通过研究和赞助节目的中西部高校办公室资助研究的支持。还要感谢的中西部大学动物基金和埃里克·沃伦。

注：所描述的实验方案，在中西部的大学在此进行批准了中西部大学IACUC。 1.房间要求<ol><li>安全过程的房间已运行冷热水，一种从水箱中取出的水，通常是漏在地板上。 </li><li>使用在其上放置潜水坦克的研究表。由于配备了地漏一般轮廓供排水，减少橡胶塞，并把它们放在桌子腿，保证了桌面，并在坦克，因此水面，处于水平状态。 </li></ol> 2.潜水坦克<ol><li>跳水池的施工<ol><li>构造中厚有机玻璃在有机玻璃的底部和1/2的侧壁（ 图1）用3/4的矩形槽（100×60×15厘米）。永久在一起使用氰基丙烯酸酯水泥或瑟尔连接外部的有机玻璃片罐TS，如三氯甲烷。 </li><li>通过使用可移动的1/2英寸厚的有机玻璃切成85×15厘米的块分割槽成5个信道的每个约100厘米的长度，。永久地放置在底部的槽和罐进入这些通道分频器能插位置的边缘。 </li><li>暂停从通道分频器水平的有机玻璃片，以创建一个“屋顶”的水下潜水“的隧道。 </li><li>在坦克的地方一个可移动的平台提出，“完成区”，老鼠拖拉出水面，游泳和跳水选拔赛之间休息的一个角落。老鼠通常新郎自己，并从它们的皮毛，而在终点区除去水分。放置高大两侧围绕终点区，以保证鼠保持训练试验之间的平台上的边缘。 </li><li>在从完成区域罐的相对的角放置的腔室只允许水下进入迷宫。这种可移动的“启动REA“仅用于潜水试验。 注：在潜水试验大鼠降入水中，而坐在起动室内的平台上的开头;因此，该平台被称为“电梯”。 注：使用模块化设计，其中起步区，终点区，和所有通道分频器是从水箱可拆卸。 1）以促进填充和排空水的水箱除去所有信道进行分割后，和2）如果一个鼠被迷失方向，特别是在水中，所述块可快速拆卸营救从大鼠：这是由于两个原因做水。 </li></ol></li><li>填充和排空水的水箱<ol><li>灌装水的水箱<ol><li>在每次训练的开始，注入新鲜的32℃的自来水中的罐，以约12cm的深度。搅拌该水箱中的水，以减少温暖或凉爽的口袋。 注：通常情况下，大约需要2个小时训练12只。在这期间水的温度约4℃的降低。通过培训课程结束时，水冷却至约28℃。由于30℃的水是热中性大鼠，采用这种温水，而不是常温清水，将最大限度地减少重复训练的热量损失。 </li></ol></li><li>排空水的水箱<ol><li>在每个训练段结束时，从水箱采用大软管，如4米长的一块1英寸内径管排出水。完全浸没在油箱管路，确保没有气泡在管。快速地将管插入地漏的一端。将所得的虹吸效应将沥干水从水箱。 </li><li>在保持管的罐端的水面之下，以保持虹吸，刀尖边缘的槽以促进水的完全排水。 注意：可替换地，而非虹吸，安装槽的底部附近的一个抽头，并连接从水箱软管挖掘的地漏。 </li></ol></li></ol></li></ol> 3.鼠过敏原的注意事项<ol><li>戴一次性手套操作和训练的老鼠时。的手套将限制与水箱中的水，这必然将包含鼠尿接触。 注：在培训期间的老鼠会变得潮湿，房间就会充满了“湿鼠”的气味。因此抗组胺剂预防性方案可以防止或限制老鼠过敏的影响。培训师还需要戴一次性防尘口罩或N95口罩，他们应该对大鼠过敏原敏感。 </li></ol> 4.游泳训练<ol><li>每天游泳训练 注：个人的经验表明，年轻大鼠学习迷宫更好，更快，从而开始了训练，35克刚断奶的老鼠最好用成年鼠。 <ol><li>补水箱加水。参见2.2.1。 </li><li>插入完成区域和通道分频器创建5个游泳池的通道。 </li><li>在第一堂训练课轻轻地放下老鼠手入水约3-5厘米的终点区。 注：在他们的介绍，水老鼠就会出现不稳定，因为他们第一次遇到漂浮的感觉。老鼠会划约以不协调的方式，同时试图找到一种方式来退出水，最终到达终点附近的区域。由于游泳他们与生俱来的能力，在以后的试验中老鼠会朝着终点区域更加协调地游泳。 注：从下面支持的老鼠让他们的脚都在调查人员手中。轻轻地降低大鼠入水，并让他们从手游开，而不是丢弃它们放入水中。 </li><li>试验间让大鼠保持在终点区为至少1分钟。这是为了让鼠新郎和探索，使他们把终点区为“安全”的地方试验间去了。这是相对于抓住刚刚完成了试验的老鼠，并立即定位其放回水中，开始下试。 注意：在训练过程中不要使用外部奖励（ 即食物），从而防止激活奖赏回路在大脑中。 </li><li>在终点区等待一段时间后，开始下一次开庭前轻轻按住老鼠，持续1分钟。轻轻擦干​​大鼠用毛巾，因为这种重复入水预审期间举行会导致老鼠变得非常潮湿，干燥它们将防止体温过低。 </li><li>重复4.1.3。通过4.1.5。完成3〜5次试验为每天游泳训练期间训练的每只大鼠。 </li><li>删除所有通道分频器和从油箱终点区。 </li><li>从水箱将水倒出。参见2.2.2。 </li></ol></li><li>每周游泳训练日程<ol><li>进行日常的培训课程（见4.1），每周5天，最好在同一时间的每一天。 </li><li>从开始谈判成功前一天的距离一审判决。从增加的距离开始第二和第三测试。 </li><li>与每个随后的日常训练增加，其中将大鼠置于水中和终点区之间的距离。对于最初的几个培训班由5-10厘米增加这个距离。之后，老鼠看起来更舒服游泳，30-50厘米加大距离。 </li><li>用在游泳的距离较大升幅，而大鼠学习游泳游泳道的直线部分。相比之下，学游泳的发夹通道之间变成可能需要2-3个培训班。 注：通常情况下，当等待在终点区，老鼠会重新进入水中游泳，和/或淹没他们的头在水下，同时仍坐在终点区的平台上。 </li><li>重复此日常训练方案3周以上，以保证整个5频道的成功游泳。这种重复的训练方案，确保圆满完成迷宫的，特别是因为迷宫包括交替向左和向右急转弯。 </li></ol></li></ol> 5.潜水培训注：在老鼠学会了谈判，成功地通过迷宫游泳，他们已经准备好开始跳水训练。 <ol><li>最初的潜水培训（第一天） <ol><li>将启动室。装满水的水箱（见2.2.1）。确保水位为1cm的开口到起始区域的下方。 </li><li>将完成区域和通道分频器创建5个游泳池的通道。 </li><li>在第一个潜水会话训练大鼠要在电梯入水开始腔内降低。进入迷宫是从一开始就腔室的底部。在此第一次会议中的水位低到足以使大鼠可以很容易地从一开始就室进入迷宫游泳。让老鼠在迷宫中穿行，以继续游泳在终点区。 </li><li>试验间让大鼠保持在终点区为至少1分钟。 注意：在训练过程中不要使用外部奖励（ 即食物），从而防止激活奖赏回路在大脑中。 </li><li>在终点区等待一段时间后，开始下一次开庭前轻轻按住老鼠，持续1分钟。这预审过程中保持轻柔地擦干大鼠用毛巾。 </li><li>重复5.1.3。通过5.1.5。完成3项试验的初始潜水训练期间训练的每只大鼠。 </li><li>删除所有通道分频器，开始腔和从油箱终点区。 </li><li>从水箱将水倒出。参见2.2.2。 </li></ol></li><li>最初的潜水培训（第二天） <ol><li>将启动室。装满水的水箱（见2.2.1）。确保水位稍高于开口的开始区域。 </li><li>将完成区域和通道分频器创建5个游泳池的通道。 </li><li>在第二潜水训练会话中的水位已经升高，使得退出起动室中的老鼠具有起动室进入迷宫的边缘下沾其头部。认为这是老鼠的第一次潜水。让老鼠然后通过迷宫的终点区继续游泳。 </li><li>让大鼠保持在终点区为试验之间的至少1分钟。 注意：在训练过程中不要使用外部奖励（ 即食物），从而防止激活奖赏回路在大脑中。 </li><li>在终点区等待一段时间后，开始下一次开庭前轻轻按住老鼠，持续1分钟。这预审过程中保持轻柔地擦干大鼠用毛巾。 </li><li>重复5.2.3。通过5.2.5。完成3项试验为这个潜水训练期间训练的每只大鼠。 </li><li>删除所有通道分频器，开始腔和从油箱终点区。 </li><li>从水箱将水倒出。参见2.2.2。 </li></ol></li><li>最初的潜水训练（T赫德日） <ol><li>将启动室。充满水（见2.2.1。）罐，确保水位开口到起始区的上方。 </li><li>将完成区域和通道分频器创建5个游泳池的通道。将水平件有机玻璃立刻起步区创建5厘米长的潜水隧道外。 </li><li>在第三次下潜训练的老鼠有沾它的头开始腔的下缘和游泳5厘米水下到达游泳通道的开放水域。让老鼠然后通过迷宫的终点区继续游泳。 注意：一旦放置在启动区，大鼠发起自己的水下浸没，因此这些被认为是“自愿”潜水。 </li><li>让大鼠保持在终点区为试验之间的至少1分钟。 注意：在训练过程中不要使用外部奖励（ 即食物），从而防止激活奖赏回路在大脑中。 </li><li>在终点区等待一段时间后，开始下一次开庭前轻轻按住老鼠，持续1分钟。这预审过程中保持轻柔地擦干大鼠用毛巾。 </li><li>重复5.3.3。通过5.3.5。完成3〜5个试验这个潜水训练期间训练的每只大鼠。 </li><li>删除所有通道分频器，开始腔和从油箱终点区。 </li><li>从水箱将水倒出。参见2.2.2。 </li></ol></li><li>每周潜水训练计划<ol><li>进行日常的培训课程（见5.3）每周5天，最好在每天同一时间。 </li><li>从开始谈判成功前一天的距离一审判决。开始具有增加潜水距离在第二和第三测试。 </li><li>随着每个后续的日常训练提高潜水隧道的长度增加额外的横向分隔线，延长距离的老鼠要在水下游泳。对于最初的几个培训课程增加THI设计的远程5〜10厘米。之后，老鼠看起来更舒服跳水，30-50厘米加大距离。 注意：不要透支潜水距离在连续试验尝试。如果有必要，抬起水平通道罩的末端，以提供更短的潜水距离。如果潜水试验过程中的老鼠没有达到下潜隧道的尽头，开始掉头水下，急忙扶起声道盖的末端，使老鼠浮出水面，并继续他的游泳。这将使老鼠成功地完成一个较长的潜水距离。这种积极的强化将让老鼠向前移动的海底隧道每下潜试验。 </li><li>使用潜水距离较大升幅，而大鼠学习潜水游泳道的直线部分。与此相反，学习潜水周围的发夹通道之间变成可能需要2-3个培训班。 注：通常情况下，当等待在终点区，老鼠会重新进入次辰水游泳，和/或淹没他们的头在水下，同时仍坐在终点区的平台上。 </li><li>重复此日常训练方案3周以上，以保证整个5频道的成功潜水。这种重复的训练方案，确保圆满完成迷宫的，特别是因为迷宫包括交替向左和向右急转弯。 注：全6周游泳和潜水训练计划与35克刚断奶的大鼠生长相吻合，达到体重为300g。 </li><li>经过每天的训练课程完成后，和老鼠已经回到了自己的家笼，保证只通过梳理迅速干透它们的皮毛，或者如果有必要，将加热垫笼子下保持温暖的老鼠，直到它们的皮毛干燥。 </li></ol></li></ol> 6.实验变奏曲注：基本实验装置和动物训练上面已经描述了。然而，行为训练只有PRovides能与其他实验技术用于收集感兴趣的数据模型。基本协议被修改，以调查潜水响应的特定方面。这些修饰的实例，以及一些考虑使用这些生理和神经解剖学的技术数据的收集，在下面给出。 <ol><li>植入式遥测发射机<ol><li>培训结束后，用市售的遥测发射器传送搏动性动脉血压从游泳和跳水的影响。获得本地IACUC批准关于手术和手术后的恢复过程。按照该公司为其发射器建议植入程序，并确保在返回大鼠入水之前，从手术完全恢复。 </li><li>确保天线接收到的无线电信号是在附近时，鼠是在水中。 注：水衰减无线电信号，所以距离的无线信号NE编行进通过水变得罐尺寸的限制因素。 </li><li>使用一个手持式天线棒，而不是鼠笼尺寸天线，遵循鼠因为它的进展通过迷宫。保持30厘米以内的大鼠的棒天线，以确保无线信号不会丢失，而大鼠是在水下。 </li></ol></li><li>尾随插管<ol><li>重新设计的通道分频器和横向件创造了跳水隧道的屋顶，使套管沿着小径穿过迷宫其进展过程中的老鼠的后面。 注：该通道的模块化设计是最重要的，也是必须允许快速从水下迷宫恢复大鼠。如果插管碰壁，并成为从大鼠脱落，老鼠很快就会渗出，而在水下，如果插管不能迅速重新连接。 </li><li>使用后动脉插管在自愿的潜水记录大鼠动脉血压和心脏率。用90厘米长PE50为尾随插管。 注：该套管长度足够长，以大鼠连接到压力换能器，同时仍允许大鼠通过迷宫进步，但足够短以尽量减少插管死空间和足够接近的压力换能器，以允许足够的保真度动脉压力信号。 </li><li>使用尾部静脉（或动脉）插管注入药剂，如副交感神经和交感神经激动剂和拮抗剂，或注入确定心输出量的分布的示踪剂或染料。 </li><li>抽血样本使用静脉（或动脉）插管老鼠，而老鼠是在水下以确定水下儿茶酚胺水平或血液化学。 注：最小化所述后套管的长度和占所述套管的死空间中抽血。 </li></ol></li><li>检测激活脑干神经元<ol><li>使用免疫学检测Fos蛋白来identi脑干风云特定区域，它们是潜水响应的一部分。 注：在心肺反射的重复启动，脑干神经元是反射电路可以被激活，并产生一种叫Fos蛋白的一部分。 </li><li>通过迷宫重复潜水大鼠每隔5分钟2小时，共24次下潜。 注：也诱导神经元的Fos生产其它协议也可以使用。为了避免激活参与应激反应脑干神经元的，重复的行为应该被包括在行为训练的一部分。 </li></ol></li><li>潜水过程中血皮质酮水平<ol><li>使用皮质酮作为应激大鼠的经验水平的游泳和潜水时的一个指标。 </li><li>要获得血皮质酮分析，得出0.1毫升血液样本从大鼠尾静脉15分钟后，3自愿游泳或潜水。 注：初步实验表明，15分钟是苏fficient时间，以允许用于生产和皮质酮的释放进入循环，并导致血浆皮质酮水平的峰值。由于皮质酮水平有昼夜节律，安排所有抽血在每天的同一时间和对应的培训课程的时间安排。 </li></ol></li></ol>

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笔者是一个顾问Stoelting公司，并提供了整体设计和麦卡洛克潜水坦克迷宫的规格作商业用途。

在他们的野性表格可老鼠做利用半水生环境，而且会经常潜入水下，而觅食6。因此，这是不是太奇怪，老鼠可以很容易训练自愿潜水水下。所描述的训练过程可以持续长达6周，这将带来新断奶的大鼠在大多数成年大鼠脑图谱中使用的主体大小（〜300克）。因而从这些受过训练的动物的大脑将更加容易地比得上在这些图谱确定的解剖结构。 之后被放置在起步区最老鼠?...

The diving response consists of a suite of autonomic reflexes seen in animals of all vertebrate classes 1. In response to submersion under water, this reflexly-induced response consists of apnea, bradycardia and an alteration of blood flow that maintains flow to the heart, brain and exercising muscles while limiting flow to viscera and non-exercising muscles 2. Many of the metabolic and cardiorespiratory aspects of the mammalian diving response have been well investigated 2,3, including those in humans 4,5. However, what has been relatively lacking, until recently, is investigation of the central integrative aspects of the diving response. What happens within the brainstem, and what is the neuronal step-by-step pathway, that connects afferent inputs to efferent outputs during this autonomic reflex? Answering these questions will require an appropriate animal model 6. An adage in comparative physiology, the Krogh principle 7, is that for every research question there is some animal of choice on which the problem can be most conveniently studied. A most appropriate animal for studying the central aspects of the diving response is the rat 6,8. In large part this is due to the fact that the brains of rats have been very well characterized, both anatomically and functionally, and many rat brain atlases are available 6. Additionally the rat is particularly useful in cardiorespiratory research, because the physiology of the rat is well known across all major organ systems, and the rat is well regarded as an animal model in systems biology 6. Finally, the laboratory rat is the domesticated version of the wild Rattus norvegicus, an animal that routinely swims and dives underwater 6. Based on these considerations, the rat is a good choice for studies investigating the central aspects of the mammalian diving response. In comparison, using marine animals to investigate the central aspects of the mammalian diving response would be much more difficult. This is due in large part to marine animals having comparatively large and non-uniformly sized brains, and the relative difficulty and high cost of housing these animals.
Rats have previously been used to investigate many aspects of the mammalian diving response, primarily in situations involving forced underwater submergence 9-12. However many studies in marine and aquatic animals have shown that there can be a differential response to diving based upon whether the submergence was forced or voluntary 2,13. Diving animals may show an extremely intense bradycardia during forced diving but a much less intense bradycardia during voluntary diving. The &#8220;stress&#8221; of forced submergence can significantly change the cardiovascular responses of diving in many animals 14. Small rodents such as muskrats also show a more intense diving response during forced submergence than during voluntary diving 15,16. Thus, if rats are to be used to investigate the central aspects of the mammalian diving response, investigators should be aware that a rat forcibly submerged underwater may produce a response different from that of a voluntarily diving rat. 
The goal of this article is to provide detailed instructions on how to train rats to voluntarily dive underwater. These procedures are not technically difficult, but can be time-consuming. The training is conducted in such a way as to reduce the stressfulness that could otherwise be associated with forced underwater submergence. This voluntary diving technique should minimize activation of central stress pathways and thus better allow investigation of the central aspects of the diving response. By itself, training rats to voluntarily dive underwater generates no data that can be used to investigate the central aspects of the mammalian diving response. Therefore examples of how voluntarily diving rats have been used in conjunction with other physiological and neuroanatomical research techniques, and how the basic dive training may need to be modified to accommodate theses other techniques, are also provided.

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	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Name of Material/ Equipment</td>
			<td style="width:71px;">Company</td>
			<td style="width:159px;">Catalog Number</td>
			<td style="width:200px;">Comments/Description</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">1 in internal diameter tubing&#160;</td>
			<td style="width:71px;">Fisher</td>
			<td style="width:159px;">14-169-63</td>
			<td>Used to fill or drain tank</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;">N95 mask - Moldex #2300N Series</td>
			<td style="width:71px;">Fisher</td>
			<td style="width:159px;">19-003-246D</td>
			<td style="width:200px;">Used to limit inhalation of rat allergens</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:247px;">Plexiglas rodent restraint device (Economy flat bottomed restrainer)</td>
			<td style="width:71px;">Braintree</td>
			<td style="width:159px;">FB-M/L&#160;</td>
			<td>For forced dives</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;">Telemetric transmitters&#160;</td>
			<td style="width:71px;">DSI</td>
			<td style="width:159px;">Model PA-C40 (270-0040-008)</td>
			<td style="width:200px;">Used to transmit pulsatile arterial blood pressure</td>
		</tr>
		<tr height="105">
			<td height="105" style="height:105px;">Hand-held antenna wand</td>
			<td style="width:71px;">DSI</td>
			<td style="width:159px;">Model RLA 3000 (272-5007)</td>
			<td style="width:200px;">Used to ensure radio antenna is near to transmitter while rat is negotiating underwater maze</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;">Intramedic PE50, 0.023&#34; ID</td>
			<td style="width:71px;">Fisher</td>
			<td style="width:159px;">14-170-12B</td>
			<td style="width:200px;">Used as trailing arterial cannula</td>
		</tr>
	</tbody>
</table>

training rats to voluntarily dive underwater: investigations of the mammalian diving response

Underwater submergence produces autonomic changes that are observed in virtually all diving animals. This reflexly-induced response consists of apnea, a parasympathetically-induced bradycardia and a sympathetically-induced alteration of vascular resistance that maintains blood flow to the heart, brain and exercising muscles. While many of the metabolic and cardiorespiratory aspects of the diving response have been studied in marine animals, investigations of the central integrative aspects of this brainstem reflex have been relatively lacking. Because the physiology and neuroanatomy of the rat are well characterized, the rat can be used to help ascertain the central pathways of the mammalian diving response. Detailed instructions are provided on how to train rats to swim and voluntarily dive underwater through a 5 m long Plexiglas maze. Considerations regarding tank design and procedure room requirements are also given. The behavioral training is conducted in such a way as to reduce the stressfulness that could otherwise be associated with forced underwater submergence, thus minimizing activation of central stress pathways. The training procedures are not technically difficult, but they can be time-consuming. Since behavioral training of animals can only provide a model to be used with other experimental techniques, examples of how voluntarily diving rats have been used in conjunction with other physiological and neuroanatomical research techniques, and how the basic training procedures may need to be modified to accommodate these techniques, are also provided. These experiments show that voluntarily diving rats exhibit the same cardiorespiratory changes typically seen in other diving animals. The ease with which rats can be trained to voluntarily dive underwater, and the already available data from rats collected in other neurophysiological studies, makes voluntarily diving rats a good behavioral model to be used in studies investigating the central aspects of the mammalian diving response.

圆满完成了所描述的游泳和潜水训练过程可以减少老鼠经历时，在水中潜水的压力。血液中皮质酮水平表明，重复的日常训练降低自愿跳水有关的心理压力，训练大鼠找到潜水不超过每天由一个人（ 图2; 17）正在处理压力。相反，只在跳水协议没有经过培训找到自愿跳水压力（ 图2; 17）。此外，无论是训练，训练组找到被迫跳水是最紧张（ 图2;</stro...

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Training Rats to Voluntarily Dive Underwater: Investigations of the Mammalian Diving Response

Detailed instructions are provided on how to train rats to voluntarily dive underwater through a 5 m long Plexiglas maze. Because the brains of rats have been very well characterized, voluntarily diving rats may help elucidate the central pathways of the mammalian diving response.

训练大鼠自愿潜水水下：哺乳动物潜水响应调查

Underwater submergence produces autonomic changes that are observed in virtually all diving animals. This reflexly-induced response consists of apnea, a ...

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12.1K 次观看.  Midwestern University. Detailed instructions are provided on how to train rats to voluntarily dive underwater through a 5 m long Plexiglas maze. Because the brains of rats have been very well characterized, voluntarily diving rats may help elucidate the central pathways of the mammalian diving response.