Source Paper
The Monoiodoacetate Model of Osteoarthritis Pain in the Mouse
Thomas Pitcher, João Sousa-Valente, Marzia Malcangio
Journal of Visualized Experiments • 2016
Source Paper
Thomas Pitcher, João Sousa-Valente, Marzia Malcangio
Journal of Visualized Experiments • 2016
A major symptom of patients with osteoarthritis (OA) is pain that is triggered by peripheral as well as central changes within the pain pathways. The current treatments for OA pain such as NSAIDS or opiates are neither sufficiently effective nor devoid of detrimental side effects. Animal models of OA are being developed to improve our understanding of OA-related pain mechanisms and define novel pharmacological targets for therapy. Currently available models of OA in rodents include surgical and chemical interventions into one knee joint. The monoiodoacetate (MIA) model has become a standard for modelling joint disruption in OA in both rats and mice. The model, which is easier to perform in the rat, involves injection of MIA into a knee joint that induces rapid pain-like responses in the ipsilateral limb, the level of which can be controlled by injection of different doses. Intra-articular injection of MIA disrupts chondrocyte glycolysis by inhibiting glyceraldehyde-3-phosphatase dehydrogenase and results in chondrocyte death, neovascularization, subchondral bone necrosis and collapse, as well as inflammation. The morphological changes of the articular cartilage and bone disruption are reflective of some aspects of patient pathology. Along with joint damage, MIA injection induces referred mechanical sensitivity in the ipsilateral hind paw and weight bearing deficits that are measurable and quantifiable. These behavioral changes resemble some of the symptoms reported by the patient population, thereby validating the MIA injection in the knee as a useful and relevant pre-clinical model of OA pain. The aim of this article is to describe the methodology of intra-articular injections of MIA and the behavioral recordings of the associated development of hypersensitivity with a mind to highlight the necessary steps to give consistent and reliable recordings.
Objective: Measurement of weight distribution between ipsilateral and contralateral hind paws using a weight incapacitance tester to assess gait changes and pain-related weight shifting in mice following monoiodoacetate (MIA) injection
This is a Weight Bearing Deficit Assessment protocol using mouse as the model organism. The procedure involves 14 procedural steps, 9 equipment items, 11 materials. Extracted from a 2016 paper published in Journal of Visualized Experiments.
Model and subjects
mouse • Not specified • unknown • 8-10 weeks • Not specified • 8
Study window
~10 week study window | ~12 hours hands-on
Core workflow
Animal housing and acclimatization • Prepare MIA solution • Anesthetize mice
Primary readouts
Key equipment and reagents
Verified items
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House 8-10 week-old mice in groups of 5 under a 12 hr light/dark cycle (lights on at 7:00 AM) with food and water ad libitum. Allow mice to acclimatize for 1 week prior to starting the experiment.
Note: Randomize and cage mice in groups of 5. Use animal numbers as codes to blind the experimenter to treatments. Use body weights as parameters for randomization.
“House 8-10 week-old mice in groups of 5 under a 12 hr light/dark cycle (lights on at 7:00 AM) with food and water ad libitum. Let the mice acclimatize for 1 week prior to starting the experiment.”
On the day of injection, freshly prepare the solution of monoiodoacetate in sterile saline (0.9% NaCl) at the desired concentrations. The solution should be sterile filtered with a 0.22 µm filter. Prepare sterile saline for control group injections.
Note: Highest recommended dose is 1 mg in 10 µl. Monoiodoacetate is very toxic - wear gloves and mask when handling powder and preparing solution.
“On the day of injection, freshly prepare the solution of monoiodoacetate in sterile saline (0.9% NaCl) at the desired concentrations. The highest recommend dose of MIA is 1 mg in 10 µl. The solution should be sterile filtered with a 0.22 µm filter.”
Anesthetize mice using an anesthetic trolley by first placing them in a chamber delivering 2% isoflurane in O2 mixture (flow rate 1.5 L/min) and then transfer mice to the nose cone section, which also delivers the 2% isoflurane-O2 mixture to maintain anesthesia during injection. Place vet ointment on the eyes to avoid drying out.
Note: Confirm anesthesia by checking the animal's lack of response to a pinch stimulus on the hind paws.
“Anesthetize mice using an anesthetic trolley by first placing them in a chamber delivering 2% isoflurane in O2 mixture (flow rate 1.5 L/min) and then transfer mice to the nose cone section, which also delivers the 2% isoflurane-O2 mixture, and as such, maintains anesthesia during injection.”
Once the animal is under anesthesia, place it on its back. Trim and wipe the area surrounding the knee joint with alcohol. Povidone iodine or chlorhexidine can be used as alternative disinfectants. The patellar tendon (white line below the patella) will become visible.
Note: Wear surgical gown, gloves, and mask while performing injection procedure.
“Once the animal is under anesthesia, place it on its back. Trim and wipe the area surrounding the knee joint with alcohol. Povidone iodine or chlorhexidine can be used as well for disinfection. The patellar tendon (white line bellow the patella) will become visible.”
In order to stabilize the injection site, keep the knee still in a bent position by placing the index finger beneath the knee joint and the thumb above the anterior surface of the ankle joint. Joint preference is not required.
Note: Proper stabilization is critical for accurate injection.
“In order to stabilize the injection site, keep the knee still, in a bent position, by placing the index finger beneath the knee joint and the thumb above the anterior surface of the ankle joint.”
To find the precise site of injection, run a 26 G needle attached to a syringe horizontally along the knee (so as not to pierce the skin with the tip) until it finds the gap beneath the patella. Apply gentle pressure to mark the area and then lift the needle and syringe vertically for the injection.
Note: This step ensures accurate intra-articular placement.
“To find the precise site of injection, run a 26 G needle attached to a syringe horizontally along the knee (so as not to pierce the skin with the tip) until it finds the gap beneath the patella. Apply gentle pressure to mark the area and then lift the needle and syringe vertically for the injection.”
Insert the needle in the marked area, through the patellar tendon, perpendicular to the tibia. No resistance should be felt. Use thumb as a guide and inject superficial to the site of entry. Inject 10 µl of MIA solution (or saline for controls) into the knee joint.
Note: Proper technique is critical to ensure MIA enters the joint capsule and does not leak outside.
“Insert the needle in the marked area, through the patellar tendon, perpendicular to the tibia. No resistance should be felt. Use thumb as a guide and inject superficial to the site of entry.”
After injection, massage the knee to ensure even distribution of the solution. Discard the needle immediately in the sharps bin.
Note: Massage helps distribute MIA throughout the joint space.
“After injection, massage the knee to ensure even distribution of the solution. Discard the needle immediately in the sharps bin.”
Place mice back into a clean home cage on a heated mat and allow them to recover. Keep constant vigilance on the animals until they regain suitable consciousness, which is measured by them regaining sternal recumbency. Once animals are recovered, return to their cage.
Note: Continuous monitoring is essential during recovery from anesthesia.
“Place mice back into a clean home cage on a heated mat and allow them to recover. Keep constant vigilance on the animals until they regain suitable consciousness, which is measured by them regaining sternal recumbency.”
Train each mouse to walk into a Plexiglass chamber on the weight incapacitance tester apparatus and sit in the holding box. Place the mouse in front of the holding box, lift the entrance up 45°, and allow the mouse to walk in and close the box. Allow the animals to move freely until they adopt a sitting posture.
Note: This training guarantees that the animal is still and not leaning on either side of the chamber during measurement.
“Train each mouse to walk into a Plexiglass chamber on the apparatus and sit in the holding box. Place the mouse in front of the holding box, lift the entrance up 45°, and allow the mouse to walk in and close the box. This training takes at least two days and guarantees that the animal is still and not leaning on either side of the chamber.”
Calibrate the instrument before use with a 100 g check weight or according to equipment instruction. Make sure that each hind paw is placed on the appropriate recording pad.
Note: Proper calibration is essential for accurate measurements.
“Calibrate the instrument before use with a 100 g check weight (or according to equipment instruction). Make sure that each hind paw is placed on the appropriate recording pad.”
Assess weight bearing changes before MIA injection as baseline values. Collect three measurements of the weight borne on each hind paw from the recording pad for each recording session and use the mean value to calculate the difference in weight borne by ipsilateral and contralateral paws.
Note: The duration of each measurement takes 1 sec, as per the manufacturer's instructions. Express values as the difference between contralateral and ipsilateral paws in grams.
“Assess weight bearing changes before MIA injection as baseline values. The duration of each measurement takes 1 sec, as per the manufacturer's instructions. Collect three measurements of the weight borne on each hind paw from the recording pad for each recording session and use the mean value to calculate the difference in weight borne by ipsilateral and contralateral paws.”
Repeat weight bearing assessments at regular intervals over several weeks to ascertain the development of gait changes. For example, measurements can be taken on days 0, 3, 5, 7, 10, 14, 21, and 28 after MIA injection.
Note: A normal weight bearing value of 50% represents equal weight distribution across ipsilateral and contralateral hindlimb. Animals considered hypersensitive display a weight bearing change of approximately 45%.
“Then, repeat assessments at regular intervals over several weeks to ascertain the development of gate changes. For example, we report thresholds measured on 0, 3, 5, 7, 10, 14, 21, and 28 days after MIA injection.”
For best practice and training purposes, use a dye and perform immediate post-mortem dissection to confirm correct localization of injection.
Note: This step is optional but recommended for validation of injection technique.
“It is suggested for best practice and training purposes that a dye is used and immediate post-mortem dissection performed to confirm correct localization of injection.”
This section explains what the experiment is doing, which readouts matter, what the data artifacts usually look like, and how the analysis should flow from raw capture to reported result.
Measurement of weight distribution between ipsilateral and contralateral hind paws using a weight incapacitance tester to assess gait changes and pain-related weight shifting in mice following monoiodoacetate (MIA) injection
Objective
Measurement of weight distribution between ipsilateral and contralateral hind paws using a weight incapacitance tester to assess gait changes and pain-related weight shifting in mice following monoiodoacetate (MIA) injection
Subjects
From papermouse • Not specified • unknown • 8-10 weeks • Not specified
Sample count
From paper8
Cohort notes
From paperHoused in groups of 5 under 12 hr light/dark cycle (lights on at 7:00 AM) with food and water ad libitum.
Animal housing and acclimatization (1 week)
Prepare MIA solution (Not specified)
Anesthetize mice (Not specified)
Prepare injection site (Not specified)
Weight borne on ipsilateral hind paw (grams)
From paperCollect three measurements of weight borne on each hind paw for each recording session and use the mean value to calculate the difference in weight borne by ipsilateral and contralateral paws.
Artifact type
Endpoint measurements summarized by group or timepoint
Comparison focus
Compare endpoint magnitude between groups, timepoints, or both
Weight borne on contralateral hind paw (grams)
From paperCollect three measurements of weight borne on each hind paw for each recording session and use the mean value to calculate the difference in weight borne by ipsilateral and contralateral paws.
Artifact type
Endpoint measurements summarized by group or timepoint
Comparison focus
Compare endpoint magnitude between groups, timepoints, or both
Difference in weight distribution between ipsilateral and contralateral paws (grams)
From paperCollect three measurements of weight borne on each hind paw for each recording session and use the mean value to calculate the difference in weight borne by ipsilateral and contralateral paws.
Artifact type
Endpoint measurements summarized by group or timepoint
Comparison focus
Compare endpoint magnitude between groups, timepoints, or both
Percentage of weight borne on ipsilateral paw relative to total weight
From paperCollect three measurements of weight borne on each hind paw for each recording session and use the mean value to calculate the difference in weight borne by ipsilateral and contralateral paws.
Artifact type
Endpoint measurements summarized by group or timepoint
Comparison focus
Compare endpoint magnitude between groups, timepoints, or both
Weight borne on ipsilateral hind paw (grams)
From paperRaw artifact
Per-sample or per-animal endpoint measurements collected during the experiment
Processed artifact
Structured table with cleaned measurements ready for comparison
Final reported form
Summary statistics and between-group or across-timepoint comparisons
Weight borne on contralateral hind paw (grams)
From paperRaw artifact
Per-sample or per-animal endpoint measurements collected during the experiment
Processed artifact
Structured table with cleaned measurements ready for comparison
Final reported form
Summary statistics and between-group or across-timepoint comparisons
Difference in weight distribution between ipsilateral and contralateral paws (grams)
From paperRaw artifact
Per-sample or per-animal endpoint measurements collected during the experiment
Processed artifact
Structured table with cleaned measurements ready for comparison
Final reported form
Summary statistics and between-group or across-timepoint comparisons
Percentage of weight borne on ipsilateral paw relative to total weight
From paperRaw artifact
Per-sample or per-animal endpoint measurements collected during the experiment
Processed artifact
Structured table with cleaned measurements ready for comparison
Final reported form
Summary statistics and between-group or across-timepoint comparisons
Acquisition
Collect raw experimental outputs with enough metadata to preserve sample identity, condition, and timing.
Preprocessing / cleaning
Collect three measurements of weight borne on each hind paw for each recording session and use the mean value to calculate the difference in weight borne by ipsilateral and contralateral paws.
Scoring or quantification
Quantify the primary readouts for this experiment: Weight borne on ipsilateral hind paw (grams); Weight borne on contralateral hind paw (grams); Difference in weight distribution between ipsilateral and contralateral paws (grams); Percentage of weight borne on ipsilateral paw relative to total weight.
Statistical comparison
Statistical method not yet structured for this page.
Reporting output
Report representative outputs alongside summary comparisons for Weight borne on ipsilateral hind paw (grams), Weight borne on contralateral hind paw (grams), Difference in weight distribution between ipsilateral and contralateral paws (grams), Percentage of weight borne on ipsilateral paw relative to total weight.
Source links and direct wording from the methods section for validation and deeper review.
Citation
Thomas Pitcher et al. (2016). The Monoiodoacetate Model of Osteoarthritis Pain in the Mouse. Journal of Visualized Experiments
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