Effect of video angle on detection of induced front limb lameness in horses.

BACKGROUND: Lameness examinations are commonly performed in equine medicine. Advancements in digital technology have increased the use of video recordings for lameness assessment, however, standardization of ideal video angle is not available yielding videos of poor diagnostic quality. The objective of this study was to evaluate the effect of video angle on the subjective assessment of front limb lameness. A randomized, blinded, crossover study was performed. Six horses with and without mechanically induced forelimb solar pain were recorded using 9 video angles including horses trotting directly away and towards the video camera, horses trotting away and towards a video camera placed to the left and right side of midline, and horses trotting in a circle with the video camera placed on the inside and outside of the circle. Videos were randomized and assessed by three expert equine veterinarians using a 0-5 point scoring system. Objective lameness parameters were collected using a body-mounted inertial sensor system (Lameness Locator®, Equinosis LLC). Interobserver agreement for subjective lameness scores and ease of grading scores were determined. RESULTS: Induction of lameness was successful in all horses. There was excellent agreement between objective lameness parameters and subjective lameness scores (AUC of the ROC = 0.87). For horses in the "lame" trials, interobserver agreement was moderate for video angle 2 when degree of lameness was considered and perfect for video angle 2 and 9 when lameness was considered as a binary outcome. All other angles had no to fair agreement. For horses in the "sound" trials, interobserver agreement was perfect for video angle 5. All other video angles had slight to moderate agreement. CONCLUSIONS: When video assessment of forelimb lameness is required, a video of the horse trotting directly towards the video camera at a minimum is recommended. Other video angles may provide supportive information regarding lameness characteristics.

Selective plasticity of layer 2/3 inputs onto distal forelimb controlling layer 5 corticospinal neurons with skilled grasp motor training.

Layer 5 neurons of the neocortex receive their principal inputs from layer 2/3 neurons. We seek to identify the nature and extent of the plasticity of these projections with motor learning. Using optogenetic and viral intersectional tools to selectively stimulate distinct neuronal subsets in rat primary motor cortex, we simultaneously record from pairs of corticospinal neurons associated with distinct features of motor output control: distal forelimb vs. proximal forelimb. Activation of Channelrhodopsin2-expressing layer 2/3 afferents onto layer 5 in untrained animals produces greater monosynaptic excitation of neurons controlling the proximal forelimb. Following skilled grasp training, layer 2/3 inputs onto corticospinal neurons controlling the distal forelimb associated with skilled grasping become significantly stronger. Moreover, peak excitatory response amplitude nearly doubles while latency shortens, and excitatory-to-inhibitory latencies become significantly prolonged. These findings demonstrate distinct, highly segregated, and cell-specific plasticity of layer 2/3 projections during skilled grasp motor learning.

Cortical cellular encoding of thermotactile integration.

Recent evidence suggests that primary sensory cortical regions play a role in the integration of information from multiple sensory modalities. How primary cortical neurons integrate different sources of sensory information is unclear, partly because non-primary sensory input to a cortical sensory region is often weak or modulatory. To address this question, we take advantage of the robust representation of thermal (cooling) and tactile stimuli in mouse forelimb primary somatosensory cortex (fS1). Using a thermotactile detection task, we show that the perception of threshold-level cool or tactile information is enhanced when they are presented simultaneously, compared with presentation alone. To investigate the cortical cellular correlates of thermotactile integration, we performed in vivo extracellular recordings from fS1 in awake resting and anesthetized mice during unimodal and bimodal stimulation of the forepaw. Unimodal stimulation evoked thermal- or tactile- specific excitatory and inhibitory responses of fS1 neurons. The most prominent features of combined thermotactile stimulation are the recruitment of unimodally silent fS1 neurons, non-linear integration features, and response dynamics that favor longer response durations with additional spikes. Together, we identify quantitative and qualitative changes in cortical encoding that may underlie the improvement in perception of thermotactile surfaces during haptic exploration.

Reach-to-Grasp and tactile discrimination task: A new task for the study of sensory-motor learning.

Although active touch in rodents arises from the forepaws as well as whiskers, most research on active touch only focuses on whiskers. This results in a paucity of tasks designed to assess the process of active touch with a forepaw. We develop a new experimental task, the Reach-to-Grasp and Tactile Discrimination task (RGTD task), to examine active touch with a forepaw in rodents, particularly changes in processes of active touch during motor skill learning. In the RGTD task, animals are required to (1) extend their forelimb to an object, (2) grasp the object, and (3) manipulate the grasped object with the forelimb. The animals must determine the direction of the manipulation based on active touch sensations arising during the period of the grasping. In experiment 1 of the present study, we showed that rats can learn the RGTD task. In experiment 2, we confirmed that the rats are capable of reversal learning of the RGTD task. The RGTD task shared most of the reaching movements involved with conventional forelimb reaching tasks. From the standpoint of a discrimination task, the RGTD task enables rigorous experimental control, for example by removing bias in the stimulus-response correspondence, and makes it possible to utilize diverse experimental procedures that have been difficult in prior tasks.

A modified rehabilitation paradigm bilaterally increased rat extensor digitorum communis muscle size but did not improve forelimb function after stroke.

Malnutrition after stroke may lessen the beneficial effects of rehabilitation on motor recovery through influences on both brain and skeletal muscle. Enriched rehabilitation (ER), a combination of environmental enrichment and forelimb reaching practice, is used preclinically to study recovery of skilled reaching after stroke. However, the chronic food restriction typically used to motivate engagement in reaching practice is a barrier to using ER to investigate interactions between nutritional status and rehabilitation. Thus, our objectives were to determine if a modified ER program comprised of environmental enrichment and skilled reaching practice motivated by a short fast would enhance post-stroke forelimb motor recovery and preserve forelimb muscle size and metabolic fiber type, relative to a group exposed to stroke without ER. At one week after photothrombotic cortical stroke, male, Sprague-Dawley rats were assigned to modified ER or standard care for 2 weeks. Forelimb recovery was assessed in the Montoya staircase and cylinder task before stroke and on days 5-6, 22-23, and 33-34 after stroke. ER failed to improve forelimb function in either task (p > 0.05). Atrophy of extensor digitorum communis (EDC) and triceps brachii long head (TBL) muscles was not evident in the stroke-targeted forelimb on day 35, but the area occupied by hybrid fibers was increased in the EDC muscle (p = 0.038). ER bilaterally increased EDC (p = 0.046), but not TBL, muscle size; EDC muscle fiber type was unchanged by ER. While the modified ER did not promote forelimb motor recovery, it does appear to have utility for studying the role of skeletal muscle plasticity in post-stroke recovery.

Recording Forelimb Muscle Activity in Head-Fixed Mice with Chronically Implanted EMG Electrodes.

Powerful genetic and molecular tools available in mouse systems neuroscience research have enabled researchers to interrogate motor system function with unprecedented precision in head-fixed mice performing a variety of tasks. The small size of the mouse makes the measurement of motor output difficult, as the traditional method of electromyographic (EMG) recording of muscle activity was designed for larger animals like cats and primates. Pending commercially available EMG electrodes for mice, the current gold-standard method for recording muscle activity in mice is to make electrode sets in-house. This article describes a refinement of established procedures for hand fabrication of an electrode set, implantation of electrodes in the same surgery as headplate implantation, fixation of a connector on the headplate, and post-operative recovery care. Following recovery, millisecond-resolution EMG recordings can be obtained during head-fixed behavior for several weeks without noticeable changes in signal quality. These recordings enable precise measurement of forelimb muscle activity alongside in vivo neural recording and/or perturbation to probe mechanisms of motor control in mice.

Morphological and radiographic studies on the Manus region in the Arabian one-humped camel (Camelus dromedaries).

The study aims to analyse the normal anatomical and radiographical features of the Manus of the southern Aswanian-adapted Arabian one-humped camel, providing crucial data for diagnosing and treating various ailments. Our study was applied to 10 cadaver forelimbs of adult male one-humped camels (4-5 years old) for an explanation of the gross anatomy of the bones of the Manus region from under the carpal bones by using traditional techniques, including the gross anatomical, radiographic and x-ray (at the dorsopalmar and lateral planes) of the preparation of Manus bones. Our results showed that the large fused (third and fourth) metacarpal bones, in which the fusion extended along the entire length of the bone except at the distal end, diverged to form separate articulations with cross-ponding digits. As described in all ruminant species, especially the camel, there were two digits, and each digit consisted of three phalanges and two proximal sesamoid bones. Our radiographic x-ray data revealed that the complete radiopaque septum that completely divided the medullary cavity into two separate parts was clear from the dorsopalmar view, while the lateral view showed the proximal sesamoid bones that were placed over each other and located palmar to the head of the large metacarpal bone. In conclusion, our study reveals the adaptations of the Arabian one-humped camel to Egyptian conditions, aiding in the early diagnosis of lameness and digit problems and enabling veterinarians and camel owners to better address these issues, thereby improving the overall health and well-being of these animals.

Gathering Self-Initiated Rat Behavioral Data to Characterize Post-Stroke Deficits.

Behavioral testing in rat models is frequently utilized for diverse purposes, including psychological, biomedical, and behavioral research. Many traditional approaches involve individual, one-on-one testing sessions between a single researcher and each animal in an experiment. This setup can be very time consuming for the researcher, and their presence may impact the behavioral data in unwanted ways. Additionally, traditional caging for rat research imposes a lack of enrichment, exercise, and socialization that would normally be typical for the species, and this context may also skew the results of behavioral data. Overcoming these limitations may be worthwhile for several research applications, including the study of acquired brain injury. Here, an example method is presented for automatically training and testing individual rat behavior in a colony cage without the presence of humans. Radio frequency identification can be utilized to tailor sessions to the individual rat. The validation of this system occurred in the example context of measuring skilled forelimb motor performance before and after stroke. Traditional characteristics of post-stroke behavioral impairments and novel measures enabled by the system are measured, including success rate, various aspects of pull force, bout analysis, initiation rate and patterns, session duration, and circadian patterns. These variables can be collected automatically with few limitations; though the apparatus removes experimental control of exposure, timing and practice, the validation produced reasonable consistency in these variables from animal to animal.

Finite element study on post-screw removal stress in toy poodle radius with different plate designs and screw arrangements.

Background: The study employs finite element analysis to investigate stress distribution in the radius of toy poodles after screw removal. The examination focuses on the biomechanical implications of varied screw hole configurations using 1.5 and 2.0-mm locking compression plates (LCPs) with notched head T-Plates. Aim: To provide a noninvasive approach to analyzing the immediate consequences of screw removal from the radius bone in toy poodles. Specifically, it explores the impact of varied plate designs and screw arrangements on stress distribution within the forelimb bones. Methods: The study constructs a three-dimensional bone model of the toy poodle's forelimb based on computed tomography (CT) images. Simulations were designed to replicate jumping and landing from a 40 cm height, comparing stress distribution in the radius post-screw removal. Results: The analysis reveals significant variations in stress distribution patterns between the two LCPs. The radius implanted with the 2.0-mm LCP displays a uniform stress distribution, contrasting with the 1.5-mm plates. Localized stress concentration is observed around the screw holes, while trabecular bone regions near the screw holes exhibit lower stress levels. Conclusion: The study highlights the plate designs and screw configurations that affect bone stress in toy poodle forelimbs post-screw removal. The findings provide valuable insights for veterinarians, aiding informed decisions in veterinary orthopedic practices.

Changes in intra- and interlimb reflexes from hindlimb cutaneous afferents after staggered thoracic lateral hemisections during locomotion in cats.

When the foot dorsum contacts an obstacle during locomotion, cutaneous afferents signal central circuits to coordinate muscle activity in the four limbs. Spinal cord injury disrupts these interactions, impairing balance and interlimb coordination. We evoked cutaneous reflexes by electrically stimulating left and right superficial peroneal nerves before and after two thoracic lateral hemisections placed on opposite sides of the cord at 9- to 13-week interval in seven adult cats (4 males and 3 females). We recorded reflex responses in ten hindlimb and five forelimb muscles bilaterally. After the first (right T5-T6) and second (left T10-T11) hemisections, coordination of the fore- and hindlimbs was altered and/or became less consistent. After the second hemisection, cats required balance assistance to perform quadrupedal locomotion. Short-latency reflex responses in homonymous and crossed hindlimb muscles largely remained unaffected after staggered hemisections. However, mid- and long-latency homonymous and crossed responses in both hindlimbs occurred less frequently after staggered hemisections. In forelimb muscles, homolateral and diagonal mid- and long-latency response occurrence significantly decreased after the first and second hemisections. In all four limbs, however, when present, short-, mid- and long-latency responses maintained their phase-dependent modulation. We also observed reduced durations of short-latency inhibitory homonymous responses in left hindlimb extensors early after the first hemisection and delayed short-latency responses in the right ipsilesional hindlimb after the first hemisection. Therefore, changes in cutaneous reflex responses correlated with impaired balance/stability and interlimb coordination during locomotion after spinal cord injury. Restoring reflex transmission could be used as a biomarker to facilitate locomotor recovery. KEY POINTS: Cutaneous afferent inputs coordinate muscle activity in the four limbs during locomotion when the foot dorsum contacts an obstacle. Thoracic spinal cord injury disrupts communication between spinal locomotor centres located at cervical and lumbar levels, impairing balance and limb coordination. We investigated cutaneous reflexes during quadrupedal locomotion by electrically stimulating the superficial peroneal nerve bilaterally, before and after staggered lateral thoracic hemisections of the spinal cord in cats. We showed a loss/reduction of mid- and long-latency responses in all four limbs after staggered hemisections, which correlated with altered coordination of the fore- and hindlimbs and impaired balance. Targeting cutaneous reflex pathways projecting to the four limbs could help develop therapeutic approaches aimed at restoring transmission in ascending and descending spinal pathways.

Patterns of integrated growth of body parts in Rook (Corvus frugilegus) ontogeny.

The early period of ontogeny is key to understanding the patterns of body plan formation in birds. Most studies of avian development have focused on the development of individual avian characters, leaving their developmental integration understudied. We explored the dynamics and integration of relative percentage increments in body mass, lengths of head, skeletal elements of wing and leg, and primary flight feathers in the embryonic and postnatal development of the Rook (Corvus frugilegus). The relative percentage increments were calculated according to Brody's equation. Groups of similar growing traits (modules) were determined using hierarchical cluster analysis, and the degree of correlation between modules was estimated by PLS analysis. The embryonic and postnatal periods demonstrate significant consistency both in the dynamics of changes in relative percentage increments of studied traits as well as in the clustering of individual modules. The modules mainly include the body mass and head length, as well as the elements that form the fore- and hind limbs. Differences were revealed in the combination of modules into clusters in embryonic and postnatal periods. Hind limb elements clustered together with wing elements in the embryonic period but with body mass and the head in the postnatal period. The strongest modularity was noted for the leg in embryogenesis, and for the wing in postnatal development. The forelimb and especially the primary feathers had more distinctive growth patterns. We suggest the changes in the degree of integration between locomotor modules in ontogenesis are connected with the earlier functioning of the legs in the postnatal period and with the preparation of the wings for functioning after a chick leaves the nest.

The effects of a synthetic epidermis spray on secondary intention wound healing in adult horses.

OBJECTIVES: To evaluate secondary intention wound healing in the horse's limbs when treated with the synthetic epidermis spray (Novacika®, Cohesive S.A.S, France) or with a standard bandaging technique. METHODS: Six Standardbred mares were included in the study. Four 2.5 x 2.5 cm full-thickness skin wounds were created on each thoracic limb. Two wounds were located on the dorsoproximal aspect of the cannon bone and the other two at the dorsoproximal aspect of the fetlock. Six hours after creation, wounds were randomly treated with synthetic epidermis spray or standard bandaging. The wounds were assessed every 4 days by gross visual assessment and using a 3D imaging camera. Analysis was performed with a 3D imaging application. RESULTS: Out of 46 wounds, 22 showed exuberant granulation tissue and were part of the standard bandaging group. Whether the wounds were treated with synthetic epidermis spray or standard bandaging, the time for healing was the same. CONCLUSION: The synthetic epidermis spray studied in this model has allowed healing without the production of exuberant granulation tissue but did not reduce the median wound healing time compared to a standard bandaging technique. The synthetic epidermis spray is potentially an interesting alternative for the management of secondary intention wound healing of superficial and non-infected distal limb wounds in adult horses on economical and practical aspects. However, all statistical inference (p-values especially) must be interpreted with caution, given the size of the sample.

Exploration biases forelimb reaching strategies.

The brain can generate actions, such as reaching to a target, using different movement strategies. We investigate how such strategies are learned in a task where perched head-fixed mice learn to reach to an invisible target area from a set start position using a joystick. This can be achieved by learning to move in a specific direction or to a specific endpoint location. As mice learn to reach the target, they refine their variable joystick trajectories into controlled reaches, which depend on the sensorimotor cortex. We show that individual mice learned strategies biased to either direction- or endpoint-based movements. This endpoint/direction bias correlates with spatial directional variability with which the workspace was explored during training. Model-free reinforcement learning agents can generate both strategies with similar correlation between variability during training and learning bias. These results provide evidence that reinforcement of individual exploratory behavior during training biases the reaching strategies that mice learn.

Topographical and cell type-specific connectivity of rostral and caudal forelimb corticospinal neuron populations.

Corticospinal neurons (CSNs) synapse directly on spinal neurons, a diverse assortment of cells with unique structural and functional properties necessary for body movements. CSNs modulating forelimb behavior fractionate into caudal forelimb area (CFA) and rostral forelimb area (RFA) motor cortical populations. Despite their prominence, the full diversity of spinal neurons targeted by CFA and RFA CSNs is uncharted. Here, we use anatomical and RNA sequencing methods to show that CSNs synapse onto a remarkably selective group of spinal cell types, favoring inhibitory populations that regulate motoneuron activity and gate sensory feedback. CFA and RFA CSNs target similar spinal neuron types, with notable exceptions that suggest that these populations differ in how they influence behavior. Finally, axon collaterals of CFA and RFA CSNs target similar brain regions yet receive highly divergent inputs. These results detail the rules of CSN connectivity throughout the brain and spinal cord for two regions critical for forelimb behavior.

Transformation of the pectoral girdle in pennaraptorans: critical steps in the formation of the modern avian shoulder joint.

Important transformations of the pectoral girdle are related to the appearance of flight capabilities in the Dinosauria. Previous studies on this topic focused mainly on paravians yet recent data suggests flight evolved in dinosaurs several times, including at least once among non-avialan paravians. Thus, to fully explore the evolution of flight-related avian shoulder girdle characteristics, it is necessary to compare morphology more broadly. Here, we present information from pennaraptoran specimens preserving pectoral girdle elements, including all purportedly volant taxa, and extensively compare aspects of the shoulder joint. The results show that many pectoral girdle modifications appear during the evolution from basal pennaraptorans to paravians, including changes in the orientation of the coracoid body and the location of the articulation between the furcula and scapula. These modifications suggest a change in forelimb range of motion preceded the origin of flight in paravians. During the evolution of early avialans, additional flight adaptive transformations occur, such as the separation of the scapula and coracoid and reduction of the articular surface between these two bones, reduction in the angle between these two elements, and elongation of the coracoid. The diversity of coracoid morphologies and types of articulations joining the scapula-coracoid suggest that each early avialan lineage evolved these features in parallel as they independently evolved more refined flight capabilities. In early ornithothoracines, the orientation of the glenoid fossa and location of the acrocoracoid approaches the condition in extant birds, suggesting a greater range of motion in the flight stroke, which may represent the acquisition of improved powered flight capabilities, such as ground take-off. The formation of a new articulation between the coracoid and furcula in the Ornithuromorpha is the last step in the formation of an osseous triosseal canal, which may indicate the complete acquisition of the modern flight apparatus. These morphological transitions equipped birds with a greater range of motion, increased and more efficient muscular output and while at the same time transmitting the increased pressure being generated by ever more powerful flapping movements in such a way as to protect the organs. The driving factors and functional adaptations of many of these transitional morphologies are as yet unclear although ontogenetic transitions in forelimb function observed in extant birds provide an excellent framework through which we can explore the behavior of Mesozoic pennaraptorans.

Alternate Day Fasting Leads to Improved Post-Stroke Motor Recovery in Mice.

BACKGROUND: Caloric restriction promotes neuroplasticity and recovery after neurological injury. In mice, we tested the hypothesis that caloric restriction can act post-stroke to enhance training-associated motor recovery. METHODS: Mice were trained to perform a skilled prehension task. We then induced a photothrombotic stroke in the caudal forelimb area, after which we retrained animals on the prehension task following an 8-day delay. Mice underwent either ad libitum feeding or alternate day fasting beginning 1-day after stroke and persisting for either 7 days or the entire post-stroke training period until sacrifice. RESULTS: Prior studies have shown that post-stroke recovery of prehension can occur if animals receive rehabilitative training during an early sensitive period but is incomplete if rehabilitative training is delayed. In contrast, we show complete recovery of prehension, despite a delay in rehabilitative training, when mice underwent alternate day fasting beginning 1-day post-stroke and persisting for either 7 days or the entire post-stroke training period until sacrifice. Recovery was independent of weight loss. Stroke volumes were similar across groups. CONCLUSIONS: Post-stroke caloric restriction led to recovery of motor function independent of a protective effect on stroke volume. Prehension recovery improved even after ad libitum feeding was reinstituted suggesting that the observed motor recovery was not merely a motivational response. These data add to the growing evidence that post-stroke caloric restriction can enhance recovery.

Dorsal-to-palmar branch neuroanastomosis in horses undergoing palmar digital neurectomy does not reduce neuroma formation or improve outcome.

OBJECTIVE: Chronic foot pain, a common cause of forelimb lameness, can be treated by palmar digital neurectomy (PDN). Complications include neuroma formation and lameness recurrence. In humans, neuroanastomoses are performed to prevent neuroma formation. The aim of the study was to evaluate the outcome of horses undergoing dorsal-to-palmar branch neuroanastomosis following PDN. STUDY DESIGN: Retrospective case series. ANIMALS: Eighty-five horses with PDN and dorsal-to-palmar branch neuroanastomosis. METHODS: Medical records for horses undergoing this procedure at two hospitals between 2015 and 2020 were reviewed. Palmar and dorsal nerve branches of the PDN were transected and end-to-end neuroanastomosis was performed by apposition of the perineurium. Follow-up was obtained from medical records and telephone interviews. Success was defined as resolution of lameness for at least one year. RESULTS: Lameness resolved following surgery in 81/85 (95%) horses with 57/84 (68%) sound at one year. Postoperative complications occurred in 19/85 (22%) cases. The main limitations of the study were an incomplete data set, inaccurate owner recall, and variations in procedure. CONCLUSION: Compared to previous studies, this technique resulted in similar numbers of horses sound immediately after surgery, a comparable rate of postoperative neuroma formation but a higher recurrence of lameness rate at 1 year postoperatively. CLINICAL SIGNIFICANCE: End-to-end neuroanastomosis of the dorsal and palmar branches of the PDN does not reduce the rate of neuroma formation in horses. Long-term outcome was less favorable compared to previously reported PDN techniques.

The use of lateral arthroscopy portals for the management of a fragmented lateral coronoid process in an English Bulldog.

OBJECTIVE: To report a case of unilateral lateral coronoid process fragmentation in a dog treated via lateral elbow arthroscopy portals. ANIMAL: A 9-month old male intact English Bulldog. STUDY DESIGN: Case report. METHODS: The dog presented with a history of intermittent right thoracic limb lameness. Orthopedic examination on presentation was unremarkable. Computed tomography of the right thoracic limb was pursued and revealed a mineralized focus along the lateral margin of the lateral coronoid process as well as sclerosis of the medial coronoid process and subtrochlear region of the ulna. Elbow arthroscopy was performed via a lateral approach and revealed chondromalacia of the entire lateral coronoid process. Abrasion arthroplasty of the lateral coronoid process was performed. RESULTS: Complete resolution of the lameness was achieved within two weeks of surgery. At 6 weeks postoperatively, the dog remained sound and a gradual return to normal activity was recommended. At the final follow up assessment, 5 months after surgery, no abnormalities were found on orthopedic examination and the owners reported excellent limb function with no observable lameness. CONCLUSION: Lateral coronoid disease can occur as a rare component of elbow dysplasia in dogs. Abrasion arthroplasty via lateral arthroscopic portals may have resulted in a successful outcome in this case and may form an effective treatment option for lateral coronoid disease in dogs.

A microglial activation cascade across cortical regions underlies secondary mechanical hypersensitivity to amputation.

Neural mechanisms underlying amputation-related secondary pain are unclear. Using in vivo two-photon imaging, three-dimensional reconstruction, and fiber photometry recording, we show that a microglial activation cascade from the primary somatosensory cortex of forelimb (S1FL) to the primary somatosensory cortex of hindlimb (S1HL) mediates the disinhibition and subsequent hyperexcitation of glutamatergic neurons in the S1HL (S1HLGlu), which then drives secondary mechanical hypersensitivity development in ipsilateral hindpaws of mice with forepaw amputation. Forepaw amputation induces rapid S1FL microglial activation that further activates S1HL microglia via the CCL2-CCR2 signaling pathway. Increased engulfment of GABAergic presynapses by activated microglia stimulates S1HLGlu neuronal activity, ultimately leading to secondary mechanical hypersensitivity of hindpaws. It is widely believed direct neuronal projection drives interactions between distinct brain regions to prime specific behaviors. Our study reveals microglial interactions spanning different subregions of the somatosensory cortex to drive a maladaptive neuronal response underlying secondary mechanical hypersensitivity at non-injured sites.