Temporomandibular Joint Pain Measurement by Bite Force and Von Frey Filament Assays in Mice

Joint disorder and TMD are a group of about 30 human disease that affect joint muscle and surrounding tissue pain is the major reason why TMD patient go to hospital so far. There is no effective treatment for TMD pain due to poor understanding of the pain mechanism. To investigate the pain mechanism we have developed a mouse model of painful TMD followed by The pain behavior analysis.

Pain assessment in Artificial pain research is quite a challenge due to intrinsic variability of behavior analysis. Also, different lab use, different protocol to evaluate the pain, make it challenge to cross reference among different studies. In this JOE article we described a visualized version of one flame, filament and back force assay to major Artificial pain.

These protocols Provide detailed method for direct implementation and combined to approaches to comprehensively assess TMJ pain. They can be applied to evaluate the pain behavior in various oracal pain conditions, such as on autogenic pain, TMJ pen, muscle pen, and the neuropath pen. To begin position the anesthetized mouse on the surgical platform, prepare a sterile insulin syringe or a 31 gauge needle for injecting CFA into the temporal mandibular joint.

Cover the needle with a sheath made of polyethylene tube, leaving only two millimeters of the needle tip protruding to prevent deep insertion. Next, palpate the zygomatic arch to locate the temporal mandibular joint or TMJ injection site. Then insert the needle into the injection site, which is under the posterior end of the zygomatic arch.

Slowly inject 10 microliters of CFA into the TMJ capsule and wait for at least five seconds before gradually withdrawing the needle. To begin, establish an inflammatory TMJ pain mouse model via intraarticular injection of CFA. Once the mouse has fully recovered, place the restraining tube in the mouse cage for at least one hour in the behavioral room before testing to acclimatize the mouse to the testing environment.

Then turn on the compatible software to measure by force and open the N-B-I-T-R-S-D-V 2.6 0.3 program. Now select the appropriate communication options. Choose attended mode UDP, then select the correct local IP and search for device IP and port by clicking on search.

Once the target IP and target port appears, click open. Set the Y axis by clicking on 10 in range Y.Then click on set. Finally click on acquisition.

Next, select only channel one for the blue graph and turn off other channels not connected to the device. Click on play to start recording bite force. Now place the mouse in a 50 milliliter plastic tube modified to loosely restrain the mouse and move the bite plate toward the mouse's mouth.

Record the voluntary biting for two minutes per session. Then click on pause to stop recording and click on save. Now rank the data in the second column of the Excel file from maximum to minimum, select the top five values of bite force amplitude for further analysis.

Record the value of channel one from the program and subtract or add it to the top five values of bite force amplitude. To obtain the relative values of bite force from this assay, take the average of these five values to determine the average relative bite force for one mouse. CFA treated mice displayed a significant reduction in relative bite force on day one post injection compared to the PBS group with gradual recovery over time, bite force change percentage significantly decreased in CFA treated mice on day one with a trend toward recovery by day 12.

To begin, put a wire mesh cage in the mouse cage and habituate the mice in the behavioral room for at least one hour before the testing procedures. Then turn on the electronic von Fray test machine. To measure the head withdrawal threshold, press maximum and set the unit to 500 grams, which is an arbitrary unit set by the machine.

Set the value to zero to start recording now. Place the mice in the wire mesh cage. Apply the von fray filament perpendicularly to the temporomandibular joint area.

Record the value of force applied from the filament or the stimulus intensity in grams that triggers the reaction as the first test. Then set the value of force on the screen to zero and wait for 10 seconds while the mouse remains inside the cage. Start applying the filament again and record the value head withdrawal threshold was significantly lower in CFA treated mice on day one compared to baseline followed by partial recovery.

Over time, the percentage change in head withdrawal threshold markedly decreased in CFA treated mice on day one with improvement observed through day 12.