A Scalable Model to Study the Effects of Blunt-Force Injury in Adult Zebrafish

Hentig, James; Cloghessy, Kaylee; Dunseath, Chloe; Hyde, David R.

doi:10.3791/62709

Cited by 4 publications

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“…While a blunt-force zebrafish TBI model has rarely been examined [32] it offers the unique opportunity to also examine the regenerative response. To develop a scalable blunt-force TBI model, we modified the commonly used Marmarou weight drop (Figure 1A, created with BioRender.com) [10,36], with the impact zone centered at the intersection of the midbrain (mesencephalon and diencephalon; optic tectal lobes) and hindbrain (rhombencephalon; cerebellum, Figure 1A).…”

Section: Modified Marmarou Weight Drop Results In a Reproducible And Scalable Tbimentioning

confidence: 99%

“…Prior to inducing a blunt-force traumatic brain injury by a modified Marmarou weight drop [10,36], zebrafish were anesthetized in 1:1000 2-phenoxyethanol (2-PE, Sigma-Aldrich, St. Louis, MO, USA) until unresponsive to tail pinch. Anesthetized fish were secured onto a clay mold that stabilized the body and exposed the zebrafish head.…”

Section: Tbi Induction Via Modified Marmarou Weight Dropmentioning

confidence: 99%

“…Undamaged control and blunt-force damaged fish were euthanized by exposure to 1:500 2-PE and brains were removed as described previously [36]. Briefly, fish were pinned dorsal side up on a clay dish, eyes were enucleated, and forceps were inserted behind the lambda suture, removing the right parietal calvarial bone along the sagittal suture.…”

Section: Tissue Processingmentioning

confidence: 99%

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Zebrafish Blunt-Force TBI Induces Heterogenous Injury Pathologies That Mimic Human TBI and Responds with Sonic Hedgehog-Dependent Cell Proliferation across the Neuroaxis

et al. 2021

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Blunt-force traumatic brain injury (TBI) affects an increasing number of people worldwide as the range of injury severity and heterogeneity of injury pathologies have been recognized. Most current damage models utilize non-regenerative organisms, less common TBI mechanisms (penetrating, chemical, blast), and are limited in scalability of injury severity. We describe a scalable blunt-force TBI model that exhibits a wide range of human clinical pathologies and allows for the study of both injury pathology/progression and mechanisms of regenerative recovery. We modified the Marmarou weight drop model for adult zebrafish, which delivers a scalable injury spanning mild, moderate, and severe phenotypes. Following injury, zebrafish display a wide range of severity-dependent, injury-induced pathologies, including seizures, blood–brain barrier disruption, neuroinflammation, edema, vascular injury, decreased recovery rate, neuronal cell death, sensorimotor difficulties, and cognitive deficits. Injury-induced pathologies rapidly dissipate 4–7 days post-injury as robust cell proliferation is observed across the neuroaxis. In the cerebellum, proliferating nestin:GFP-positive cells originated from the cerebellar crest by 60 h post-injury, which then infiltrated into the granule cell layer and differentiated into neurons. Shh pathway genes increased in expression shortly following injury. Injection of the Shh agonist purmorphamine in undamaged fish induced a significant proliferative response, while the proliferative response was inhibited in injured fish treated with cyclopamine, a Shh antagonist. Collectively, these data demonstrate that a scalable blunt-force TBI to adult zebrafish results in many pathologies similar to human TBI, followed by recovery, and neuronal regeneration in a Shh-dependent manner.

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Section: Modified Marmarou Weight Drop Results In a Reproducible And Scalable Tbimentioning

confidence: 99%

Section: Tbi Induction Via Modified Marmarou Weight Dropmentioning

confidence: 99%

Section: Tissue Processingmentioning

confidence: 99%

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Zebrafish Blunt-Force TBI Induces Heterogenous Injury Pathologies That Mimic Human TBI and Responds with Sonic Hedgehog-Dependent Cell Proliferation across the Neuroaxis

et al. 2021

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“…2-PE was used because it is approved for anesthesia in zebrafish and has no effect on neuronal function even at higher concentrations [ 25 ]. A modified Marmarou weight drop protocol for zebrafish [ 17 , 26 ] was used to administer a blunt-force injury. Following anesthetic, fish were secured onto a clay mold that stabilized the body and exposed the zebrafish head.…”

Section: Methodsmentioning

confidence: 99%

“…Edema was measured using a modified Hoshi protocol [ 26 , 27 ]. Whole brains (undamaged, sTBI, or sTBI treated fish n = 9 fish control/experimental group) were isolated from control and injured fish at 1 or 3 dpi, weighed, dried at 60 °C for 12 h, and weighed again, with the percent fluid calculated using the following formula:

…”

Section: Methodsmentioning

confidence: 99%

Prophylactic Activation of Shh Signaling Attenuates TBI-Induced Seizures in Zebrafish by Modulating Glutamate Excitotoxicity through Eaat2a

et al. 2021

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Approximately 2 million individuals experience a traumatic brain injury (TBI) every year in the United States. Secondary injury begins within minutes after TBI, with alterations in cellular function and chemical signaling that contribute to excitotoxicity. Post-traumatic seizures (PTS) are experienced in an increasing number of TBI individuals that also display resistance to traditional anti-seizure medications (ASMs). Sonic hedgehog (Shh) is a signaling pathway that is upregulated following central nervous system damage in zebrafish and aids injury-induced regeneration. Using a modified Marmarou weight drop on adult zebrafish, we examined PTS following TBI and Shh modulation. We found that inhibiting Shh signaling by cyclopamine significantly increased PTS in TBI fish, prolonged the timeframe PTS was observed, and decreased survival across all TBI severities. Shh-inhibited TBI fish failed to respond to traditional ASMs, but were attenuated when treated with CNQX, which blocks ionotropic glutamate receptors. We found that the Smoothened agonist, purmorphamine, increased Eaat2a expression in undamaged brains compared to untreated controls, and purmorphamine treatment reduced glutamate excitotoxicity following TBI. Similarly, purmorphamine reduced PTS, edema, and cognitive deficits in TBI fish, while these pathologies were increased and/or prolonged in cyclopamine-treated TBI fish. However, the increased severity of TBI phenotypes with cyclopamine was reduced by cotreating fish with ceftriaxone, which induces Eaat2a expression. Collectively, these data suggest that Shh signaling induces Eaat2a expression and plays a role in regulating TBI-induced glutamate excitotoxicity and TBI sequelae.

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A Robust and Reproducible Method to Study Neurorepair after Stab Injury in the African Turquoise Killifish Telencephalon

Mariën

Arckens

houcke

2023

Cold Spring Harb Protoc

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The aging population (people >60 yr old) is steadily increasing worldwide, resulting in an increased prevalence of age-related neurodegenerative diseases. Despite intensive research efforts in the past decades, there are still no therapies available to stop, cure, or prevent these diseases. Induction of successful neuroregeneration (i.e., the production of new neurons that can functionally integrate into the existing neural circuitry) could represent a therapy to replace neurons lost by injury or disease in the aged central nervous system. The African turquoise killifish, with its particularly short life span, has emerged as a useful model to study how aging influences neuroregeneration. Here, we describe a robust and reproducible stab-injury protocol to study regeneration in the telencephalon of the African turquoise killifish. After the injury, newborn cells are traced by conducting a BrdU pulse-chase experiment. To identify newborn neurons, a double immunohistochemical staining for BrdU and HuCD is carried out. Techniques such as bromodeoxyuridine (BrdU) labeling, intracardial perfusion, cryosectioning, and immunofluorescence staining are described as separate sections.

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A Scalable Model to Study the Effects of Blunt-Force Injury in Adult Zebrafish

Cited by 4 publications

References 0 publications

Zebrafish Blunt-Force TBI Induces Heterogenous Injury Pathologies That Mimic Human TBI and Responds with Sonic Hedgehog-Dependent Cell Proliferation across the Neuroaxis

Zebrafish Blunt-Force TBI Induces Heterogenous Injury Pathologies That Mimic Human TBI and Responds with Sonic Hedgehog-Dependent Cell Proliferation across the Neuroaxis

Prophylactic Activation of Shh Signaling Attenuates TBI-Induced Seizures in Zebrafish by Modulating Glutamate Excitotoxicity through Eaat2a

A Robust and Reproducible Method to Study Neurorepair after Stab Injury in the African Turquoise Killifish Telencephalon

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