The authors would like to express their gratitude to the Figdraw scientific research drawing platform of the Home for Researchers for providing the material used to create Figure 2 in this paper, with authorized license ID: URUUO5699e. This work has received support from grants provided by the National Natural Science Foundation of China (81873380, 82074566), the Natural Science Foundation of Hubei Province (2022CFCO43), and the National Famous Traditional Chinese Medicine Heritage Studio Construction Project of the National Administration of Traditional Chinese Medicine ([2022] No.5, [2022] No.75).

All procedures conducted on the rats in this study adhered to the National Standard for Laboratory Animal Welfare and Ethics Review Guidelines (2018) officially adopted by the People&#39;s Republic of China and Guide for the Care and Use of Laboratory Animals from the National Academies Press15. This protocol is composed of three main steps. Firstly, the rat jackets are tailored and fitted based on the individual body shapes of rats with different weights. Next, acupuncture manipulation is carried out following standard procedures. Lastly, the rats are suspended in their jackets while the needles remain in place to accommodate the need for simultaneous interventions among rats within the same group.
1. Pre-needling preparation
<ol>
	<li>Production of the rat jacket
	<ol>
		<li>Weigh the rats and measure their body shape before making their jackets. To verify the suitability of this protocol for rats of varying weights, select rats weighing 220 g, 300 g, 400 g, and 520 g for independent measurements of their bodily dimensions, encompassing measurements from a to f, as depicted in Figure 2. 
		NOTE: The correlation between the bodily proportions of rats and their corresponding jacket sizes is also presented in Figure 2. The pertinent body length parameters for rats of different weights are recorded in Table 1.</li>
		<li>Cut a rectangular piece of thick, dark fabric to make a rat jacket. Use the fabric with unfolded width equal to the circumference measured around the thickest part of the rat&#39;s abdomen. The length of the fabric is equal to the distance from the rat&#39;s nose tip to the base of its tail (see Figure 2).</li>
		<li>Fold the fabric in half and cut off the top left corner at an angle of about 45&#176;-65&#176; to form a right trapezoid (see Figure 3A).</li>
		<li>Sew the two sides on the left tightly with cotton thread. Refer to Figure 1B and Figure 3B for guidance.</li>
		<li>Cut two small holes, one on each side of the fabric, for the rat&#39;s upper and lower limbs to pass through (see Figure 3C). The distance between the two upper limbs determines the spacing between the upper holes of the rat jacket. The distance between the two lower limbs determines the spacing between the lower holes of the rat jacket.</li>
		<li>Use the longitudinal distance between the rat&#39;s armpit and the front of the ilium to serve as the spacing between the upper and lower holes of the rat jacket. Use the circumference of the shoulder joint to determine the circumference of the upper hole in the rat jacket. Use the circumference of the hip joint for the circumference of the lower hole in the rat jacket.</li>
		<li>Beside each hole, sew a long cloth strap that can be crossed and wrapped tightly around the body from the opening end of the jacket to fix the rat.</li>
	</ol>
	</li>
	<li>Rat jacket-wearing method
	<ol>
		<li>Wear gloves before proceeding to dress the rat in the jacket. Begin by holding the middle and back part of the rat jacket with one hand on the table. With the other hand, hold the rat&#39;s tail from its feeding cage and gently place it into the jacket.</li>
		<li>As the rat moves forward, its head will reach the headgear of the jacket. At this point, use both hands to carefully pull its left and right upper limbs out of the holes in the jacket.</li>
		<li>Take the tying rope from the opening end of the jacket, cross it over from left to right, and wrap it securely around the rat&#39;s trunk. Tie it tightly on the opposite side of the trunk to ensure proper fixation. Following the restraint of the rat&#39;s upper limbs, repeat the same procedure to fix its lower limbs.</li>
		<li>Take precautions to avoid getting bitten or scratched by the rat&#39;s claws when pulling its limbs out of the jacket holes. Ensure that the tightness of the rope is appropriately adjusted, i.e., not too tight to cause local limb ischemia and hypoxia in rats, and not too loose, allowing the rat to escape easily (see Video 1).</li>
	</ol>
	</li>
	<li>Fixation rack and S-shaped iron hooks
	<ol>
		<li>The fixation rack consists of two heavy iron trays (24 cm in length, 12 cm in width, and 2 cm in height) and an iron column (60 cm in length) that is firmly welded on each tray. Fix a solid iron stick (145 cm in length) at the same height on the two iron columns using a screw connection structure that can be adjusted and maintained at a fixed height (see Figure 4A).</li>
		<li>For connecting the rat wearing the jacket to the iron stick, use the S-shaped hooks (11 cm in diameter). Wrap one end of the hook around the strap on the jacket while the other end is hung on the iron stick. Use two S-shaped hooks to secure the rat onto the stick. Please refer to Figure 1D and Figure 4B for the overall structure.</li>
	</ol>
	</li>
</ol>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/65652/65652fig02.jpg" /> 
Figure 2: Corresponding diagram of rat body shape and matched rat jacket size. In the diagram, the parameter a represents the transverse distance at the back of the joint between the upper limbs and the rat&#39;s trunk. This value is utilized as the spacing measurement between the penetrating holes for the upper limbs of the rat jacket. Similarly, parameter b represents the transverse distance on the back where the bilateral lower limbs of the rat connect with the trunk. This measurement determines the spacing of the penetrating holes for the lower limbs of the rat jacket. The parameter c signifies the circumference of the thickest part of the rat&#39;s abdomen, which can be utilized as the width measurement for the fabric of the rat&#39;s jacket. The parameter d represents the distance from the rat&#39;s armpit to the front of its ilium, determining the longitudinal distance between the upper and lower holes of the jacket. The parameter e corresponds to the longitudinal distance from the rat&#39;s nose tip to the base of its tail, serving as the length measurement for the rat&#39;s jacket cloth. The parameter f denotes the circumference of the articulation connecting the upper limb and the rat&#39;s trunk, serving as the circumference of the hole through which the upper limbs pass. The parameter g represents the circumference of the joint connecting the lower limbs and the rat&#39;s trunk, determining the circumference of the hole through which the lower limbs pass. <a href="https://www.jove.com/files/ftp_upload/65652/65652fig02large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td rowspan="2">Body Weight (g)</td>
			<td colspan="7">&#160;Body Shape Parameters (cm)</td>
		</tr>
		<tr>
			<td>a</td>
			<td>b</td>
			<td>c</td>
			<td>d</td>
			<td>e</td>
			<td>f</td>
			<td>g</td>
		</tr>
		<tr>
			<td>220</td>
			<td>6.0</td>
			<td>7.0</td>
			<td>15.0</td>
			<td>8.0</td>
			<td>19.0</td>
			<td>5.0</td>
			<td>7.0</td>
		</tr>
		<tr>
			<td>300</td>
			<td>7.0</td>
			<td>7.5</td>
			<td>17.0</td>
			<td>10.0</td>
			<td>21.0</td>
			<td>6.0</td>
			<td>8.0</td>
		</tr>
		<tr>
			<td>400</td>
			<td>8.0</td>
			<td>8.5</td>
			<td>19.0</td>
			<td>11.5</td>
			<td>23.0</td>
			<td>6.5</td>
			<td>9.0</td>
		</tr>
		<tr>
			<td>520</td>
			<td>8.0</td>
			<td>10.0</td>
			<td>20.0</td>
			<td>12.5</td>
			<td>26.0</td>
			<td>7.0</td>
			<td>9.5</td>
		</tr>
	</tbody>
</table>
Table 1. The measurement results of body shape parameters of rats with different body weights.
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/65652/65652fig03.jpg" /> 
Figure 3: Diagram depicting the physical appearance and usage of the rat jacket. (A) Plane exterior view of the rat jacket. The shape of the rat&#39;s jacket resembles a right-angled trapezoid, with separate openings on the left and right sides for the rat&#39;s upper and lower limbs to pass through. (B) Unfolded exterior view of the rat jacket.&#160;The closure of the headgear creates a semi-closed cavity, which serves as a crucial component to entice rats to enter. (C) Side view of a rat clothed with the jacket. Once the rat&#39;s limbs pass through the openings in the clothing, the straps are crossed over the ventral surface of the trunk and securely fastened at the back with appropriate tension. This ensures that the rat is relatively immobilized. (D) Abdominal view of a rat clothed with the rat jacket. The ventral surface of the jacket exposes the rat&#39;s abdominal skin, enabling convenient access for acupuncture at ventral points. <a href="https://www.jove.com/files/ftp_upload/65652/65652fig03large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
Video 1: Putting on the rat jacket. This video shows a typical process of dressing the rat in the jacket. <a href="https://www.jove.com/files/ftp_upload/65652/Video 1.Putting on the rat jacket.mp4" target="_blank">Please click here to download this Video.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/65652/65652fig04.jpg" /> 
Figure 4: Diagram depicting the physical appearance and usage of the elevated fixation rack. (A) Elevated fixation rack. This rack consists primarily of two vertical iron columns (60 cm in length) welded to each tray, along with a long horizontal iron stick (145 cm in length) placed parallel to the ground. Screws are used to secure the horizontal stick at the same height as the vertical columns. (B) Rat suspension on the fixation rack. Rats wearing the rat jackets are securely fastened to the fixation rack using two S-shaped hooks per rat. (C) Simultaneous electroacupuncture treatment in multiple rats. To prevent any potential interference resulting from tail movement, it was necessary to maintain proper spacing among the rats, equivalent to the length of a rat&#39;s tail. <a href="https://www.jove.com/files/ftp_upload/65652/65652fig04large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
2. Electroacupuncture procedure
<ol>
	<li>Routine preparation
	<ol>
		<li>Place the fixation rack on a horizontal desktop or ground.</li>
		<li>Prior to the needling procedure, remove the fur around the acupoints using an electric shaver to ensure complete exposure. When the acupoint to be punctured is located on the abdomen of the rat, such as Guanyuan (CV4, about 25 mm below the navel), shorten the width of rat clothing appropriately to expose abdominal acupoints.</li>
		<li>Locate the acupoint from the gap on the midline of the abdomen that is not covered by the rat jacket (see Figure 3D) and then inject the needle.</li>
		<li>When the acupoints to be punctured are on the head of the rat, such as Baihui (GV20, located at the middle of the parietal bone), make a small incision at the corresponding position on the head to fully expose the skin around GV20 (See Figure 1C).</li>
		<li>When acupuncturing acupoints on the limbs of rats, such as Neiguan (PC6, on the inner side of the forelimb, between the radial and ulnar sutures about 3 mm away from the wrist joint) in the upper limb or Zusanli (ST36, on the posterolateral side of the knee joint, about 2 mm below the fibula head) in the lower limb, due to the obvious exposure of the acupoints, use one hand to assist in fixing the limb, while the other can directly perform the acupuncture operation.</li>
		<li>When the acupoint is located on the back or waist of a rat, such as Shenshu (BL23, below the 2nd lumbar vertebrae, 6 mm apart from the midline of the back), cut an opening approximately 2 cm wide in the center of the rat&#39;s jacket to fully expose the skin of the corresponding site (See Figure 1C).</li>
		<li>Dress the rat in the jacket. After disinfecting the local skin of acupoints with alcohol swabs, begin the acupuncture procedure.</li>
	</ol>
	</li>
	<li>Needling technique: Locate the acupoint accurately. Insert a sterile disposable acupuncture needle (0.16 mm x 7 mm) into the rat&#39;s acupoint at the appropriate depth.</li>
	<li>Electroacupuncture operation
	<ol>
		<li>Wrap the electrode wire around the horizontal iron rod. This step reduces the risk of the needle falling out due to gravitational force acting on the electroacupuncture clip. Then, attach the end of the electrode wire securely onto the needle handle.</li>
		<li>Switch on the power and select the appropriate frequency, current, and time parameters. In this experiment, the parameters were set to 2 Hz, 1 mA, and 15 min. Once the parameters are adjusted, commence the electroacupuncture treatment process (See Video 2 and Video 3).</li>
		<li>Simultaneous electroacupuncture on rats: During treatment, suspend the rats in the same group together on the iron stick using the S-shaped hooks and let them undergo electroacupuncture intervention simultaneously.</li>
		<li>NOTE: The iron stick should not only be sturdy but also long enough to treat a group of rats at the same time. In this experiment, while ensuring appropriate spacing and no interference among the rats, 3-4 rats can be treated simultaneously (see Figure 4C).</li>
	</ol>
	</li>
	<li>Observation of rat responses during electroacupuncture treatment: Evaluate the suitability of this fixation method for electroacupuncture in rats based on the following factors, such as the number of personnel required to complete the operation, time taken to fix a rat, the rats&#39; response during the electroacupuncture treatment, the number of needles dropped during the procedure, and any accidental deaths that might occur during the operation.</li>
	<li>Removal of the needles
	<ol>
		<li>Once the pre-set time is completed, the electroacupuncture apparatus will automatically shut off. Remove the small clip that is connected to the needle handle and carefully remove the acupuncture needle from beneath the rat&#39;s skin, thereby concluding the electroacupuncture treatment process.</li>
		<li>Loosen the straps, lift the head cover of the rat&#39;s jacket, and position the rat above the cage. The rat will then be free to release itself from the jacket and enter the cage independently (see Video 4).</li>
	</ol>
	</li>
</ol>
Video 2. Needling acupoints in the lumbar region. This video demonstrates the comprehensive and coherent procedure encompassing the attiring of the rat in the jacket, the insertion of acupuncture needles into acupoints at the waist, and the subsequent connection of an electroacupuncture apparatus. <a href="https://www.jove.com/files/ftp_upload/65652/Video 2. Needling acupoints in lumbar region.mp4" target="_blank">Please click here to download this Video.</a>
Video 3. Needling acupoints in the abdomen and lower limb. This video demonstrates the comprehensive and coherent procedure encompassing the attiring of the rat in the jacket, the insertion of acupuncture needles into acupoints located in the abdomen and the lower limb, and the subsequent connection of an electroacupuncture apparatus. <a href="https://www.jove.com/files/ftp_upload/65652/Video 3.Needling acupoints in abdomen and lower limb.mp4" target="_blank">Please click here to download this Video.</a>
Video 4. Removing the rat jacket. This video showcases the customary practice of removing the jacket from the rat. <a href="https://www.jove.com/files/ftp_upload/65652/Video 4.Removing the rat jacket.mp4" target="_blank">Please click here to download this Video.</a>

Rat Jackets to Enhance Suspension Fixation During Electroacupuncture on Rats

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All authors have declared no potential conflicts of interest.

Electroacupuncture has been shown to have good therapeutic effects on various diseases such as cognitive impairment16,17, myocardial injury18, and inflammatory pain19. To understand its mechanism of action, animal experiments primarily using rats have been conducted. Fixation of the animals during the procedures is crucial for accurate results. While many studies have administered electroacupuncture subsequent to an...

Acupuncture is a traditional therapy characterized by needling at specific acupoints or other positions to prevent and treat diseases. Electroacupuncture combines filiform needles with electrical stimulation1, which allows for precise control over the amount of stimulation by adjusting relevant parameters, promoting the standardization of acupuncture treatment2. In recent years, research on the mechanisms of electroacupuncture has continued to expand and deepen3,4, which has led to an annual increase in studies focusing on the mechanisms of electroacupuncture5.
In basic research on electroacupuncture, rats are the most commonly used animals. During electroacupuncture treatment, the recipient must remain still to ensure a smooth procedure. Unlike in clinical trials with human subjects, small animals used in animal experiments cannot follow commands consciously during 15-30 min lengthy electroacupuncture procedures. This presents researchers with a primary challenge of achieving stable and reliable fixation. Improper fixation can lead to negative emotions such as anxiety, depression, and fear in animals6, which not only increases the risk of injury for rats and the chance of being bitten for the experimental personnel but also affects the stability and persuasiveness of research results.
In previous studies, researchers have often used anesthesia fixation7, high platform fixation8, binding fixation6,9, or rat coat fixation10 before acupuncture or electroacupuncture operations. However, given the effects of anesthesia on the body11 and brain function12, and considering that acupuncture is performed on conscious patients in clinical cases, anesthesia may impact research results to a certain extent. Therefore, compared with anesthesia fixation, comfortable fixation of rats in an awake state is a better option.
However, in the non-anesthetized state, various fixation methods also possess their respective limitations, including restricted acupoint exposure due to coverage of the rat coat on rats10, the necessity of at least two individuals to perform the operation13, non-adjustable fixator size14, or the stressed state of rats in a forced position6. To address these technical issues, we designed a rat jacket and a fixation rack suitable for acupuncture manipulation in most parts of the rat body. This can help researchers efficiently carry out acupuncture procedures with rats.
The rationale for this protocol is as follows: firstly, the closed headgear on the rat&#39;s jacket takes advantage of the rats&#39; natural inclination for darkness and burrowing, thus restricting their range of activities. Secondly, the binding ropes near the joints of the rats&#39; limbs provide physical compression restraint, imposing certain limitations on their movements. However, rats are still able to jump and move horizontally even while wearing the jacket. Lastly, since rats cannot touch the ground without a fixed force point when suspended, we added a suspension fixation device to the rat jacket, further restricting their movements and ensuring needle retention. The composition of the fixation device is shown in Figure 1.
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/65652/65652fig01.jpg" /> 
Figure 1: Schematic diagram of the rat jacket and suspension fixation. (A) The general appearance of the rat jacket in a flat view. (B) Three-dimensional view of the rat jacket. (C) Specially tailored rat jacket with an incision near the acupoints on the head and back waist. (D) Illustration of multiple rats simultaneously suspended on the elevated rack. 1: Fabric overall view; 2: Forelimb outlet; 3: Hind limb outlet; 4: Tying rope; 5: Cavity formed after suturing; 6: Incision for exposing head acupoints; 7: Incision for exposing back and waist acupoints. 8: Suture position. <a href="https://www.jove.com/files/ftp_upload/65652/65652fig01large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

<table><tbody><tr><td>75% alcohol</td><td>Shandong Lircon medical technology co., ltd</td><td>2002-0059</td><td>Disinfection</td></tr><tr><td>acupuncture needles</td><td>Beijing Luo ya shan chuan medical equipment co., Ltd.</td><td></td><td>needling</td></tr><tr><td>Adult Sprague-Dawley (SD) rats</td><td>Liaoning Changsheng biotechnology co., Ltd.</td><td>SCXK 2020-0001</td><td>The recipient&amp;nbsp; subjected to fixation</td></tr><tr><td>an elevated fixation rack&amp;nbsp;</td><td>Taizhou shiai instruments and equipments for scientific and teaching research&amp;nbsp; Co., ltd</td><td>/</td><td>Fix rats</td></tr><tr><td>Hans electronic acupuncture apparatus</td><td>Beijing Huayun Ante Science and Technology Co. Ltd.&amp;nbsp;</td><td>HANS-200A</td><td>electroacupuncture</td></tr><tr><td>Medical cotton ball</td><td>Jiangsu Misawa Medical Supplies Co., Ltd.</td><td>20172142357</td><td>Disinfection</td></tr><tr><td>Pet electric clipper</td><td>Fenghua Xikou Lubao Knives &amp;amp; Scissors Electrical Appliance Factory</td><td>/</td><td>Shave the hair of rats</td></tr><tr><td>Tweezers</td><td>Qingdao Schultz Biotechnology Co., ltd</td><td></td><td>Pinch cotton balls</td></tr></tbody></table>

improved suspension fixation using rat jacket in rat acupuncture experiments

In basic experimental acupuncture research, rats are commonly used as laboratory animals. However, it is difficult for them to maintain a fixed posture. During electroacupuncture procedures, proper immobilization of rats is essential. Various methods of rat fixation are currently used, including anesthesia fixation, high-platform fixation, binding fixation, and fixation with a self-made rat coat. However, these methods have their limitations, which may affect the efficiency and operability of the experiment to some extent. This protocol introduces a method of suspending and fixing rats using rat clothes. Firstly, rats are clothed with rat jackets that match their body shape, taking advantage of their preference for darkness and burrowing. The needling operation can then be carried out after the rats have worn rat clothes. When suspended, the rats are relatively still, as their limbs cannot move. This fixation method offers not only economical and user-friendly benefits but also ensures a stable and reliable fixation of the rats in a comfortably relaxed position. It also effectively minimizes time consumption, experimental space, and manpower resources. Additionally, this method allows for the exposure of most acupoints used for acupuncture in rats. This article primarily concentrates on the device's composition, encompassing a specially designed rat jacket, an elevated fixation rack, and their connecting structures. Additionally, an illustrative example will be presented to demonstrate the application of the rat clothing-based suspension fixation method in rat electroacupuncture procedures.

In this experiment, a total of 12 rats were selected for electroacupuncture treatment, and their reactions under the fixation method were observed (as shown in Table 2). Among the 12 rats, only 1 rat exhibited intense struggle during the 15 min electroacupuncture needle retention period. As a result of its vigorous movements, the needle came off midway, accounting for 8.33% of the total number of rats. This outcome suggests that the fixation method used generally demonstrates a notable degree of stabilit...

Watch this Scientific Journal Video about Efficient Rat Clothing for Improved Rat Fixation During Electroacupuncture at JoVE.com

Author Spotlight: Efficient Rat Clothing for Improved Rat Fixation During Electroacupuncture 

Here, we present an improved suspension fixation method using a rat jacket to effectively immobilize rats during acupuncture experiments. Furthermore, the structure and application of this fixation apparatus, as well as instructions and evaluation for its utilization in electroacupuncture therapy, is elucidated.

Improved Suspension Fixation Using Rat Jacket in Rat Acupuncture Experiments

In basic experimental acupuncture research, rats are commonly used as laboratory animals. However, it is difficult for them to maintain a fixed posture. ...

improved-suspension-fixation-using-rat-jacket-rat-acupuncture

Research

JoVE Journal

Medicine

1.3K Views.  Hubei University of Chinese Medicine. In een acupunctuurexperiment ontwikkelden we een verbeterde methode om ratten op te hangen en te fixeren met rattenkleren. Dit protocol heeft tot doel gedetailleerde beschrijvingen te geven van de procedurele stappen, inclusief belangrijke voorzorgsmaatregelen, en biedt een referentie voor onderzoekers die fundamentele experimenten uitvoeren op het gebied van acupunctuur en moxibustie. In eerder gerapporteerde onderzoeken hebben onderzoekers vaak anesthesiefixatie, hoge platformfixatie, binde...

Video: Auteur in de schijnwerpers: efficiënte rattenkleding voor een betere fixatie van ratten tijdens elektro-acupunctuur 

A Mini-Invasive Internal Fixation Technique for Studying Immobilization-Induced Knee Flexion Contracture in Rats

Joint contracture, resulting from a prolonged joint immobilization, is a common complication in orthopedics. Currently, utilizing an internal fixation to restrict knee joint mobility is a widely accepted model to generate experimental contracture. However, implanting application will inevitably cause surgical trauma to the animals. Aiming to develop a less invasive approach, we combined a muscle-gap separation modus with a previously reported mini-incision skill during the surgical procedure: Two mini skin incisions were made on the lateral thigh and leg, followed by performing muscle-gap separation to expose the bone surface. The rat knee joint was gradually immobilized by a preconstructed internal fixation at approximately 135&#176; knee flexion without interfering essential nerves or blood vessels. As expected, this simple technique permits rapid postoperative rehabilitation in animals. The correct position of the internal fixation was confirmed by an x-ray or micro-CT scanning analysis. The range of motion was significantly restricted in the immobilized knee joint than that observed in the contralateral knee joint demonstrating the effectiveness of this model. Besides, histological analysis revealed the development of fibrous deposition and adhesion in the posterior-superior knee joint capsule over time. Thus, this mini-invasive model may be suitable for mimicking the development of immobilized knee joint contracture.

Here, we present a protocol to describe a minimally invasive technique for knee joint immobilization in a rat model. This reproducible protocol, basing on muscle-gap separation modus and the mini-incision skill, is suitable for studying the underlying molecular mechanism of acquired joint contracture.

Joint contracture, resulting from a prolonged joint immobilization, is a common complication in orthopedics. Currently, utilizing an internal fixation to ...

Developmental Biology

Acrylic Resin Molding Based Head Fixation Technique in Rodents

Head fixation is a technique of immobilizing animal's head by attaching a head-post on the skull for rigid clamping. Traditional head fixation requires surgical attachment of metallic frames on the skull. The attached frames are then clamped to a stationary platform resulting in immobilization of the head. However, metallic frames for head fixation have been technically difficult to design and implement in general laboratory environment. In this study, we provide a novel head fixation method. Using a custom-made head fixation bar, head mounter is constructed during implantation surgery. After the application of acrylic resin for affixing implants such as electrodes and cannula on the skull, additional resins applied on top of that to build a mold matching to the port of the fixation bar. The molded head mounter serves as a guide rails, investigators conveniently fixate the animal&#8217;s head by inserting the head mounter into the port of the fixation bar. This method could be easily applicable if implantation surgery using dental acrylics is necessary and might be useful for laboratories that cannot easily fabricate CNC machined metal head-posts.

Traditional head fixation techniques have used metallic frames attached on the skull as an interface for head fixation apparatus. This protocol demonstrates a frameless, acrylic resin molding based head fixation technique for behavioral tasks in rodents.

Head fixation is a technique of immobilizing animal's head by attaching a head-post on the skull for rigid clamping. Traditional head fixation requires ...

Neuroscience

Rat Model of Adhesive Capsulitis of the Shoulder

This proposal aims to create an in vivo rat model of adhesive capsulitis for researching potential treatment options for this condition and other etiologies of comparable arthrofibrosis. The model includes extra-articular fixation of the shoulder in rats via scapular to humeral suturing, resulting in a secondary contracture without invading the intra-articular space and resulting in decreased rotational ROM and increased joint stiffness.
We used 10 Sprague-Dawley rats for the purpose of this study. Baseline ROM measurements were taken before glenohumeral immobilization. The rats were subjected to 8 weeks of immobilization before the fixation sutures were removed and changes in ROM and joint stiffness were evaluated. To evaluate whether immobilization resulted in a significant reduction in ROM, changes in kinematics were calculated. ROM was measured at each time point in the follow-up period and was compared to the baseline internal and external ROM measurements. In order to evaluate the stiffness, joint kinetics were calculated by determining the differences in torque (text and tint ) needed to reach the initial external rotation of 60&#176; and initial internal rotation of 80&#176;.
After the removal of the extra-articular suture fixation on follow-up day 0, we found a 63% decrease in total ROM compared to baseline. We observed continuous improvement until week 5 of follow-up, with the progress slowing down around a 19% restriction. On week 8 of follow-up, there was still an 18% restriction of ROM. Additionally, on follow-up day 0, we found the torque increased by 13.3 Nmm when compared to baseline. On week 8, the total torque was measured to be 1.4 &#177; 0.2 Nmm higher than initial measurements. This work introduces a rat model of shoulder adhesive capsulitis with lasting reduced ROM and increased stiffness.

This protocol presents an in vivo rat model of adhesive capsulitis. The model includes an internal fixation of the glenohumeral joint with extra-articular suture fixation for an extended time, resulting in a decreased rotational range of motion (ROM) and increased joint stiffness.

This proposal aims to create an in vivo rat model of adhesive capsulitis for researching potential treatment options for this condition and other ...

Bioengineering

Acupuncture in a Rat Model of Asthma

A high platform can fix rats without restriction and completely expose the acupoints on the back during acupuncture manipulation. This article describes methods for the fabrication of the high platform, establishes a rat model of asthma and measures changes in respiratory function using a noninvasive and real-time whole-body plethysmography (WBP) system.

A high platform can fix rats without restriction and completely expose the acupoints on the back during acupuncture manipulation. This article describes ...

Development of a Uterosacral Ligament Suspension Rat Model

Pelvic organ prolapse (POP) is a common pelvic floor disorder (PFD) with the potential to significantly impact a woman's quality of life. Approximately 10%-20% of women undergo pelvic floor repair surgery to treat prolapse in the United States. PFD cases result in an overall $26.3 billion annual cost in the United States alone. This multifactorial condition has a negative impact on the quality of life and yet the treatment options have only dwindled in the recent past. One common surgical option is uterosacral ligament suspension (USLS), which is typically performed by affixing the vaginal vault to the uterosacral ligament in the pelvis. This repair has a lower incidence of complications compared to those with mesh augmentation, but is notable for a relatively high failure rate of up to 40%. Considering the lack of standard animal models to study pelvic floor dysfunction, there is an urgent clinical need for innovation in this field with a focus on developing cost-effective and accessible animal models. In this manuscript, we describe a rat model of USLS involving a complete hysterectomy followed by fixation of the remaining vaginal vault to the uterosacral ligament. The goal of this model is to mimic the procedure performed on women to be able to use the model to then investigate reparative strategies that improve the mechanical integrity of the ligament attachment. Importantly, we also describe the development of an in situ tensile testing procedure to characterize interface integrity at chosen time points following surgical intervention. Overall, this model will be a useful tool for future studies that investigate treatment options for POP repair via USLS.

Pelvic organ prolapse affects millions of women worldwide and yet some common surgical interventions have failure rates as high as 40%. The lack of standard animal models to investigate this condition impedes progress. We propose the following protocol as a model for uterosacral ligament suspension and in vivo tensile testing.

Pelvic organ prolapse (POP) is a common pelvic floor disorder (PFD) with the potential to significantly impact a woman's quality of life. Approximately ...

Standardized Tuina Manipulation for Knee Osteoarthritis in Rats

Tuina Manipulation to Reduce Inflammation and Cartilage Loss in Knee Osteoarthritis Rats

Clinical trials suggest that Tuina manipulation is effective in treating knee osteoarthritis (KOA), while further studies are required to discover its mechanism. Therefore, the manipulation of animal models of knee osteoarthritis is critical. This protocol provides a standard process for Tuina manipulation on KOA rats and a preliminary exploration of the mechanism of Tuina for KOA. The press and kneading manipulation method (a kind of Tuina manipulation that refers to pressing and kneading the specific area of the body surface) is applied on 5 acupoints around the knee joint of rats. The force and frequency of the manipulation were standardized by finger pressure recordings, and the position of the rat during manipulation is described in detail in the protocol. The effect of manipulation can be measured by pain behavior tests and microscopic findings in synovial and cartilage. KOA rats showed significant improvement in pain behavior. The synovial tissue inflammatory infiltration was reduced in the Tuina group, and the expression of tumor necrosis factor (TNF)-&#945; was significantly lower. Compared to the control group, chondrocyte apoptosis was less in the Tuina group. This study provides a standardized protocol for Tuina manipulation on KOA rats and preliminary proof that the therapeutic effects of Tuina may be related to reducing synovial inflammation and delayed chondrocyte apoptosis.

Author Spotlight: Understanding the Mechanism of Tuina Manipulation in Rats 

We present a protocol for Tuina manipulation performed on plaster-immobilized-induced knee osteoarthritis rats. Based on the preliminary results, it suggested that the efficacy of the method relied on the reduction of inflammation and cartilage loss.

Clinical trials suggest that Tuina manipulation is effective in treating knee osteoarthritis (KOA), while further studies are required to discover its ...

Tuina Therapy for Frozen Shoulders in a Rat Model

Tuina in a Frozen Shoulder Rat Model: An Efficient and Reproducible Protocol

Frozen shoulder (FS) is a common condition with no defined optimal therapy. Tuina therapy, a traditional Chinese medicine (TCM) technique used to treat FS patients in Chinese hospitals, has demonstrated excellent results, but its mechanisms are not fully understood. Building on a previous study, this work aimed to develop a Tuina protocol for an FS rat model. We randomly divided 20 SD rats into control (C; n = 5), FS model (M; n = 5), FS model Tuina treatment (MT; n = 5), and FS model oral treatment (MO; n = 5) groups. This study used the cast immobilization method to establish the FS rat model. The effect of Tuina and oral dexamethasone on the glenohumeral range of motion (ROM) was evaluated, and the histological findings were assessed. Our study showed that Tuina and oral dexamethasone were able to improve shoulder active ROM and preserve the structure of the capsule, with Tuina therapy proving to be more effective than oral dexamethasone. In conclusion, the Tuina protocol established in this study was highly effective for FS.

Author Spotlight: Investigating the Effectiveness of Tuina Therapy for Frozen Shoulder 

This study develops an efficient and reproducible Tuina protocol for treating frozen shoulder established in a rat model. This approach will help to study the Tuina therapy treatment method for frozen shoulders.

Frozen shoulder (FS) is a common condition with no defined optimal therapy. Tuina therapy, a traditional Chinese medicine (TCM) technique used to treat FS ...

Therapeutic Potential of Tuina Massage for Knee Osteoarthritis

Tuina Intervention in Sodium Monoiodoacetate Injection-Induced Rat Model of Knee Osteoarthritis

Knee osteoarthritis (KOA), a common degenerative joint disorder, is characterized by chronic pain and disability, which can progress to irreparable structural damage of the joint. Investigations into the link between articular cartilage, muscles, synovium, and other tissues surrounding the knee joint in KOA are of great importance. Currently, managing KOA includes lifestyle modifications, exercise, medication, and surgical interventions; however, the elucidation of the intricate mechanisms underlying KOA-related pain is still lacking. Consequently, KOA pain remains a key clinical challenge and a therapeutic priority. Tuina has been found to have a regulatory effect on the motor, immune, and endocrine systems, prompting the exploration of whether Tuina could alleviate KOA symptoms, caused by the upregulation of inflammatory factors, and further, if the inflammatory factors in skeletal muscle can augment the progression of KOA.
We randomized 32 male Sprague Dawley (SD) rats (180-220 g) into four groups of eight animals each: antiPD-L1+Tuina (group A), model (group B), Tuina (group C), and sham surgery (group D). For groups A, B, and C, we injected 25 &#181;L of sodium monoiodoacetate (MIA) solution (4 mg MIA diluted in 25 &#181;L of sterile saline solution) into the right knee joint cavity, and for group D, the same amount of sterile physiological saline was injected. All the groups were evaluated using the least to most stressful tests (paw mechanical withdrawal threshold, paw withdrawal thermal latency, swelling of the right knee joint, Lequesne MG score, skin temperature) before injection and 2, 9, and 16 days after injection.

Author Spotlight: Treating Knee Osteoarthritis with Tuina - A New Perspective 

This protocol describes the methods of Tuina intervention in sodium monoiodoacetate injection-induced rat model of knee osteoarthritis (KOA), which provides a reference for the application of Tuina in KOA animal models. This protocol also studies the effective mechanism of Tuina for KOA, and the results will help promote its application.

Knee osteoarthritis (KOA), a common degenerative joint disorder, is characterized by chronic pain and disability, which can progress to irreparable ...

Osteoporosis Management in Rats Through Lingnan Fire-Needle Therapy

Operation Procedure and Precautions of Lingnan Fire-Needle Therapy in Osteoporosis Model Rats

Compared to filiform needle therapy, fire-needle therapy has both the stimulation of needles and the warming effect of heat, making it have unexpected effects on some chronic diseases and incurable diseases. Osteoporosis (OP) has a high incidence in postmenopausal women and middle-aged and elderly men, and the treatment cycle is long. According to Traditional Chinese Medicine (TCM), Lingnan fire-needle therapy has shown potential in treating osteoporosis. However, there is still a long way to go before it can be widely used. This article focuses on the application of Lingnan fire-needle therapy in the intervention of OP in rats. It covers the selection of needle tools, acupuncture point selection, positioning of rats' bodies, and fixation methods. We also outline the steps and precautions to be taken during and after needling with fire needles. The experiment was done with three groups: a normal group, a model group, and a fire-needle group, each containing 10 rats. The rats in the fire-needle group were treated with fire-needle intervention for six sessions. After the intervention period, we collected femoral specimens and performed micro-CT scans. The results suggest that fire needling can enhance bone morphology and mineral density in OP rats. This information can serve as a methodological basis for conducting basic research on fire-needle therapy.

Author Spotlight: Exploring the Impact of Lingnan Fire-Needle Therapy in Osteoporosis Intervention

Here, we present a protocol for the application of Lingnan Fire-Needle therapy in the intervention of osteoporosis model rats.

Compared to filiform needle therapy, fire-needle therapy has both the stimulation of needles and the warming effect of heat, making it have unexpected ...

Using a 3D Printed Holder for Bulk Electroacupuncture in Small Animals

Bulk Electroacupuncture Operation for Mice or Young Rats

Electroacupuncture (EA) is widely used to treat various health conditions. However, the underlying mechanism of EA treatment remains unclear, hindering its promotion. The mechanistic study requires mouse or rat models to address this issue. However, these animals are not obedient to the experimental process, which is time-consuming. To solve these problems, we designed a 3D-printed small animal body bulk fixator to improve the efficiency of EA's animal experiments. This video shows in detail how to use the fixator to perform bulk EA on mice or young rats. For the selection of acupoints, the anterior oblique line of vertex temporal (MS6 head) and Tianshu point (ST25 belly) were chosen to verify the effect of the fixation device with prone positioning and supine positioning. Using the 3D-printed small animal holder allows multiple rodents to be immobilized and treated simultaneously, reducing the time and resources required for the experiment. This technique could be applied to other animal models by 3D printing different sizes and could potentially be used for various fixing conditions. The device is beneficial for the promotion of experimental scientific research in EA.

Author Spotlight: Using a 3D-Printed Holder for Simultaneous Multiple Electroacupuncture Operation

Here, we present a protocol for efficient and rapid electroacupuncture (EA) in mice or young rats using a three-dimensional (3D) printed holder. This technique allows simultaneous operation on multiple animals, saving time and increasing experimental efficiency.

Electroacupuncture (EA) is widely used to treat various health conditions. However, the underlying mechanism of EA treatment remains unclear, hindering ...

Improved Suspension Fixation Using Rat Jacket in Rat Acupuncture Experiments

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