three-step procedure. For a more complete description of carbon fiber electrode fabrication, see a recent JO...

<ol>
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No conflicts of interest declared.

The protocol presented here demonstrates how to prepare and use mouse coronal brain slices for FSCV experiments. Although this method is specific to obtaining and measuring dopamine dynamics, other neurotransmitters such as adenosine, hydrogen peroxide, norepinephrine, and serotonin have been monitored in vivo or in vitro with FSCV 3, 8 - 11. FSCV can be used to monitor some of these other neurochemicals by simple modifications of the waveform applied to the working electrode 3, 11...

<table border="1">
<tbody>
<tr>
<td>Name</td>
<td>Company</td>
<td>CAS number</td>
</tr>
<tr>
<td width="274">Reagent</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Potassium Chloride</td>
<td>Fisher </td>
<td width="142" valign="top">7447407</td>
</tr>
<tr>
<td width="274">Sodium chloride</td>
<td>EMD Chemicals</td>
<td width="142" valign="top">7647145</td>
</tr>
<tr>
<td width="274">Magnesium chloride</td>
<td width="194" valign="top">Fisher</td>
<td width="142" valign="top">7791186</td>
</tr>
<tr>
<td width="274">Calcium chloride</td>
<td width="194" valign="top">Fisher</td>
<td width="142" valign="top">10035048</td>
</tr>
<tr>
<td width="274">Sodium bicarbonate </td>
<td width="194" valign="top">EMD Chemicals</td>
<td width="142" valign="top">144558</td>
</tr>
<tr>
<td width="274">Sodium phosphate,Dibasic</td>
<td width="194" valign="top">EMD Chemicals</td>
<td width="142" valign="top">7558794</td>
</tr>
<tr>
<td width="274">D-glucose</td>
<td width="194" valign="top">Fisher</td>
<td width="142" valign="top">50997</td>
</tr>
<tr>
<td width="274">Ascorbic acid</td>
<td width="194" valign="top">Fisher</td>
<td width="142" valign="top">50817</td>
</tr>
<tr>
<td width="274">Sucrose</td>
<td width="194" valign="top">Fisher</td>
<td>57501</td>
</tr>
<tr>
<td width="274">&nbsp;</td>
<td width="194" valign="top">&nbsp;</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Materials</td>
<td width="194" valign="top">Vendor</td>
<td>Catalogue number </td>
</tr>
<tr>
<td width="274">Carbon fiber</td>
<td>Goodfellow Oakdale, PA</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Glass capillary </td>
<td>A-M Systems Carlsborg&nbsp; WA.</td>
<td>602000</td>
</tr>
<tr>
<td width="274">Silver wire</td>
<td>A-M Systems Carlsborg&nbsp; WA.</td>
<td>787000</td>
</tr>
<tr>
<td width="274">Tungsten stimulating electrode </td>
<td>Plastics One, Roanoke, VA </td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Platinum wire </td>
<td>&nbsp;</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Lead wire</td>
<td>Squires Electronics, Cornelius, OR</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Loctite 404 instant adhesive </td>
<td>Hankal Corp. Rocky Hill 
CT.</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Razor blade</td>
<td>World precision Instruments Inc. FL.</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">BD Spinal needle</td>
<td>BD Medical systems, 
Franlin&nbsp; Lake, NJ</td>
<td>REF 405234</td>
</tr>
<tr>
<td width="274">Surgical Blade </td>
<td>Feather&nbsp; Safety razor Co. LTD. Japan</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Software </td>
<td width="194" valign="top">Vendor</td>
<td>Catalogue number </td>
</tr>
<tr>
<td width="274">TH software</td>
<td>ESA Inc.,Chelmsford, MA</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Instrument</td>
<td width="194" valign="top">Vendor</td>
<td>Catalogue number </td>
</tr>
<tr>
<td width="274">Submersion recording chamber </td>
<td>Custom Scientific, Denver, CO </td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Neorolog stimulus isolator</td>
<td>Digitmeter, Hertfordshire, England </td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Automatic&nbsp; temperature controller</td>
<td>Warner Instrument Corporation </td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Microscope (SZX7)</td>
<td>Olympus</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Microscope </td>
<td>Fisher</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Vibratome 3000 sectioning&nbsp; system</td>
<td>St. Louis , MO.</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">Perfusion pump </td>
<td>Watson Marlow Limited, Falmouth Cornwall &ndash; England</td>
<td>H110708</td>
</tr>
<tr>
<td width="274">Micropipette puller</td>
<td>Narishige, Tokyo, Japan</td>
<td>&nbsp;</td>
</tr>
<tr>
<td width="274">ChemClamp potentiostat </td>
<td>Dagan Corporations, 
Minneapolis, MN. </td>
<td>&nbsp;</td>
</tr>
</tbody>
</table>

presynaptic dopamine dynamics in striatal brain slices with fast-scan cyclic voltammetry

Extensive research has focused on the neurotransmitter dopamine because of its importance in the mechanism of action of drugs of abuse (e.g. cocaine and amphetamine), the role it plays in psychiatric illnesses (e.g. schizophrenia and Attention Deficit Hyperactivity Disorder), and its involvement in degenerative disorders like Parkinson's and Huntington's disease. Under normal physiological conditions, dopamine is known to regulate locomotor activity, cognition, learning, emotional affect, and neuroendocrine hormone secretion. One of the largest densities of dopamine neurons is within the striatum, which can be divided in two distinct neuroanatomical regions known as the nucleus accumbens and the caudate-putamen. The objective is to illustrate a general protocol for slice fast-scan cyclic voltammetry (FSCV) within the mouse striatum. FSCV is a well-defined electrochemical technique providing the opportunity to measure dopamine release and uptake in real time in discrete brain regions. Carbon fiber microelectrodes (diameter of ~ 7 &mu;m) are used in FSCV to detect dopamine oxidation. The analytical advantage of using FSCV to detect dopamine is its enhanced temporal resolution of 100 milliseconds and spatial resolution of less than ten microns, providing complementary information to in vivo microdialysis.

Watch this Scientific Journal Video about Presynaptic Dopamine Dynamics in Striatal Brain Slices with Fast-scan Cyclic Voltammetry at JoVE.com

Presynaptic Dopamine Dynamics in Striatal Brain Slices with Fast-scan Cyclic Voltammetry

Using fast-scan cyclic voltammetry to measure electrically evoked presynaptic dopamine dynamics in striatal brain slices.

Extensive research has focused on the neurotransmitter dopamine because of its importance in the mechanism of action of drugs of abuse (e.g. cocaine and ...

presynaptic-dopamine-dynamics-striatal-brain-slices-with-fast-scan

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JoVE Journal

Neuroscience

21.7K ビュー.  Wayne State University. 線条体の脳切片に電気的に誘発シナプス前ドーパミンのダイナミクスを測定するための高速スキャンサイクリックボルタンメトリーを使用する。