This work is in part supported by grants from National Institutes of Health (RO1 NS059043 and RO1 ES015988), National Multiple Sclerosis Society, Roche Foundation for Anemia Research, Feldstein Medical Foundation, and Shriners Hospitals for Children.
We would like to thank Dr. David Pleasure for his suggestions. This work was in part supported by grants to W.D. from National Institutes of Health (RO1 NS059043 and RO1 ES015988), National Multiple Sclerosis Society, and Shriners Hospitals for Children. We declare no competing interests related to this article.

Detailed procedures of creating neonatal brain injury in P6 mice:
<ol>
<li>Postnatal day 6 (P6) mice are anesthetized with indirect cooling on ice to the point of unresponsiveness to noxious stimulation (deep anesthesia).</li>
 <li>Following cleaning the skin with alcohol, a midline ventral incision is made in the anterior neck.</li>
 <li>Under a dissecting microscope, the bifurcation of the right common carotid artery is approached by gently retracting the omohyoid and sternocleidomastoid muscles.</li>
 <li>The fascia around the carotid sheath is removed, and the proximal internal branch isolated from the nearby vagus nerve and sympathetic ganglia with a hook. The internal carotid artery is then cauterized using a cauterizing tip.</li>
 <li>Following cauterization, the skin incision is sutured, and the animal is kept warm until fully awake and then returned to the dam.</li>
 <li>One hour after ligation, the animal is placed in a sealed chamber infused with nitrogen until a level of 6.0% O2 is reached. The animal is then exposed to 35 min of hypoxia.</li>
 <li>After hypoxia exposure, the mouse recovers on a heating pad (33&deg;C) for 30 min and then is returned to the dam. For creation of brain injury with combined hypoxia/ischemia and infection/inflammation, the animal is then injected intraperitoneally with 0.015 ml of lipopolysaccharide (LPS, 1 mg/kg). </li>
 <li> Four days post-hypoxia/ischemia, mice are anesthetized with indirect cooling on ice to the point of unresponsiveness to noxious stimuli, then perfused with saline and then 4% paraformaldehyde. Brains are removed and cryoprotected.</li>
</ol>
<img src="/files/ftp_upload/1951/1951fig1.jpg" alt="Figure 1" /> Figure 1. Characterization of brain injury in the hypoxic/ischemic mouse model with H &amp; E staining. There is focal necrosis (arrows) in the cerebral white matter ipsilateral (A) to the carotid ligation, compared to the contralateral cerebral white matter (B). Ipsilateral necrosis and focal microinfarcts are observed in the hippocampus (C) and thalamus (D) in the brains with ipsilateral cerebral white matter injury. 
Representative Results and Figures:
Mouse models of PVL with hypoxia/ischemia and/or infection/inflammation are achieved by unilateral carotid ligation followed by exposure to hypoxia and injection of LPS. We are defining the new mouse models as suitable PVL models. Neuropathological examination of coronal sections stained with hematoxylin-and-eosin (H &amp; E) through the entire forebrain reveals focal necrosis in the central white matter of the cerebral hemisphere ipsilateral to the carotid ligation (Fig. 1A), with relative sparing of neurons in the overlying cerebral cortex compared to the contralateral cerebral white matter (Fig. 1B). In the focal necrotic white matter, there is marked increase in cellularity consisting of macrophages and reactive astrocytes, as is characteristic of focal human PVL. In the contralateral central white matter, the oligodendroglial nuclei are arranged in fascicular-like bundles; this architecture is completely disrupted by the focal necrosis on the ipsilateral side. Of note, we also observe ipsilateral necrosis in the hippocampus (Fig. 1C) and thalamus (focal microinfarcts) (Fig. 1D) in the brains with ipsilateral cerebral white matter injury. Human PVL also does not occur in complete isolation from accompanying gray matter lesions. The hallmark of PVL is the relative, although not complete, sparing of the cerebral cortex overlying the injured white matter - a feature that we believe is present in our mouse models.
<img src="/files/ftp_upload/1951/1951fig2.jpg" alt="Figure 2" /> Figure 2. Scoring scale of 0-5 for evaluating white matter injury by MBP or O1 in mouse models of periventricular leukomalacia.
Our mouse models show characteristics of PVL at both the phenotypic and molecular levels. We have used immunocytochemical detection of myelin basic protein (MBP) or O1-antigen to evaluate injury, have established a standard scale from 0 to 5 to score the white matter injury (Fig. 2), and have characterized the white matter pathology (Fig. 3). More recently, a non-biased stereological assay has been implemented to substitute for the initial semi-quantitative method. In addition, to test for correlative effects of white matter injury on functional outcomes, we have performed behavioral tests for motor function. We have demonstrated that the mice exhibit striking motor deficits at P10-21 following hypoxia/ischemia plus LPS at P6. We have established scoring criteria with a scale of 0-4 to define contralateral limb dysfunction in injured mice at P10-21. Typically, normal mice can scamper up a 30&deg; incline (score of 4) and climb up a 45&deg; incline without sliding (score of 3). Injured mice have lower scores in correlation with the degrees of white matter injury.
<img src="/files/ftp_upload/1951/1951fig3.jpg" alt="Figure 3" /> Figure 3. Characterization of white matter injury by MBP or O1 staining and myelin structure by electron microscope (EM). MBP and O1 staining show myelin loss in cerebral white matter ipsilateral to the ligation, compared to contralateral white matter. EM examination shows myelinated axons in cerebral white matter contralateral to the ligation and degenerated tissue in ipsilateral white matter. 
We anticipate that the establishment of the new mouse models of PVL will allow us to study specific mechanisms of PVL-like injury to the immature brain. We will address pathogenesis using transgenic mouse strains to examine the role of individual molecules of interest and evaluate targets of neuroprotection.

<ol>
	<li>Volpe, J. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Neurology of newborn. , 217-276 (2001).</li><li>Deng, W., Pleasure, J., Pleasure, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Progress+in+periventricular+leukomalacia.">Progress in periventricular leukomalacia.</a> Arch. Neurol. 65, 1291-1295 (2008).</li><li>Follett, P. L., Deng, W., Dai, W., Talos, D. M., Massillon, L. J., Rosenberg, P. A., Volpe, J. J., Jensen, F. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glutamate+receptor-mediated+oligodendrocyte+toxicity+in+periventricular+leukomalacia:+a+protective+role+for+topiramate.">Glutamate receptor-mediated oligodendrocyte toxicity in periventricular leukomalacia: a protective role for topiramate.</a> J. Neurosci. 24, 4412-4420 (2004).</li></ol>

No conflicts of interest declared.

Injury to the developing brain leads to devastating neurological consequences. Strikingly, the pattern of perinatal brain injury is highly age-dependent. In term infants, it predominantly affects cerebral cortex with characteristic neuronal loss. However, in premature infants, it selectively affects cerebral white matter with prominent injury to developing oligodendrocytes, a disorder termed periventricular leukomalacia (PVL). The magnitude of the problem of brain injury in the premature infant is extraordinary. Premature delivery and improved neonatal intensive care have led to nearly 90% of survival of the 13 million premature infants worldwide (approximately 56,000 in the United States) born yearly with a birthweight under 1500 grams, i.e., very low-birth-weight infants. Approximately 10% of the survivors subsequently exhibit cerebral palsy and approximately 50% have cognitive and behavioral deficits. PVL is the most important brain pathology underlying cerebral palsy in premature infants. No specific therapy for PVL presently exists, in part because the pathogenesis has not been well understood1,2.

Although the etiology of PVL is multifactorial, perinatal hypoxia/ischemia with maternal intrauterine infection is thought to be a primary cause of PVL. A hypoxic/ischemic model of PVL in the rat at P7 has been previously established3. This model is unique in that it resembles PVL in premature infants. The hallmark of this model is selective white matter pathology, in contrast to the majority of the stroke models that are characterized by gray matter infarction. However, the limitations of the hypoxic-ischemic model in the rat should not be overlooked. First, the etiology of PVL is often multifactorial. In addition to hypoxia/ischemia, maternal-fetal infection/inflammation is strongly associated with PVL. Thus, combined hypoxia/ischemia and infection/inflammation would likely create more PVL-like models. The rodent age that best correlates with the human developmental period of greatest risk for PVL lesions (24-32 weeks of gestation) is postnatal because oligodendroglial differentiation is predominantly a postnatal event in rodents. As it is impossible to produce a maternal-fetal infection/inflammation in postnatal ages, it would be reasonable to simulate an infection at a relevant postnatal developmental stage to create a rodent model of PVL. We have sought to combine LPS injection with the hypoxic/ischemic model in mice at P6, to produce more clinically relevant PVL lesions. Second, the optimal proof-of-concept experimental approach for a causative role of a molecule necessitates the use of transgenic mice; genetic manipulations are much harder in the rat than in the mouse. Thus, we have sought to convert our experimental model of PVL from the rat to the mouse.

Mouse models of PVL will greatly facilitate 1) the study of disease biology and pathogenesis using available transgenic mouse strains; 2) conduction of drug trials to discover candidate agents with therapeutic potential; 3) testing of the utility of stem cells using immunodeficient mouse strains.

<!DOCTYPE html PUBLIC "-//W3C//DTD XHTML 1.0 Transitional//EN" "http://www.w3.org/TR/xhtml1/DTD/xhtml1-transitional.dtd">
<html xmlns="http://www.w3.org/1999/xhtml" xml:lang="en" lang="en">	
		
		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>LPS</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>L4130</td>
<td>&nbsp;</td>
<tr>
<td>Hypoxia Chamber</td>
<td>&nbsp;</td>
<td>BioSpherix</td>
<td>A-30274</td>
<td>&nbsp;</td>
<tr>
<td>Surgical hook</td>
<td>&nbsp;</td>
<td>F.S.T.</td>
<td>10064-14</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

mouse models of periventricular leukomalacia

We describe a protocol for establishing mouse models of periventricular leukomalacia (PVL). PVL is the predominant form of brain injury in premature infants and the most common antecedent of cerebral palsy. PVL is characterized by periventricular white matter damage with prominent oligodendroglial injury. Hypoxia/ischemia with or without systemic infection/inflammation are the primary causes of PVL. We use P6 mice to create models of neonatal brain injury by the induction of hypoxia/ischemia with or without systemic infection/inflammation with unilateral carotid ligation followed by exposure to hypoxia with or without injection of the endotoxin lipopolysaccharide (LPS). Immunohistochemistry of myelin basic protein (MBP) or O1 and electron microscopic examination show prominent myelin loss in cerebral white matter with additional damage to the hippocampus and thalamus. Establishment of mouse models of PVL will greatly facilitate the study of disease pathogenesis using available transgenic mouse strains, conduction of drug trials in a relatively high throughput manner to identify candidate therapeutic agents, and testing of stem cell transplantation using immunodeficiency mouse strains.

Watch this Scientific Journal Video about Mouse Models of Periventricular Leukomalacia at JoVE.com

Mouse Models of Periventricular Leukomalacia

We established mouse models of periventricular leukomalacia (PVL), the predominant brain injury in premature infants characterized by periventricular white matter lesions. Hypoxia/ischemia with/without systemic infection are the primary causes of PVL. Unilateral carotid ligation and hypoxia exposure with/without lipopolysaccharide injection creates PVL-like lesions in P6 mice.

We describe a protocol for establishing mouse models of periventricular leukomalacia (PVL). PVL is the predominant form of brain injury in premature ...

mouse-models-of-periventricular-leukomalacia

Research

JoVE Journal

Neuroscience

29.7K Views.  University of California, Davis. Мы создали мышь модели перивентрикулярной лейкомаляции (ПВЛ), преобладающим травмы головного мозга у недоношенных детей характеризуется перивентрикулярной поражения белого вещества. Гипоксия / ишемии с / без системной инфекции являются основными причинами ...

Video: Mouse Models of Periventricular Leukomalacia

A Simple Composite Phenotype Scoring System for Evaluating Mouse Models of Cerebellar Ataxia

We describe a protocol for the rapid and sensitive quantification of disease severity in mouse models of cerebella ataxia. It is derived from previously published phenotype assessments in several disease models, including spinocerebellar ataxias, Huntington s disease and spinobulbar muscular atrophy. Measures include hind limb clasping, ledge test, gait and kyphosis. Each measure is recorded on a scale of 0-3, with a combined total of 0-12 for all four measures. The results effectively discriminate between affected and non-affected individuals, while also quantifying the temporal progression of neurodegenerative disease phenotypes. Measures may be analyzed individually or combined into a composite phenotype score for greater statistical power. The ideal combination of the four described measures will depend upon the disorder in question. We present an example of the protocol used to assess disease severity in a transgenic mouse model of spinocerebellar ataxia type 7 (SCA7).
Albert R. La Spada and Gwenn A. Garden contributed to this manuscript equally.

We describe a protocol for the rapid and sensitive quantification of disease severity in mouse models of cerebellar ataxia. Measures include hind limb clasping, ledge test, gait and kyphosis. This protocol effectively discriminates between affected and non-affected individuals, and detects the progression of affected individuals over time.

We describe a protocol for the rapid and sensitive quantification of disease severity in mouse models of cerebella ataxia. It is derived from previously ...

Experimental Models for Study of Retinal Pigment Epithelial Physiology and Pathophysiology

We have developed a cell culture procedure that can produce large quantities of confluent monolayers of primary human fetal retinal pigment epithelium (hfRPE) cultures with morphological, physiological and genetic characteristics of native human RPE. These hfRPE cell cultures exhibit heavy pigmentation, and electron microscopy show extensive apical membrane microvilli. The junctional complexes were identified with immunofluorescence labeling of various tight junction proteins. Epithelial polarity and function of these easily reproducible primary cultures closely resemble previously studied mammalian models of native RPE, including human. These results were extended by the development of therapeutic interventions in several animal models of human eye disease. We have focused on strategies for the removal of abnormal fluid accumulation in the retina or subretinal space. The extracellular subretinal space separates the photoreceptor outer segments and the apical membrane of the RPE and is critical for maintenance of retinal attachments and a whole host of RPE/retina interactions.

We provide a reproducible method for culturing confluent monolayers of human fetal retinal pigment epithelial cells (hfRPE) cells that exhibit morphology, physiology, polarity, and protein and gene expression patterns of adult native tissue. This work has been extended to an animal model of several eye diseases.

We have developed a cell culture procedure that can produce large quantities of confluent monolayers of primary human fetal retinal pigment epithelium ...

Multiple-mouse Neuroanatomical Magnetic Resonance Imaging

The field of mouse phenotyping with magnetic resonance imaging (MRI) is rapidly growing, motivated by the need for improved tools for characterizing and evaluating mouse models of human disease. MRI is an excellent modality for investigating genetically altered animals. It is capable of whole brain coverage, can be used in vivo, and provides multiple contrast mechanisms for investigating different aspects of neuranatomy and physiology. The advent of high-field scanners along with the ability to scan multiple mice simultaneously allows for rapid phenotyping of novel mutations.
Effective mouse MRI studies require attention to many aspects of experiment design. In this article, we will describe general methods to acquire quality images for mouse phenotyping using a system that images mice concurrently in shielded transmit/receive radio frequency (RF) coils in a common magnet (Bock et al., 2003). We focus particularly on anatomical phenotyping, an important and accessible application that has shown a high potential for impact in many mouse models at our imaging centre. Before we can provide the detailed steps to acquire such images, there are important practical considerations for both in vivo brain imaging (Dazai et al., 2004) and ex vivo brain imaging (Spring et al., 2007) that should be noted. These are discussed below.

Magnetic resonance imaging (MRI) has become an increasingly popular tool for examining the phenotype of genetically altered mice. This article illustrates the methods necessary to achieve high-throughput phenotyping of genetically altered mice using multiple-mouse MRI.

The field of mouse phenotyping with magnetic resonance imaging (MRI) is rapidly growing, motivated by the need for improved tools for characterizing and ...

Olfactory Assays for Mouse Models of Neurodegenerative Disease

In many neurodegenerative diseases and particularly in Parkinson&#8217;s disease, deficits in olfaction are reported to occur early in the disease process and may be a useful behavioral marker for early detection. Earlier detection in neurodegenerative disease is a major goal in the field because this is when neuroprotective therapies have the best potential to be effective. Therefore, in preclinical studies testing novel neuroprotective strategies in rodent models of neurodegenerative disease, olfactory assessment could be highly useful in determining therapeutic potential of compounds and translation to the clinic. In the present study we describe a battery of olfactory assays that are useful in measuring olfactory function in mice. The tests presented in this study were chosen because they measure olfaction abilities in mice related to food odors, social odors, and non-social odors. These tests have proven useful in characterizing novel genetic mouse models of Parkinson&#8217;s disease as well as in testing potential disease-modifying therapies.

Impairment in olfactory function is a common feature in many neurodegenerative disorders including Parkinson, Alzheimer, and Huntington diseases. In the present article, we describe a set of tests for assessing olfaction discrimination and detection in mice that can be used to measure olfactory abilities in mouse models of neurodegenerative diseases.

In many neurodegenerative diseases and particularly in Parkinson&#8217;s disease, deficits in olfaction are reported to occur early in the disease process ...

3D Modeling of the Lateral Ventricles and Histological Characterization of Periventricular Tissue in Humans and Mouse

The ventricular system carries and circulates cerebral spinal fluid (CSF) and facilitates clearance of solutes and toxins from the brain. The functional units of the ventricles are ciliated epithelial cells termed ependymal cells, which line the ventricles and through ciliary action are capable of generating laminar flow of CSF at the ventricle surface. This monolayer of ependymal cells also provides barrier and filtration functions that promote exchange between brain interstitial fluids (ISF) and circulating CSF. Biochemical changes in the brain are thereby reflected in the composition of the CSF and destruction of the ependyma can disrupt the delicate balance of CSF and ISF exchange. In humans there is a strong correlation between lateral ventricle expansion and aging. Age-associated ventriculomegaly can occur even in the absence of dementia or obstruction of CSF flow. The exact cause and progression of ventriculomegaly is often unknown; however, enlarged ventricles can show regional and, often, extensive loss of ependymal cell coverage with ventricle surface astrogliosis and associated periventricular edema replacing the functional ependymal cell monolayer. Using MRI scans together with postmortem human brain tissue, we describe how to prepare, image and compile 3D renderings of lateral ventricle volumes, calculate lateral ventricle volumes, and characterize periventricular tissue through immunohistochemical analysis of en face lateral ventricle wall tissue preparations. Corresponding analyses of mouse brain tissue are also presented supporting the use of mouse models as a means to evaluate changes to the lateral ventricles and periventricular tissue found in human aging and disease. Together, these protocols allow investigations into the cause and effect of ventriculomegaly and highlight techniques to study ventricular system health and its important barrier and filtration functions within the brain.

Using MRI scans (human), 3D imaging software, and immunohistological analysis, we document changes to the brain&#8217;s lateral ventricles. Longitudinal 3D mapping of lateral ventricle volume changes and characterization of periventricular cellular changes that occur in the human brain due to aging or disease are then modeled in mice.

The ventricular system carries and circulates cerebral spinal fluid (CSF) and facilitates clearance of solutes and toxins from the brain. The functional ...

Using Enzyme-based Biosensors to Measure Tonic and Phasic Glutamate in Alzheimer's Mouse Models

Neurotransmitter disruption is often a key component of diseases of the central nervous system (CNS), playing a role in the pathology underlying Alzheimer's disease, Parkinson's disease, depression, and anxiety. Traditionally, microdialysis has been the most common (lauded) technique to examine neurotransmitter changes that occur in these disorders. But because microdialysis has the ability to measure slow 1-20 minute changes across large areas of tissue, it has the disadvantage of invasiveness, potentially destroying intrinsic connections within the brain and a slow sampling capability. A relatively newer technique, the microelectrode array (MEA), has numerous advantages for measuring specific neurotransmitter changes within discrete brain regions as they occur, making for a spatially and temporally precise approach. In addition, using MEAs is minimally invasive, allowing for measurement of neurotransmitter alterations in vivo. In our laboratory, we have been specifically interested in changes in the neurotransmitter, glutamate, related to Alzheimer's disease pathology. As such, the method described here has been used to assess potential hippocampal disruptions in glutamate in a transgenic mouse model of Alzheimer's disease. Briefly, the method used involves coating a multi-site microelectrode with an enzyme very selective for the neurotransmitter of interest and using self-referencing sites to subtract out background noise and interferents. After plating and calibration, the MEA can be constructed with a micropipette and lowered into the brain region of interest using a stereotaxic device. Here, the method described involves anesthetizing rTg(TauP301L)4510 mice and using a stereotaxic device to precisely target sub-regions (DG, CA1, and CA3) of the hippocampus.

Using Enzyme-based Biosensors to Measure Tonic and Phasic Glutamate in Alzheimer&#039;s Mouse Models

Here, we describe the setup, software navigation, and data analysis for a spatially and temporally precise method of measuring tonic and phasic extracellular glutamate changes in vivo using enzyme-linked microelectrode arrays (MEA).

Neurotransmitter disruption is often a key component of diseases of the central nervous system (CNS), playing a role in the pathology underlying ...

Exploring Deep Space - Uncovering the Anatomy of Periventricular Structures to Reveal the Lateral Ventricles of the Human Brain

Anatomy students are typically provided with two-dimensional (2D) sections and images when studying cerebral ventricular anatomy and students find this challenging. Because the ventricles are negative spaces located deep within the brain, the only way to understand their anatomy is by appreciating their boundaries formed by related structures. Looking at a 2D representation of these spaces, in any of the cardinal planes, will not enable visualisation of all of the structures that form the boundaries of the ventricles. Thus, using 2D sections alone requires students to compute their own mental image of the 3D ventricular spaces. The aim of this study was to develop a reproducible method for dissecting the human brain to create an educational resource to enhance student understanding of the intricate relationships between the ventricles and periventricular structures. To achieve this, we created a video resource that features a step-by-step guide using a fiber dissection method to reveal the lateral and third ventricles together with the closely related limbic system and basal ganglia structures. One of the advantages of this method is that it enables delineation of the white matter tracts that are difficult to distinguish using other dissection techniques. This video is accompanied by a written protocol that provides a systematic description of the process to aid in the reproduction of the brain dissection. This package offers a valuable anatomy teaching resource for educators and students alike. By following these instructions educators can create teaching resources and students can be guided to produce their own brain dissection as a hands-on practical activity. We recommend that this video guide be incorporated into neuroanatomy teaching to enhance student understanding of the morphology and clinical relevance of the ventricles.

This paper demonstrates the effective use of a fiber dissection method to reveal the superficial white matter tracts and periventricular structures of the human brain, in three-dimensional space, to aid student comprehension of ventricular morphology.

Anatomy students are typically provided with two-dimensional (2D) sections and images when studying cerebral ventricular anatomy and students find this ...

Establishing Mouse Models for Zika Virus-induced Neurological Disorders Using Intracerebral Injection Strategies: Embryonic, Neonatal, and Adult

The Zika virus (ZIKV) is a flavivirus currently endemic in North, Central, and South America. It is now established that the ZIKV can cause microcephaly and additional brain abnormalities. However, the mechanism underlying the pathogenesis of ZIKV in the developing brain remains unclear. Intracerebral surgical methods are frequently used in neuroscience research to address questions about both normal and abnormal brain development and brain function. This protocol utilizes classical surgical techniques and describes methods that allow one to model ZIKV-associated human neurological disease in the mouse nervous system. While direct brain inoculation does not model the normal mode of virus transmission, the method allows investigators to ask targeted questions concerning the consequence after ZIKV infection of the developing brain. This protocol describes embryonic, neonatal, and adult stages of intraventricular inoculation of ZIKV. Once mastered, this method can become a straightforward and reproducible technique that only takes a few hours to perform.

Here we describe a method for establishing a model of Zika virus-induced microcephaly in mouse. This protocol includes methods for embryonic, neonatal, and adult-stage intracerebral inoculation of the Zika virus.

The Zika virus (ZIKV) is a flavivirus currently endemic in North, Central, and South America. It is now established that the ZIKV can cause microcephaly ...

Adult Mouse DRG Explant and Dissociated Cell Models to Investigate Neuroplasticity and Responses to Environmental Insults Including Viral Infection

This protocol describes an ex vivo model of mouse-derived dorsal root ganglia (DRG) explant and in vitro DRG-derived co-culture of dissociated sensory neurons and glial satellite cells. These are useful and versatile models to investigate a variety of biological responses associated with physiological and pathological conditions of the peripheral nervous system (PNS) ranging from neuron-glial interaction, neuroplasticity, neuroinflammation, and viral infection. The usage of DRG explant is scientifically advantageous compared to simplistic single cells models for multiple reasons. For instance, as an organotypic culture, the DRG explant allows ex vivo transfer of an entire neuronal network including the extracellular microenvironment that play a significant role in all the neuronal and glial functions. Further, DRG explants can also be maintained ex vivo for several days and the culture conditions can be perturbed as desired. In addition, the harvested DRG can be further dissociated into an in vitro co-culture of primary sensory neurons and satellite glial cells to investigate neuronal-glial interaction, neuritogenesis, axonal cone interaction with the extracellular microenvironment, and more general, any aspect associated with the neuronal metabolism. Therefore, the DRG-explant system offers a great deal of flexibility to study a wide array of events related to biological, physiological, and pathological conditions in a cost-effective manner.

In this report, the advantages of organotypic cultures and dissociated primary cultures of mouse-derived dorsal root ganglia are highlighted to investigate a wide range of mechanisms associated with neuron-glial interaction, neuroplasticity, neuroinflammation, and response to viral infection.

This protocol describes an ex vivo model of mouse-derived dorsal root ganglia (DRG) explant and in vitro DRG-derived co-culture of dissociated sensory ...

In Vivo Electrophysiological Measurement of Compound Muscle Action Potential from the Forelimbs in Mouse Models of Motor Neuron Degeneration

Assessing the functionality of the nerve axon provides detailed information on the progression of neuromuscular disorders. Electrophysiological recordings provide a sensitive approach to measure nerve conduction in humans and rodent models. To broaden the technical possibilities for electromyography in mice, the measurement of compound muscle action potentials (CMAPs) from the brachial plexus nerve in the forelimb using needle electrodes is described here. CMAP recordings after stimulating the sciatic nerve in hindlimbs have been previously described. The newly introduced method here allows for the evaluation of the nerve conductivity at an additional site, and thus provides a more profound overview of the neuromuscular functionality. The technique provides information on both the relative number of functional axons and the myelination level. Thereby, this method can be applied to assess both axonal diseases as well as demyelinating conditions. This minimally invasive method does not require extraction of the nerve and therefore it is suitable for repeated measurements for longitudinal follow-up in the same animal. Similar recordings are performed in clinical setups to emphasize the translational relevance of the method.

In Vivo Electrophysiological Measurement of Compound Muscle Action Potential from the Forelimbs in Mouse Models of Motor Neuron Degeneration

The measurement of nerve conduction is a useful tool to assess mouse models of neurodegeneration but it is frequently only applied to stimulate the sciatic nerve in hindlimbs. Here, we describe a technique to measure compound muscle action potential (CMAP) in vivo in the mouse forelimb muscles innervated by the brachial plexus.

Assessing the functionality of the nerve axon provides detailed information on the progression of neuromuscular disorders. Electrophysiological recordings ...