这项工作是支持由美国国立卫生研究院拨款NS052535聚糖EY014061＆EY014997的ARL，以及UL1DE019583 - 02财团奖。

为了避免偏见，实验者进行的评估不应该有动物的基因型知识。个别措施，取得了0-3分，0代表没有相关的表型和代表的最严重的表现的3。每个测试进行多次，以确保将比分是可重复的的。肥胖会所述的所有措施的解释复杂化。研究者不妨权衡表型的得分，以评估的结果可能发挥的作用，肥胖​​的老鼠。 礁测试窗台测试是一个直接的措施，协调，这是在小脑共济失调和许多其他神经退行性疾病中受损。这项措施是最直接可比人类小脑共济失调的迹象。 <ol><li>电梯从笼子里的鼠标和它放在笼子里的窗台上。老鼠通常会沿着窗台，企图入笼下降。 </li><li>观察鼠标，因为它走笼窗台，并降低其笼。野生型的小鼠通常会沿着窗台，而不会失去其平衡，会降低入笼优雅，用它的爪子。这是分配的得分为0。如果鼠标失去其立足点而沿窗台走，但在其他方面出现的协调，它接收到1分。如果没有有效地利用它的后腿，发挥得淋漓尽致，而不是它的爪子或土地入笼下降时，它接收到2分。如果脱落的窗台，或接近，而步行或试图降低自己，或动摇，并拒绝移动尽管鼓励所有，它接收到3分。有些老鼠会需要一个温柔的微调，以鼓励他们沿着窗台，或入笼下降。 </li><li>记录窗台测试得分。 </li></ol>后肢紧握后肢紧握的病情恶化，在小鼠模型的神经退行性疾病，包括某些小脑共济失调[1]的标记。 <ol><li>抓住它的基地附近的尾巴，并解除所有周围的物体清晰的鼠标。 </li><li>观察10秒的后肢位置。如果坚持后肢叉开向外，从腹部，它是分配的得分为0。如果一个后肢是朝着腹部的暂停时间超过50％的翻供，它接收到1分。如果两个后肢部分收回对暂停的时间超过50％的腹部时，它接收一个2分。如果它的后肢完全缩回，触摸腹部的暂停时间超过50％，它接收一个3分。 </li><li>将鼠标放回笼子里，并记录其后肢紧握得分。 </li></ol>步态步态是一个协调和肌肉功能的措施。 <ol><li>鼠标从笼中取出，并放置在一个平坦的表面，其头部远离研究者面临。 </li><li>观察鼠标，从后面行走。如果鼠标移动正常，支持其体重四肢，腹部不接触地面，并与后肢均参加均匀，收到了0分。如果它显示了震颤，或出现跛行，行走时，它接收1分。如果它显示了剧烈震颤，严重跛行，降低骨盆，或从身体，脚点在运动（"鸭脚"），它接收到2分。如果鼠标有困难向前迈进，并拖动它的腹部，沿地面的，它接收一个3分。 </li><li>将鼠标放回笼子里，并记录其步态评分。 </li></ol>脊柱后凸后凸畸形的一个特点是背的脊柱弯曲，常见的表现是神经退行性疾病小鼠模型[2]。这是由于继发于神经退行性病变的脊髓肌肉中的肌张力损失。 <ol><li>鼠标从笼中取出，放置在一个平面上上。 </li><li>观察它，因为它走。如果鼠标是能够很容易地拉直脊柱，因为它散步，不具有持久性脊柱后凸的，它接收得分为0。如果鼠标表现出轻度的脊柱后凸畸形，但能够理顺其脊椎，它接收到1分。如果不能彻底整顿其脊椎，并保持持续，但轻度后凸，它接收到2分。如果鼠标保持着明显的后凸畸形，如散步或而坐，它被分配一个3分。 </li><li>将鼠标放回笼子里，并记录其后凸得分。 </li></ol>代表性的成果有了足够数量的动物，这个程序能够检测菌株间内随着时间的推移一脉相承的表型差异。进行数据分析，计算所采取的每项措施的得分平均每一个人的三个测量。措施可以单独分析，或结合成复合材料的表型得分为各个。 图表中的数据和应用适当的统计方法来确定的意义。图1显示了从转基因小鼠SCA7模型的复合的表型评估的结果。在此floxed SCA7 - 92Q的转基因模型，两侧是人类ataxin - 7基因与92个CAG重复loxP位的网站，并从细菌人工染色体表达。复合材料的表型得分的紧握着，窗台走路，步态最大可能得分12和后凸评估。逐步SCA7在floxed SCA7 - 92Q的转基因小鼠的表型表现出越来越大的复合表型的得分，这与人类疾病的逐步性质是一致的。 <img src="/files/ftp_upload/1787/1787fig1.jpg" alt="图1" /> 图1 Floxed SCA7 92Q小鼠表现出一个渐进的SCA7表型，即开始显着不同的非转基因窝，在20周（2方差分析 :Bonferroni事后; *** P &lt; 0.001）。一个窗台测试每0-3规模的小鼠进行了评估，紧握着，步态和后凸畸形。每个基因型的平均综合得分，每个年龄计算。酒吧代表教育统筹局局长。

<ol>
	<li>Chou, A. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Polyglutamine-expanded+ataxin-3+causes+cerebellar+dysfunction+of+SCA3+transgenic+mice+by+inducing+transcriptional+dysregulation.">Polyglutamine-expanded ataxin-3 causes cerebellar dysfunction of SCA3 transgenic mice by inducing transcriptional dysregulation.</a> Neurobiol Dis. 31 (1), 89-101 (2008).</li><li>Thomas, P. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Loss+of+endogenous+androgen+receptor+protein+accelerates+motor+neuron+degeneration+and+accentuates+androgen+insensitivity+in+a+mouse+model+of+X-linked+spinal+and+bulbar+muscular+atrophy.">Loss of endogenous androgen receptor protein accelerates motor neuron degeneration and accentuates androgen insensitivity in a mouse model of X-linked spinal and bulbar muscular atrophy.</a> Hum Mol Genet. 15 (14), 2225-2238 (2006).</li><li>Ditzler, S., Stoeck, J., LeBlanc, M., Kooperberg, C., Hansen, S., Coppin, L., Olson, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Rapid+Neurobehavioral+Assessment+Reveals+that+FK506+Delays+Symptom+Onset+in+R6/2+Huntington's+Disease+Mice.">A Rapid Neurobehavioral Assessment Reveals that FK506 Delays Symptom Onset in R6/2 Huntington's Disease Mice.</a> Preclinica Research Articles. 1 (3), 115-126 (2003).</li></ol>

这些研究的大学华盛顿机构动物护理和使用委员会批准。

此协议是专为小脑共济失调的小鼠模型中的疾病严重程度的一个敏感和容易执行的评价。的打分系统的各个组件将在不同的神经退行性疾病小鼠模型或多或少有效。 这个评分系统的元素都被有效地用于评估的各种人类神经退行性疾病的小鼠模型，包括小脑共济失调，亨廷顿氏病和spinobulbar性肌萎缩症[1-3]。测试的理想组合将取决于问题的障碍。该协议最初是由作者设计，评估?...

a simple composite phenotype scoring system for evaluating mouse models of cerebellar ataxia

我们描述了一种快速，灵敏的定量小脑共济失调的小鼠模型中的疾病严重程度的协议。这是来自先前公布的表型在一些疾病模型评估，包括脊髓小脑共济失调，亨廷顿氏病和spinobulbar肌肉萎缩。措施包括后肢紧握着，窗台测试，步态和后凸。记录每一个措施是0-3分，与0-12的所有四项措施的总和。结果有效区分影响和非受影响的个人，同时也量化的神经退行性疾病表型的时间进展。措施可以单独分析，或结合成一个更大的统计力量的综合表型得分。理想的组合描述的四个措施将取决于问题的障碍。我们目前用来评估疾病的严重程度在脊髓小脑性共济失调7型（SCA7）的转基因小鼠模型的协议的一个例子。 阿尔伯特R.拉斯帕达和Gwenn A.花园贡献同样这个手稿。

Watch this Scientific Journal Video about A Simple Composite Phenotype Scoring System for Evaluating Mouse Models of Cerebellar Ataxia at JoVE.com

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

我们描述了一种快速，灵敏的定量小脑共济失调的小鼠模型中的疾病严重程度的协议。措施包括后肢紧握着，窗台测试，步态和后凸。该协议有效地区别对待受影响的和非受影响的个人，以及检测，随着时间的推移受影响的个人的进展。

一个简单的综合评分系统评估小脑共济失调的小鼠模型的表型

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

a-simple-composite-phenotype-scoring-system-for-evaluating-mouse

Research

JoVE Journal

Neuroscience

35.3K 次观看.  University of Washington. 我们描述了一种快速，灵敏的定量小脑共济失调的小鼠模型中的疾病严重程度的协议。措施包括后肢紧握着，窗台测试，步态和后凸。该协议有效地区别对待受影响的和非受影响的个人，以及检测，随着时间的推移受影响的个人的进展。

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

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.

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.

脑室周围白质软化的小鼠模型

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

科研

神经科学

Organotypic Cerebellar Cultures: Apoptotic Challenges and Detection

Organotypic cultures of neuronal tissue were first introduced by Hogue in 1947 1,2 and have constituted a major breakthrough in the field of neuroscience. Since then, the technique was developed further and currently there are many different ways to prepare organotypic cultures. The method presented here was adapted from the one described by Stoppini et al. for the preparation of the slices and from Gogolla et al. for the staining procedure 3,4.
A unique feature of this technique is that it allows you to study different parts of the brain such as hippocampus or cerebellum in their original structure, providing a big advantage over dissociated cultures in which all the cellular organization and neuronal networks are disrupted. In the case of the cerebellum it is even more advantageous because it allows the study of Purkinje cells, extremely difficult to obtain as dissociated primary culture. This method can be used to study certain developmental features of the cerebellum in vitro, as well as for electrophysiological and pharmacological experiments in both wild type and mutant mice.
The method described here was designed to study the effect of apoptotic stimuli such as Fas ligand in the developing cerebellum, using TUNEL staining to measure apoptotic cell death. If TUNEL staining is combined with cell type specific markers, such as Calbindin for Purkinje cells, it is possible to evaluate cell death in a cell population specific manner. The Calbindin staining also serves the purpose of evaluating the quality of the cerebellar cultures.

This method describes the generation of organotypic cerebellar cultures and the effect of certain apoptotic stimuli on the viability of different cerebellar cell types.

器官型小脑文化:细胞凋亡的挑战和检测

Organotypic cultures of neuronal tissue were first introduced by Hogue in 1947 1,2 and have constituted a major breakthrough in the field of neuroscience. ...

Quantitative Analysis of Synaptic Vesicle Pool Replenishment in Cultured Cerebellar Granule Neurons using FM Dyes

After neurotransmitter release in central nerve terminals, SVs are rapidly retrieved by endocytosis. Retrieved SVs are then refilled with neurotransmitter and rejoin the recycling pool, defined as SVs that are available for exocytosis1,2. The recycling pool can generally be subdivided into two distinct pools - the readily releasable pool (RRP) and the reserve pool (RP). As their names imply, the RRP consists of SVs that are immediately available for fusion while RP SVs are released only during intense stimulation1,2. It is important to have a reliable assay that reports the differential replenishment of these SV pools in order to understand 1) how SVs traffic after different modes of endocytosis (such as clathrin-dependent endocytosis and activity-dependent bulk endocytosis) and 2) the mechanisms controlling the mobilisation of both the RRP and RP in response to different stimuli.
FM dyes are routinely employed to quantitatively report SV turnover in central nerve terminals3-8. They have a hydrophobic hydrocarbon tail that allows reversible partitioning in the lipid bilayer, and a hydrophilic head group that blocks passage across membranes. The dyes have little fluorescence in aqueous solution, but their quantum yield increases dramatically when partitioned in membrane9. Thus FM dyes are ideal fluorescent probes for tracking actively recycling SVs. The standard protocol for use of FM dye is as follows. First they are applied to neurons and are taken up during endocytosis (Figure 1). After non-internalised dye is washed away from the plasma membrane, recycled SVs redistribute within the recycling pool. These SVs are then depleted using unloading stimuli (Figure 1). Since FM dye labelling of SVs is quantal10, the resulting fluorescence drop is proportional to the amount of vesicles released. Thus, the recycling and fusion of SVs generated from the previous round of endocytosis can be reliably quantified.
Here, we present a protocol that has been modified to obtain two additional elements of information. Firstly, sequential unloading stimuli are used to differentially unload the RRP and the RP, to allow quantification of the replenishment of specific SV pools. Secondly, each nerve terminal undergoes the protocol twice. Thus, the response of the same nerve terminal at S1 can be compared against the presence of a test substance at phase S2 (Figure 2), providing an internal control. This is important, since the extent of SV recycling across different nerve terminals is highly variable11.
Any adherent primary neuronal cultures may be used for this protocol, however the plating density, solutions and stimulation conditions are optimised for cerebellar granule neurons (CGNs)12,13.

A live fluorescence imaging technique to quantify the replenishment and mobilisation of specific synaptic vesicle (SV) pools in central nerve terminals is described. Two rounds of SV recycling are monitored in the same nerve terminals providing an internal control.

使用调频染料培养的小脑颗粒神经突触小泡池补充的定量分析

After neurotransmitter release in central nerve terminals, SVs are rapidly retrieved by endocytosis.  Retrieved SVs are then refilled with ...

Laser Nanosurgery of Cerebellar Axons In Vivo

Only a few neuronal populations in the central nervous system (CNS) of adult mammals show local regrowth upon dissection of their axon. In order to understand the mechanism that promotes neuronal regeneration, an in-depth analysis of the neuronal types that can remodel after injury is needed. Several studies showed that damaged climbing fibers are capable of regrowing also in adult animals1,2. The investigation of the time-lapse dynamics of degeneration and regeneration of these axons within their complex environment can be performed by time-lapse two-photon fluorescence (TPF) imaging in vivo3,4. This technique is here combined with laser surgery, which proved to be a highly selective tool to disrupt fluorescent structures in the intact mouse cortex5-9.
This protocol describes how to perform TPF time-lapse imaging and laser nanosurgery of single axonal branches in the cerebellum in vivo. Olivocerebellar neurons are labeled by anterograde tracing with a dextran-conjugated dye and then monitored by TPF imaging through a cranial window. The terminal portion of their axons are then dissected by irradiation with a Ti:Sapphire laser at high power. The degeneration and potential regrowth of the damaged neuron are monitored by TPF in vivo imaging during the days following the injury.

Laser Nanosurgery of Cerebellar Axons In Vivo

Two-photon imaging, coupled to laser nanodissection, are useful tools to study degenerative and regenerative processes in the central nervous system with subcellular resolution. This protocol shows how to label, image, and dissect single climbing fibers in the cerebellar cortex in vivo.

小脑轴突的激光Nanosurgery<em&gt;在体内</em

Only a few neuronal populations in the central nervous system (CNS) of adult mammals show local regrowth upon dissection of their axon. In order to ...

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.

嗅觉测定的神经退行性疾病的小鼠模型

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

Modeling Neuronal Death and Degeneration in Mouse Primary Cerebellar Granule Neurons

Cerebellar granule neurons (CGNs) are a commonly used neuronal model, forming an abundant homogeneous population in the cerebellum. In light of their post-natal development, abundance, and accessibility, CGNs are an ideal model to study neuronal processes, including neuronal development, neuronal migration, and physiological neuronal activity stimulation. In addition, CGN cultures provide an excellent model for studying different modes of cell death including excitotoxicity and apoptosis. Within a week in culture, CGNs express N-methyl-D-aspartate (NMDA) receptors, a specific ionotropic glutamate receptor with many critical functions in neuronal health and disease. The addition of low concentrations of NMDA in conjunction with membrane depolarization to rodent primary CGN cultures has been used to model physiological neuronal activity stimulation while the addition of high concentrations of NMDA can be employed to model excitotoxic neuronal injury. Here, a method of isolation and culturing of CGNs from 6 day old pups as well as genetic manipulation of CGNs by adenoviruses and lentiviruses are described. We also present optimized protocols on how to stimulate NMDA-induced excitotoxicity, low-potassium-induced apoptosis, oxidative stress and DNA damage following transduction of these neurons.

This protocol describes a simple method for isolating and culturing primary mouse cerebral granule neurons (CGNs) from 6-7 day old pups, efficient transduction of CGNs for loss and gain of function studies, and modelling NMDA-induced neuronal excitotoxicity, low-potassium-induced cell death, DNA-damage, and oxidative stress using the same culture model.

小鼠小脑颗粒神经元神经元死亡和变性的模型研究

Cerebellar granule neurons (CGNs) are a commonly used neuronal model, forming an abundant homogeneous population in the cerebellum. In light of their ...

Assessment of Long-term Depression Induction in Adult Cerebellar Slices

Synaptic plasticity provides a mechanism for learning and memory. For cerebellar motor learning, long-term depression (LTD) of synaptic transmissions from parallel fibers (PF) to Purkinje cells (PC) is considered the basis for motor learning, and deficiencies of both LTD and motor learning are observed in various gene-manipulated animals. Common motor learning sets, such as adaptation of the optokinetic reflex (OKR), the vestibular-ocular reflex (VOR), and rotarod test were used for evaluation of motor learning ability. However, results obtained from the GluA2-carboxy terminus modified knock-in mice demonstrated normal adaptation of the VOR and the OKR, despite lacking PF-LTD. In that report, induction of LTD was only attempted using one type of stimulation protocol at room temperature. Thus, conditions to induce cerebellar LTD were explored in the same knock-in mutants using various protocols at near physiological temperature. Finally, we found stimulation protocols, by which LTD could be induced in these gene-manipulated mice. In this study, a set of protocols are proposed to evaluate LTD-induction, which will more accurately allow examination of the causal relationship between LTD and motor learning. In conclusion, experimental conditions are crucial when evaluating LTD in gene-manipulated mice.

In some gene-manipulated animals, using a single protocol may fail to induce LTD in cerebellar Purkinje cells, and there may be a discrepancy between LTD and motor learning. Multiple protocols are necessary to assess LTD-induction in gene-manipulated animals. Standard protocols are shown.

成人脑切片长期抑郁症诱导评估

Synaptic plasticity provides a mechanism for learning and memory. For cerebellar motor learning, long-term depression (LTD) of synaptic transmissions from ...

Cerebellar Regional Dissection for Molecular Analysis

Cerebellum plays an important role in several key functions including control of movement, balance, cognition, reward, and affect. Imaging studies indicate that distinct cerebellar regions contribute to these different functions. Molecular studies examining regional cerebellar differences are lagging as they are mostly done on whole cerebellar extracts thereby masking any distinctions across specific cerebellar regions. Here we describe a technique to reproducibly and quickly dissect four different cerebellar regions: the deep cerebellar nuclei (DCN), anterior and posterior vermal cerebellar cortex, and the cerebellar cortex of the hemispheres. Dissecting out these distinct regions allows for the exploration of molecular mechanisms that may underlie their unique contributions to balance, movement, affect and cognition. This technique may also be used to explore differences in pathological susceptibility of these specific regions across various mouse disease models.

Different cerebellar regions have been implicated to play a role in distinct behavioral outputs, yet the underlying molecular mechanisms remain unknown. This work describes a method to reproducibly and quickly dissect cerebellar cortex of the hemispheres, anterior and posterior regions of the vermis, and the deep cerebellar nuclei in order to probe for molecular differences by isolating RNA and testing for differences in gene expression.&#160;

分子分析的谷类区域解剖

Cerebellum plays an important role in several key functions including control of movement, balance, cognition, reward, and affect. Imaging studies ...

Evaluation of Motor Impairment in C. elegans Models of Amyotrophic Lateral Sclerosis

The neurodegenerative disease amyotrophic lateral sclerosis (ALS) features progressive loss of motor neurons accompanied by muscle weakness and motor impairment that worsens with time. While considerable advances have been made in determining genetic drivers of ALS for a subset of patients, the majority of cases have an unknown etiology. Further, the mechanisms underlying motor neuron dysfunction and degeneration are not well understood; therefore, there is an ongoing need to develop and characterize representative models to study these processes. Caenorhabditis elegans can adapt their movement to the physical constraints of their surroundings, with two primary movement paradigms studied in a laboratory environment- crawling on a solid surface and swimming in liquid. These represent a complex interplay between sensation, motor neurons, and muscles. C. elegans models of ALS can exhibit impairment in one or both of these movement paradigms. This protocol describes two sensitive assays for evaluating motility in C. elegans: an optimized radial locomotion assay measuring crawling on a solid surface and an automated method for tracking and analyzing swimming in liquid (thrashing). In addition to the characterization of baseline motor impairment of ALS models, these assays can detect suppression or enhancement of the phenotypes from genetic or small molecule interventions. Thus, these methods have utility for studying ALS&#160;models and any C. elegans strain that exhibits altered motility.

Evaluation of Motor Impairment in C. elegans Models of Amyotrophic Lateral Sclerosis

This protocol describes two sensitive assays for discriminating among mild, moderate, and severe motor impairment in C. elegans models of amyotrophic lateral sclerosis, with general utility for C. elegans strains, with altered motility.

秀丽隐杆线虫运动障碍评估 肌萎缩性侧索硬化症模型

The neurodegenerative disease amyotrophic lateral sclerosis (ALS) features progressive loss of motor neurons accompanied by muscle weakness and motor ...

A Behavioral Screen for Heat-Induced Seizures in Mouse Models of Epilepsy

Transgenic mouse models have proved to be powerful tools in studying various aspects of human neurological disorders, including epilepsy. The SCN1A-associated genetic epilepsies comprise a wide spectrum of seizure disorders with incomplete penetrance and clinical variability. SCN1A mutations can result in a large variety of seizure phenotype ranging from simple, self-limited fever-associated febrile seizures (FS), moderate-level genetic epilepsy with febrile seizures plus (GEFS+) to more severe Dravet Syndrome (DS). Although FS are commonly seen in children below 6-7 years of age who do not have genetic epilepsy, FS in GEFS+ patients continue to occur into adulthood. Traditionally, experimental FS have been induced in mice by exposing the animal to a stream of dry air or heating lamps, and the rate of change in body temperature is often not well controlled. Here, we describe a custom-built heating chamber, with a plexiglass front, that is fitted with a digital temperature controller and a heater-equipped electric fan, which can send heated forced air into the test arena in a temperature-controlled manner. The body temperature of a mouse placed in the chamber, monitored through a rectal probe, can be increased to 40-42 &#176;C in a reproducible manner by increasing the temperature inside the chamber. Continual visual monitoring of the animals during the heating period demonstrates induction of heat-induced seizures in mice carrying an FS mutation at a body temperature that does not elicit behavioral seizures in wild-type litter mates. Animals can be easily removed from the chamber and placed on a cooling pad to rapidly return body temperature to normal. This method provides for a simple, rapid, and reproducible screening protocol for the occurrence of heat-induced seizures in epilepsy mouse models.

The goal of the method is to screen for hyperthermia or heat-induced seizures in mouse models. The protocol describes the use of a custom-built chamber with continuous monitoring of the body temperature to determine whether elevated body temperature leads to seizures.