Name: testing animal anxiety in rats: effects of open arm ledges and closed arm wall transparency in elevated plus maze test
Uploaded: 2018-06-29T12:30:00
Description: Watch this Scientific Journal Video about Testing Animal Anxiety in Rats: Effects of Open Arm Ledges and Closed Arm Wall Transparency in Elevated Plus Maze Test at JoVE.com

The authors wish to thank Mr. Akihiro Hirano for his assistance in producing the customized apparatus. The authors would like to thank Mr. Kosuke Takahashi and Mr. Shingo Nakajima for their technical assistance. This work was supported by Research on Risk of Chemical Substances, Health and Laboratory Sciences Research Grants, Ministry of Health, Labour and Welfare, Japan (H25-KAGAKU-IPPANN-003) and Research Project Grant A by Institute of Science and Technology Meiji University, Japan.

All experimental procedures described here have been approved by the Institutional Animal Care and Use Committee of Meiji University (IACUC 14-0002(1)). Animals used in the representative results were prepared as follows: Male Sprague-Dawley rats were purchased from a laboratory animal company and housed in groups of 4 animals per standard rat cage (W200 &#215; D410 &#215; H250 mm) for at least one week before the start of the behavioral experiment.&#160;The animal housing room was maintained at standard ambient conditio...

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As the number of patients suffering from psychiatric disorders, including anxiety-disorder, increases, it is important to develop cutting-edge studies to improve understanding of the basic mechanism of anxiety. To achieve this, it is also important to optimize the experimental procedures. Therefore, to improve the application of the elevated plus maze test, we have demonstrated the effect of arm features on experimental outcomes using rats.
The elevated plus maze was originally established to ...

The number of patients suffering from mood disorders, including anxiety-disorder and depression, is increasing. Although the development of these disorders involves both genetic and environmental factors, the detailed pathogenic mechanisms are not yet fully understood1. Therefore, research needs to focus on uncovering the underlying mechanisms of mood disorders, as well as the development of effective drugs that have fewer side effects. 
Behavioral tests for laboratory animals are useful for investigating brain higher function, including emotion. As animal behavior is highly variable, experimental outcomes often differ between laboratories, even if an identical inbred strain was used2. Thus, researchers also need to understand the basic features of the experimental devices they adopt in order to provide reliable, precise outcomes in the laboratory.
The elevated plus maze test was originally established to examine emotional reactivity in rodents3,4. The entire apparatus is usually located 500 mm above the floor, and consists of two open and two closed arms that make a plus shape. Measurements of arm exploration (including duration and frequency of entries on each arm), distance traveled, and specified behaviors such as stretched-attending posture of subjected animals are recorded during observational periods. By these measures, animal anxiety is estimated as an experimental outcome. 
The elevated plus maze test is used throughout the world, but not all laboratories use the same apparatus design5,6, and these differences may affect the experimental outcomes. For example, differences include the presence or absence of ledges along the open arms and/or the use of transparent or opaque walls for closed arms5,6. The ledges along open arms and the use of transparent walls for closed arms were developed to prevent animals falling off the apparatus7, and to facilitate behavioral observations inside the closed arms, respectively8,9. Although there have been some studies that examined how individual arm structure differences may affect the ability to detect animal anxiety1,2,5,10,11, the combined effects of open (with/without short ledge) and closed (transparent/opaque walls) arm features on animal behavior has not yet been investigated. 
In a recent report, we discussed the effect of these combinations of arm designs on anxiety-like behavior in rats12. In brief, open arm ledges significantly increased anxiety-like behavior in rats, while transparent arms may also have decreased this behavior. Furthermore, we verified higher detection sensitivity of the effect of an anxiolytic drug in a combination of no-ledged open arm + opaque walled closed arm compared with a combination of ledged open arm + transparent walled closed arm. In this report, we will introduce our protocol for the elevated plus maze test, together with discussion of the key results from the previous report. We believe this report will provide useful information for those who have employed or who plan to use the elevated plus maze in their studies.

<table border="0" cellpadding="0" cellspacing="0" width="573">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Customized container (W1500 &#215; D1500 &#215; H2000 mm)</td>
			<td style="width:71px;">NA</td>
			<td style="width:119px;">NA</td>
			<td style="width:167px;">Handmade by Mr. Akihiro Hirano</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">LED lighting</td>
			<td style="width:71px;">NA</td>
			<td style="width:119px;">NA</td>
			<td>Handmade by Mr. Akihiro Hirano</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Elevated plus maze apparatus</td>
			<td style="width:71px;">NA</td>
			<td style="width:119px;">NA</td>
			<td>Handmade by Mr. Akihiro Hirano</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Digital video camera</td>
			<td style="width:71px;">Watec</td>
			<td style="width:119px;">WAT-250D2</td>
			<td>Any suitable video camera can be attached to a hard disk recorder for recording tracking profiles.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Hard disk recorder</td>
			<td style="width:71px;">TOSHIBA</td>
			<td style="width:119px;">RD-R200</td>
			<td>Any suitable hard disk recorder can be attached to the digital video camera.</td>
		</tr>
		<tr height="105">
			<td height="105" style="height:105px;width:216px;">Sprague-Dawley rats</td>
			<td style="width:71px;">Charles River Laboratories Japan, Inc.</td>
			<td style="width:119px;">NA</td>
			<td>Any suitable rats can be used for your study</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Standard rat cage</td>
			<td style="width:71px;">Toyoriko</td>
			<td style="width:119px;">TP-105</td>
			<td>Any suitable rat cage be used for your study</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Instrument to measure light intensity</td>
			<td style="width:71px;">AS ONE</td>
			<td style="width:119px;">1-5818-02</td>
			<td>Any suitable instrument to measure light intensity</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">EthoVision XT version 8.0</td>
			<td style="width:71px;">Noldus</td>
			<td style="width:119px;">NA</td>
			<td>Any suitable software to measure animal behavior</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">Diazepam</td>
			<td style="width:71px;">Taiyo Pharmaceutical Industry</td>
			<td style="width:119px;">1124402A2061</td>
			<td>Any suitable drug can be used for your study</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">Tween 80</td>
			<td style="width:71px;">Wako Pure Chemical Industries</td>
			<td style="width:119px;">9005-65-6</td>
			<td>Any suitable drug can be used for your study</td>
		</tr>
	</tbody>
</table>

testing animal anxiety in rats: effects of open arm ledges and closed arm wall transparency in elevated plus maze test

The elevated plus maze test is a behavioral test for assessing animal anxiety in rodents. Although this test is widely applied in the field of behavioral science, conflicting outcomes are often provided from different laboratories. To identify reasons for the different outcomes, we previously focused on arm features, which differ between laboratories, most notably the presence/absence of ledges at the sides of open arms and the transparency/opaqueness of closed arm walls. In a previous report, we used a custom designed container to compare rat behavior on different combinations of open and closed arm designs under otherwise identical experimental conditions, and showed that differences in arm features interfere with experimental outcomes. In brief, open arm ledges significantly increased anxiety-like behavior in rats, while transparent arms may also have decreased this behavior. Furthermore, we verified a higher detection sensitivity of the effect of an anxiolytic drug in a combination of no-ledged open arm + opaque walled closed arm compared with a combination of ledged open arm + transparent walled closed arm. In this report, we introduce our protocol for the elevated plus maze test, together with discussion of the key results from the previous report and our experimental experience. We believe this report will provide useful information for researchers who have employed or who plan to use the elevated plus maze in their studies.

In the previous study, we compared the combined effects of open arm ledges and transparency of closed arm walls on the exploratory behaviors on the maze12. Rats significantly spent much less time on open arms without ledges than on open arms with ledges (F[1, 60] = 13.49, P = 0.0005) (Figure 2A). Similar results were also found for the number of entries into open arms (F[1, 60] = 11.17, P &#60; 0.0014)<sup class="xref...

Watch this Scientific Journal Video about Testing Animal Anxiety in Rats: Effects of Open Arm Ledges and Closed Arm Wall Transparency in Elevated Plus Maze Test at JoVE.com

Testing Animal Anxiety in Rats: Effects of Open Arm Ledges and Closed Arm Wall Transparency in Elevated Plus Maze Test (Video) | JoVE

Rats' anxiety-like behavior tested by the elevated plus maze varies highly depending on the testing environment. Here we show how apparatus settings, especially arm structure, affect behavior on the maze, and we explain a reliable protocol to detect effects of anxiolytic drugs. This report will contribute to anxiety research.

Testing Animal Anxiety in Rats: Effects of Open Arm Ledges and Closed Arm Wall Transparency in Elevated Plus Maze Test

The elevated plus maze test is a behavioral test for assessing animal anxiety in rodents. Although this test is widely applied in the field of behavioral ...

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