Introduction

Traumatic brain injury (TBI) is one of the most common brain injuries caused by an external impact or pressure, such as rapid acceleration or deceleration, crushing, and projectile penetration. Following TBI, cognitive, physical, and psychosocial functions are impaired depending on the severity of the injury. To investigate TBI, several animal models of experimental traumatic brain injury have been developed. Out of these, the weight-drop injury model has been widely used to present diffused axonal injury and concussion. Weight-drop injury model provides an easy and inexpensive method for producing graded brain injuries in animals by simply dropping the weights from varying heights.

Weight drop models are relatively new techniques of TBI investigation, but the models are gaining tremendous success given their similarities to human TBI. These models could simulate the full spectrum of traumatic brain injury, ranging from mild concussion to severe TBI. Common TBI models, such as fluid percussion (FPI) and controlled cortical impact (CCI) produce focal brain contusion with little axonal injury. Whereas, weight drop models are used to reproduce diffuse brain injury (Kalish. & Whalen., 2016).

Principle

The weight-drop models use the gravitational force of the free-falling weight to generate focal or diffused brain injury. In this model, the scalp of anesthetized mice is shaved, and the periosteum is exposed by making an incision. A stainless steel helmet is fixed onto the skull with dental acrylic. This helmet distributes the kinetic energy over the brain, thereby preventing focal injury. The injury impact is delivered to the exposed skull or intact dura of the test animals. During the impact delivery, generally, silicon covered soft tips reduce the risk of skull fractures. For focal brain injury, the animals are placed on non-flexible platforms to minimize energy dissipation. In contrast, for diffused brain injury, the impact delivery is crucial as the effect is distributed over the skull and flexible platforms let the head accelerate (Albert. & Sirén., 2010).

Apparatus

The weight-drop injury device consists of a column of free-falling brass weights. Although the weights are enclosed in a Plexiglas tube, a slight left- or- right movement during its fall may potentially lateralize the impact causing uneven injury distribution. The device is equipped with an air-driven high-velocity impactor that is targeted to contact a steel disc implanted onto the rodent skull. Usually, the impactor has the same diameter as the steel disc, which is 10 mm. The top and middle surfaces of the device are made of acrylic glass and are used to hold a metal rod with a round plastic tip that penetrates to deliver the impact onto the animal’s skull. The bottom platform is constructed of iron, and a mouse’s head could be fixed on it to deliver falling weight into the targeted area of the skull (Cernak, 2005).

Protocol (Flierl. et al., 2009)

1. Anesthetize the animal.
2. Place the animal on a stereotaxic frame.
3. Inject local anesthesia under the skull and open the scalp by making a midline incision to expose the skull bone, pull out the skin to expose the skull bone.
4. Clean the skull bone by removing connective tissue and periosteum.
5. Apply the tissue adhesive to attach the disk in the preferred position while using the midline and bregma sutures as reference points.
6. Place the animal on a flexible bed and position the guide tube so that the weight hits the disk or the head of the animal.
7. Release the weight from the set height.
8. Immediately move the rat after the weight hits to prevent a second impact as the weight rebounds.
9. Remove the steel disk and suture the scalp.
10. Return the animal to a recovery cage.

Weight drop model – Surgery

Pre-operative set-up

1. Carefully check the weight-drop mechanism of the device. Lubricate the metal rod with oil to ensure smooth gliding during the weight-drop procedure.
2. Anesthetize the animals with isoflurane using a standard anesthetic machine.
3. Monitor the depth of the anesthesia by toe pinch using tweezers.
4. Weigh the animals to determine the amount of post-operative analgesics to be used.

Surgery

1. Disinfect the scalp using alcohol pads.
2. Make a longitudinal midline incision (2.0–2.5 cm) with a scalpel (blade #10).
3. Place the mouse onto the mobile platform of the weight-drop injury device. Place the device on a hard-surface bench, such as stone or marble, to minimize energy dissipation.
4. Slowly advance the tip of the rod on the exposed mouse skull to determine the exact area of impact.
5. Carefully adjust the mobile platform of the apparatus to determine the impact site.
6. Push bilateral blocks under the mobile platform to stabilize the animal’s position for impact delivery.
7. Retract the rod to the targeted position.
8. Press the pedal of the device to deliver injury. The rod will fall freely on the skull.
9. Immediately apply post-traumatic oxygen as the impact could lead to trauma-induced respiratory depression and death.

Post-operative care

1. Close the scalp by using standard suture material. The incision normally heals rapidly without wound complications.
2. Inject fentanyl (0.05 mg per kg body weight, i.p.) as post-operative analgesia. Repeat its injection every 12 hours with 0.01 mg per kg body weight i.p. for continuous analgesia.
3. Monitor initial neurological impairment per neurological severity score (NSS) 60 minutes after the injury, to assess the severity.
4. Place the animals back into the cages and kept in a temperature-controlled room with 12-hours light and dark cycles and monitor them every 6 hours.
5. Immediately return the animals to cages at the end of the surgical procedures where access to water and food are freely available.

Applications

Mimicking traumatic brain injury (Kalish. & Whalen., 2016)

Weight drop models have been widely used to advance understanding of the pathophysiology of traumatic brain injury in rodents. These models have made it possible for researchers to replicate focal cerebral contusion as well as diffused brain injury characterized by axonal damage. Recently, closed head injury models with free head rotation have also been developed to model sports concussions. The weight-drop injury model efficiently reproduces the key aspects of sports concussion for the mechanistic understanding of long-term cognitive deficits and neurological impairments.

Assessment of neuroprotective effects of Quercetin (Kom., Nageshwar, Srilatha., & Reddy., 2019)

Traumatic brain injury is followed by neuroinflammation, which could play a critical role in exasperating the progression of neurodegeneration. This study was conducted to assess the protective effects of dietary quercetin against neuroinflammation-induced changes in test subjects using the weight-drop injury model. Quercetin, a natural flavonoid, found in high quantities in edibles such as fruits is known for its anti-inflammatory, antioxidant, and free radical scavenging properties. The mice were treated for 7 days, and subsequently, behavioral studies were performed, and then their brains were collected for biochemical and histological analysis. Mice with developed neuroinflammation showed significant deficits in motor coordination, an increase in the paw withdrawal latency period, lipid peroxidation levels, as well as a decrease in antioxidant enzymes as compared to the control animals. It was observed that the quercetin treatment significantly reversed the behavioral alterations, decreased the lipid peroxidation, and increased the concentration of antioxidant enzymes, and histological alterations in the cerebral cortex. These results suggest that dietary quercetin has potential antioxidant benefits in mitigating neuroinflammation following a traumatic brain injury caused by the weight-drop injury model.

Exploring the relationship between traumatic brain injury and Alzheimer’s disease (Shishido. et al., 2019)

Traumatic brain injury (TBI) is known to cause Alzheimer’s disease (AD) later in life. It has been reported that TBI increases amyloid-β (Aβ) pathology and decreases cognitive ability in the AD model mice. This study was performed to assess the short-term and long-term effects of TBI on amyloid-β pathology and cognitive performance in weight-drop injury models. AD-related alterations and cognitive impairment following TBI were assessed in wild-type C57BL6J mice. The wild-type mice exhibited significantly decreased spatial learning as compared to the sham-treated WT mice after seven days of the injury. However, after 28 days, the cognitive impairment in the TBI-treated wild-type mice was recovered while significant accumulation of amyloid-β (Aβ) plaques and amyloid precursor protein (APP) were observed in the TBI-treated mouse hippocampus after seven days of TBI. Whereas, the Aβ deposition was no longer apparent 28 days after TBI. Therefore, it was validated that TBI induces transient amyloid-β deposition and acute cognitive impairments in mice. The results suggested that the TBI could lead to acute cognitive impairment even in the absence of genetic and hereditary predispositions. A weight-drop injury model is a useful tool for evaluating and developing a pharmacological treatment for traumatic brain injury as well as Alzheimer’s disease.

Determination of the relation between mechanical impact and neurologic dysfunction (Hsieh. et al., 2017)

Weight-drop injury model has also been used to obtain mechanistic insights about TBI. However, the relationship between the level of mechanical impact and neurological severity remained uncertain. In this study, the relationship between physical impact and graded severity was investigated at various weight-drop heights. The impact force, acceleration, and displacement during the impact were measured. Also, the longitudinal changes in cognitive deficits and balance function were monitored at 1st, 4th, and 7th days following TBI lesion. The inflammatory expression markers were also observed in the frontal cortex, hippocampus, and corpus callosum using western-blot at 1 and 7 days post-lesion. It was observed that alterations in impact pressure produced graded injuries and varying severities in the neurological score and balance function. Also, the inflammatory markers were found elevated at 1 and 7 days post-impact damage. It implied that the severity of neurologic dysfunction and excretion of inflammatory markers are strongly correlated with the graded mechanical impact levels. It was concluded that the weight-drop-induced TBI model is a useful experimental model to create graded brain injury and induce neurobehavioral deficits. This model also has translational relevance to developing therapeutic agents for TBI.

Strengths and limitations

• Weight-drop injury model efficiently induces clinically relevant behavioral outcomes representative of post-concussion symptomology. This method is rapidly applied with an extremely low mortality rate, making it suitable for therapeutic evaluation.
• As the method involves a mild injury to a closed head so that it could be easily applied for repetitive brain injury.
• The weight-drop technique is an ideal model for the study of mild traumatic brain injury and concussion as it uses a glancing impact for creating rapid rotational acceleration and deceleration to the head.
• The weight-drop injury model is inexpensive and could be carried out rapidly, allowing the examination of potential therapeutic compounds and treatment regimens for traumatic brain injury and Alzheimer’s disease.
• Weight-drop mode could lead to certain heterogeneities because of the lack of a highly precise trauma impact and the lack of animal head fixation.

References

1. A. M. Flierl., F. P. Stahel., M. K. Beauchamp., J. S. Morgan., R. W. Smith., & Shohami., E. (2009). Mouse closed head injury model induced by a weight-drop device. Nat Protoc, 4(9), 1328-37.
2. C. Albert., & Sirén., A. L. (2010). Experimental traumatic brain injury. Experimental & Translational Stroke Medicine, 2(16).
3. Cernak, I. (2005). Animal Models of Head Trauma. NeuroRX, 2(3), 410-422.
4. H. H. Kom., M. Nageshwar, K. Srilatha., & Reddy., K. P. (2019). Protective effect of quercetin on weight drop injury model-induced neuroinflammation alterations in brain of mice. Journal of Applied Pharmaceutical Science, 9(4), 96-103.
5. H. Shishido., M. Ueno., K. Sato., M. Matsumura., Y. Toyota., Y. Kirino., . . . Kishimoto., Y. (2019). Traumatic Brain Injury by Weight-Drop Method Causes Transient Amyloid-β Deposition and Acute Cognitive Deficits in Mice. Behav Neurol.
6. H. T. Hsieh., W. J. Kang., H. J. Lai., Z. Y. Huang., A. Rotenberg., Y. K. Chen., . . . Peng., W. C. (2017). Relationship of mechanical impact magnitude to neurologic dysfunction severity in a rat traumatic brain injury model. PLoS One, 12(5).
7. T. B. Kalish., & Whalen., J. M. (2016). Weight Drop Models in Traumatic Brain Injury. Methods Mol Biol, 1462, 193-209.