This work was supported in part by the Lupus Research Institute, NCI grant CA138617, NIDDK grant DK090978, Charlotte Schmidlapp Award (to E.M.J.), and the Albert J. Ryan Fellowship (to J.K.). We are grateful for the support and instrumentation provided by the Research Flow Cytometry Core in the Division of Rheumatology at Cincinnati Children&#8217;s Hospital Medical Center, supported in part by NIH AR-47363, NIH DK78392 and NIH DK90971.

<ol>
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The authors have nothing to disclose.

המודל מושרה bm12 הוא דרך קלה יחסית ויעילה ללמוד את התהליכים התאיים ומולקולריים של SLE. הפעלה כרונית של תאי CD4 T הועברו adoptively המכוון נגד אנטיגנים עצמיים מובילה להצטברות של TFH, GC תאי B, ותאי פלזמה אשר ניתן למדוד על ידי cytometry זרימה, כפי שתואר כאן. מחקרים עתידיים באמצעות מודל זה ?...

זאבת אדמנתית מערכתית (לופוס) היא מחלה אוטואימונית מורכבת המאופיינת prototypically על ידי נוגדן אנטי-גרעיני ייצור (ANA) וגלומרולונפריטיס. סיבוכים רבים אחרים, כוללים עורי, לב-ריאה, ונגעים בכבד קשורים למחלה אצל אנשים מסוימים. הערכות שכיחות בארה&quot;ב משתנות במידה רבה, מ150,000-1,500,000 1,2, עם שכיחות גבוהה במיוחד בנשים ומיעוטים 3. למרות אטיולוגיה של SLE כבר קשה להבחין, הוא חשב שינבע ממשחק הגומלין של גורמים גנטיים וסביבתיים שונים, שיגיעו לשיאו באוטואימוניות המערכתית. מודלים של בעלי חיים רבים כבר מועסקים ללמוד גורמים מובילים להופעת מחלה והתקדמות. מודלים עכבר קלאסיים של SLE כוללים זני נטייה גנטית עכבר כוללים NZB x מודל NZW F1 ונגזרי NZM, המתח / LPR הקה&quot;ע, ומתח BXSB / Yaa, ומערכות מושרה, כגון pristane ודגמי מחלת שתל נגד המארח כרוני (cGVHD) 4. דיווחים מוקדמים של ייצור נוגדנים בדגמי GVHD משמשים זנים שונים עכבר או זני אוגר להורה להעברות F1 5 - 8; שיטות נפוצות יותר בשימוש ללמוד מחלה כמו זאבת-כיום כוללות הורה DBA / 2 → (DBA / 2 x C57BL / 6) F1, ומודל העברת bm12 המתואר כאן. לכל אחד יש מודל האזהרות שלה, אבל הם בדרך כלל לשתף מערך משותף של תכונות המתואמים עם מאפיינים קליניים של מחלה אנושית. הפרמטרים לרוב דיווחו במודלים של עכברים כוללים splenomegaly, בלוטות הלימפה, דלקת כליות, ייצור ANA, וברמה התאית, ההתרחבות של תאי T זקיקי עוזר (TFH), תאי B נבטי מרכז (GC), ותאי פלזמה. מודל bm12 מושרה מושגת על ידי העברת המאמצת של לימפוציטים מIA bm12 B6 (C) - H2 - עכברים (bm12) AB1 bm12 / KhEgJ, מתח אניdentical לC57BL / 6 למעט 3 החלפות חומצת אמינו בכיתת MHC II, לIA ב C57BL / 6 עכברים (B6). Alloactivation של תורם CD4 T תאים על ידי נמען נגמ&quot;שים מוביל לcGVHD עם תסמינים דומים SLE מקרוב. באופן ספציפי, אלה כוללים הרחבת TFH תורם נגזרות, התרחבות של תאי שמקורם נמען GC B ותאי פלזמה, וייצור של אנאס כולל אנטי-dsDNA, אנטי-ssDNA, אנטי-הכרומטין, ונוגדנים נגד RBC 9. לאורך זמן, עכברי נמען לפתח גלומרולונפריטיס הקשורים פיקדונות IgG בגלומרולרי, ביניים, ואזורים של כלי דם של הכליות 10. הראינו לאחרונה כי, בדומה למחלות של בני אדם, יש גם תפקיד קריטי עבור סוג אני IFN במודל זה 11. יש לציין, הקריטריונים המגדירים עבור SLE האנושי כוללים את הפיתוח של דלקת כליות תואמות עם SLE בנוכחות נוגדנים נגד dsDNA 12, שניהם מאפיינים בולטים של מודל עכבר זה. יש seיתרונות veral של מודל bm12 על המודלים ספונטניים. מודלים קלאסיים שלפתח סימנים כמו SLE-ספונטני להסתמך על שני זנים היברידיים עכבר, זנים טהורים עכבר לא על רקע B6, או לוקוסים גנטיים גדולים על רקע B6, שהופכים את מעבר לפצצה או עכברים מהונדסים גנטי אחר קשה זמן רב. עם המודל מושרה bm12, עכברים מהונדסים גנטי יכולים לשמש גם תורם או נמען, המאפשרים זיהוי מהיר יותר של התא הסלולרי שבגנים מסוימים עשויים להיות חשובים למחלה. יתר על כן, התפתחות מחלה במודל bm12 היא הרבה יותר מהר, הדורש רק 2 שבועות עד להופעת אנאס, בהשוואה למספר חודשים לדגמים ספונטניים ביותר. יתר על כן, בניגוד לדגמים ספונטניים שמתפתחים מחלה בנקודות זמן שונה, תחילת המחלה וההתקדמות במודל B6 → bm12 מסונכרנת מאוד. זה מאפשר לדור של קבוצות בגודל המתאימה שיכולות בהדואר משמש לאסטרטגיות התערבותית או טיפוליות בכל שלב של התפתחות מחלה. להלן פרוטוקול מפורט לייזום אוטואימוניות כמו SLE-ידי העברת המאמצת של לימפוציטים bm12 לעכברי C57BL / 6, או וריאנטים גנטיים על רקע B6. בנוסף, אנו מתארים פרוטוקול מכתים הזרימה cytometric לספירת TFH, GC תאי B, וסוגי תאי פלזמה תאים הקשורים למחלות אנושיות. חשוב לציין, יכולים לשמש גם פרוטוקולים אלה לאפיין מחלה ברוב דגמי עכבר של SLE ולזהות TFH, GC תאי B ותאי פלזמה בדגמי מחלה אחרים.

<table border="0">
	<tbody>
		<tr>
			<td>B6.SJL-Ptprca Pepcb/BoyJ</td>
			<td>The Jackson Laboratory</td>
			<td>001162</td>
			<td>CD45.1+ BoyJ mouse strain</td>
		</tr>
		<tr>
			<td>B6(C)-H2-Ab1bm12/KhEgJ</td>
			<td>The Jackson Laboratory</td>
			<td>001162</td>
			<td>Bm12 mouse strain</td>
		</tr>
		<tr>
			<td>FastDigest PsuI</td>
			<td>Life Technologies</td>
			<td>FD1554</td>
			<td>Restriction digest enzyme for genotyping</td>
		</tr>
		<tr>
			<td>1X RBC Lysis Buffer</td>
			<td>eBioscience</td>
			<td>00-4333-57</td>
			<td></td>
		</tr>
		<tr>
			<td>IMDM</td>
			<td>GE Healthcare</td>
			<td>SH30228.01</td>
			<td></td>
		</tr>
		<tr>
			<td>Plasma Separation Tube (PST)</td>
			<td>BD</td>
			<td>365974</td>
			<td>Blood collection tube with Dipotassium EDTA</td>
		</tr>
		<tr>
			<td>Serum Separation Tube (SST)</td>
			<td>BD</td>
			<td>365967</td>
			<td>Blood collection tube with Clot activator / SST Gel</td>
		</tr>
		<tr>
			<td>Ficoll</td>
			<td>GE Healthcare</td>
			<td>17-1440-02&#160;</td>
			<td>High density cell separation solution</td>
		</tr>
		<tr>
			<td>Lympholyte-M</td>
			<td>Cedarlane</td>
			<td>CL5030</td>
			<td>High density cell separation solution</td>
		</tr>
		<tr>
			<td>GL-7-biotin</td>
			<td>eBioscience</td>
			<td>13-5902-82&#160;</td>
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			<td>Streptavidin-BUV395</td>
			<td>BD</td>
			<td>564176</td>
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			<td>CD138-BV421</td>
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			<td>CD4-BV510</td>
			<td>BioLegend</td>
			<td>100559</td>
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			<td>TCR&#946;-BV605</td>
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			<td>CD45.1-BV711</td>
			<td>BioLegend</td>
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			<td>CD45.2-FITC</td>
			<td>BioLegend</td>
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			<td>PD-1-PE</td>
			<td>BioLegend</td>
			<td>135206</td>
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			<td>CD19-PerCP</td>
			<td>BioLegend</td>
			<td>115532</td>
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			<td>Fas-PE-Cy7</td>
			<td>BD</td>
			<td>557653</td>
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			<td>CXCR5-APC</td>
			<td>BioLegend</td>
			<td>145506</td>
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			<td>Fixable Viability Dye ef780</td>
			<td>eBioscience</td>
			<td>65-0865-18</td>
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			<td>CD4-BV421</td>
			<td>BioLegend</td>
			<td>100443</td>
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			<td>1.2 ml FACS tube inserts, racked</td>
			<td>USA Scientific</td>
			<td>1412-1400</td>
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			<td>BD Falcon&#8482; Round-Bottom Tubes</td>
			<td>BD</td>
			<td>352017</td>
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</table>

the bm12 inducible model of systemic lupus erythematosus (sle) in c57bl/6 mice

Systemic lupus erythematosus (SLE) is an autoimmune disease with diverse clinical and immunological manifestations. Several spontaneous and inducible animal models mirror common components of human disease, including the bm12 transfer model. Upon transfer of bm12 splenocytes or purified CD4 T cells, C57BL/6 mice rapidly develop large frequencies of T follicular helper cells (Tfh), germinal center (GC) B cells, and plasma cells followed by high levels of circulating anti-nuclear antibodies. Since this model utilizes mice on a pure C57BL/6 background, researchers can quickly and easily study disease progression in transgenic or knockout mouse strains in a relatively short period of time. Here we describe protocols for the induction of the model and the quantitation Tfh, GC B cells, and plasma cells by multi-color flow cytometry. Importantly, these protocols can also be used to characterize disease in most mouse models of SLE and identify Tfh, GC B cells, and plasma cells in other disease models.

עכברים חולים לפתח splenomegaly קטן כמו 14 ימים, מציגים טחולים 2-3 פעמים בגודל של עכברים בריאים במונחים של מסה וcellularity (איור 2). Splenocytes הם מגודרת ברצף על פיזור אור (FSC-ידי SSC-), חיסול כפילויות (FSC-W או -H ידי FSC-A), תאי קיימא (הכתמה נמוכה של צ?...

Watch this Scientific Journal Video about The bm12 Inducible Model of Systemic Lupus Erythematosus SLE in C57BL/6 Mice at JoVE.com

The bm12 Inducible Model of Systemic Lupus Erythematosus SLE in C57BL/6 Mice

The transfer of bm12 lymphocytes into a C57BL/6 recipient is an established model of systemic lupus erythematosus. Here we describe how to initiate disease using this model and how to characterize T follicular helper cells, germinal center B cells and plasma cells by flow cytometry.

דגם bm12 העין מתנהל של זאבת אדמנתית המערכתית (לופוס) בC57BL עכברים / 6

Systemic lupus erythematosus (SLE) is an autoimmune disease with diverse clinical and immunological manifestations. Several spontaneous and inducible ...

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Research

JoVE Journal

Medicine

The bm12 Inducible Model of Systemic Lupus Erythematosus (SLE) in C57BL/6 Mice

17.2K Views.  Cincinnati Children's Hospital Medical Center and University of Cincinnati College of Medicine. The transfer of bm12 lymphocytes into a C57BL/6 recipient is an established model of systemic lupus erythematosus. Here we describe how to initiate disease using this model and how to characterize T follicular helper cells, germinal center B cells and plasma cells by flow cytometry.

Video: The bm12 Inducible Model of Systemic Lupus Erythematosus SLE in C57BL/6 Mice

The Spared Nerve Injury (SNI) Model of Induced Mechanical Allodynia in Mice

Peripheral neuropathic pain is a severe chronic pain condition which may result from trauma to sensory nerves in the peripheral nervous system. The spared nerve injury (SNI) model induces symptoms of neuropathic pain such as mechanical allodynia i.e. pain due to tactile stimuli that do not normally provoke a painful response [1]. 
The SNI mouse model involves ligation of two of the three branches of the sciatic nerve (the tibial nerve and the common peroneal nerve), while the sural nerve is left intact [2]. The lesion results in marked hypersensitivity in the lateral area of the paw, which is innervated by the spared sural nerve. The non-operated side of the mouse can be used as a control. The advantages of the SNI model are the robustness of the response and that it doesn&#x2019;t require expert microsurgical skills.
The threshold for mechanical pain response is determined by testing with von Frey filaments of increasing bending force, which are repetitively pressed against the lateral area of the paw [3], [4]. A positive pain reaction is defined as sudden paw withdrawal, flinching and/or paw licking induced by the filament. A positive response in three out of five repetitive stimuli is defined as the pain threshold. 
As demonstrated in the video protocol, C57BL/6 mice experience profound allodynia as early as the day following surgery and maintain this for several weeks.

The Spared Nerve Injury SNI Model of Induced Mechanical Allodynia in Mice

The Spared Nerve Injury animal model is described here as a mouse model of peripheral neuropathic pain following partial denervation of the sciatic nerve by lesioning the tibial and common peroneal nerve branches, leaving the remaining sural nerve intact. Behavioral modification resulting from mechanical allodynia is quantified by von Frey filaments.

פגיעה ניצלו עצב (SNI) דגם Allodynia מכונות המושרה עכברים

Peripheral neuropathic pain is a severe chronic pain condition which may result from trauma to sensory nerves in the peripheral nervous system. The spared ...

Tibial Nerve Transection - A Standardized Model for Denervation-induced Skeletal Muscle Atrophy in Mice

The tibial nerve transection model is a well-tolerated, validated, and reproducible model of denervation-induced skeletal muscle atrophy in rodents. Although originally developed and used extensively in the rat due to its larger size, the tibial nerve in mice is big enough that it can be easily manipulated with either crush or transection, leaving the peroneal and sural nerve branches of the sciatic nerve intact and thereby preserving their target muscles. Thus, this model offers the advantages of inducing less morbidity and impediment of ambulation than the sciatic nerve transection model and also allows investigators to study the physiologic, cellular and molecular biologic mechanisms regulating the process of muscle atrophy in genetically engineered mice. The tibial nerve supplies the gastrocnemius, soleus and plantaris muscles, so its transection permits the study of denervated skeletal muscle composed of fast twitch type II fibers and/or slow twitch type I fibers. Here we demonstrate the tibial nerve transection model in the C57Black6 mouse. We assess the atrophy of the gastrocnemius muscle, as a representative muscle, at 1, 2, and 4 weeks post-denervation by measuring muscle weights and fiber type specific cross-sectional area on paraffin-embedded histologic sections immunostained for fast twitch myosin.

The tibial nerve transection model is a well-tolerated, validated, and reproducible model of skeletal muscle atrophy. The model surgical protocol is described and demonstrated in C57Black6 mice. &#160;

חיתוך רוחב עצב הטיביאלי - מודל סטנדרטי לניוון שרירים שלד מושרה denervation בעכברים

The tibial nerve transection model is a well-tolerated, validated, and reproducible model of denervation-induced skeletal muscle atrophy in rodents. ...

Assessing Changes in Volatile General Anesthetic Sensitivity of Mice after Local or Systemic Pharmacological Intervention

One desirable endpoint of general anesthesia is the state of unconsciousness, also known as hypnosis. Defining the hypnotic state in animals is less straightforward than it is in human patients. A widely used behavioral surrogate for hypnosis in rodents is the loss of righting reflex (LORR), or the point at which the animal no longer responds to their innate instinct to avoid the vulnerability of dorsal recumbency. We have developed a system to assess LORR in 24 mice simultaneously while carefully controlling for potential confounds, including temperature fluctuations and varying gas flows. These chambers permit reliable assessment of anesthetic sensitivity as measured by latency to return of the righting reflex (RORR) following a fixed anesthetic exposure. Alternatively, using stepwise increases (or decreases) in anesthetic concentration, the chambers also enable determination of a population's sensitivity to induction (or emergence) as measured by EC50 and Hill slope. Finally, the controlled environmental chambers described here can be adapted for a variety of alternative uses, including inhaled delivery of other drugs, toxicology studies, and simultaneous real-time monitoring of vital signs.

Loss of the righting reflex has long served as a standard behavioral surrogate for unconsciousness, also called hypnosis, in laboratory animals. Alterations in volatile anesthetic sensitivity caused by pharmacological interventions can be detected with a carefully controlled high-throughput assessment system, which may be adapted for delivery of any inhaled therapeutic.

הערכת שינויים בנדיפות כללית הרדמה רגישות של עכברים לאחר התערבות תרופתי מקומית או מערכתית

One desirable endpoint of general anesthesia is  the state of unconsciousness, also known as hypnosis. Defining the hypnotic state in animals is  less ...

The Sciatic Nerve Cuffing Model of Neuropathic Pain in Mice

Neuropathic pain arises as a consequence of a lesion or a disease affecting the somatosensory system. This syndrome results from maladaptive changes in injured sensory neurons and along the entire nociceptive pathway within the central nervous system. It is usually chronic and challenging to treat. In order to study neuropathic pain and its treatments, different models have been developed in rodents. These models derive from known etiologies, thus reproducing peripheral nerve injuries, central injuries, and metabolic-, infectious- or chemotherapy-related neuropathies. Murine models of peripheral nerve injury often target the sciatic nerve which is easy to access and allows nociceptive tests on the hind paw. These models rely on a compression and/or a section. Here, the detailed surgery procedure for the &#34;cuff model&#34; of neuropathic pain in mice is described. In this model, a cuff of PE-20 polyethylene tubing of standardized length (2 mm) is unilaterally implanted around the main branch of the sciatic nerve. It induces a long-lasting mechanical allodynia, i.e., a nociceptive response to a normally non-nociceptive stimulus that can be evaluated by using von Frey filaments. Besides the detailed surgery and testing procedures, the interest of this model for the study of neuropathic pain mechanism, for the study of neuropathic pain sensory and anxiodepressive aspects, and for the study of neuropathic pain treatments are also discussed.

Neuropathic pain is a consequence of a lesion or disease affecting the somatosensory system. The &#8220;cuff model&#8221; of neuropathic pain in mice consists of the implantation of a polyethylene cuff around the main branch of the sciatic nerve. Mechanical allodynia is tested using von Frey filaments.

דגם השת עצב איזוק של כאב נוירופתי בעכברים

Neuropathic pain arises as a consequence of a lesion or a disease affecting the somatosensory system. This syndrome results from maladaptive changes in ...

Monitoring of Systemic and Hepatic Hemodynamic Parameters in Mice

The use of mouse models in experimental research is of enormous importance for the study of hepatic physiology and pathophysiological disturbances. However, due to the small size of the mouse, technical details of the intraoperative monitoring procedure suitable for the mouse were rarely described. Previously we have reported a monitoring procedure to obtain hemodynamic parameters for rats. Now, we adapted the procedure to acquire systemic and hepatic hemodynamic parameters in mice, a species ten-fold smaller than rats. This film demonstrates the instrumentation of the animals as well as the data acquisition process needed to assess systemic and hepatic hemodynamics in mice. Vital parameters, including body temperature, respiratory&#160;rate&#160;and heart&#160;rate were recorded throughout the whole procedure. Systemic hemodynamic parameters consist of carotid artery pressure (CAP) and central venous pressure (CVP). Hepatic perfusion parameters include portal vein pressure (PVP), portal flow rate as well as the flow rate of the common hepatic artery (table 1). Instrumentation and data acquisition to record the normal values was completed within 1.5 h. Systemic and hepatic hemodynamic parameters remained within normal ranges during this procedure.
This procedure is challenging but feasible. We have already applied this procedure to assess hepatic hemodynamics in normal mice as well as during 70% partial hepatectomy and in liver lobe clamping experiments. Mean PVP after resection (n= 20), was 11.41&#177;2.94 cmH2O which was significantly higher (P&#60;0.05) than before resection (6.87&#177;2.39 cmH2O). The results of liver lobe clamping experiment indicated that this monitoring procedure is sensitive and suitable for detecting small changes in portal pressure and portal flow rate. In conclusion, this procedure is reliable in the hands of an experienced micro-surgeon but should be limited to experiments where mice are absolutely needed.

This film demonstrates how to acquire systemic and hepatic hemodynamics in mice. The whole monitoring includes acquisition of vital parameters, systemic blood pressure, central venous pressure, common hepatic artery flow rate, and portal vein pressure as well as the portal flow rate in mice.

ניטור של פרמטרים מערכתיים והבדיה המודינמית בעכברים

The use of mouse models in experimental research is of enormous importance for the study of hepatic physiology and pathophysiological disturbances. ...

Utilizing Murine Inducible Telomerase Alleles in the Studies of Tissue Degeneration/Regeneration and Cancer

Telomere dysfunction-induced loss of genome integrity and its associated DNA damage signaling and checkpoint responses are well-established drivers that cause tissue degeneration during ageing. Cancer, with incidence rates greatly increasing with age, is characterized by short telomere lengths and high telomerase activity. To study the roles of telomere dysfunction and telomerase reactivation in ageing and cancer, the protocol shows how to generate two murine inducible telomerase knock-in alleles 4-Hydroxytamoxifen (4-OHT)-inducible TERT-Estrogen Receptor (mTERT-ER) and Lox-Stopper-LoxTERT (LSL-mTERT). The protocol describes the procedures to induce telomere dysfunction and reactivate telomerase activity in mTERT-ER and LSL-mTERT mice in vivo. The representative data show that reactivation of telomerase activity can ameliorate the tissue degenerative phenotypes induced by telomere dysfunction. In order to determine the impact of telomerase reactivation on tumorigenesis, we generated prostate tumor model G4 PB-Cre4 PtenL/L p53L/L LSL-mTERTL/L and thymic T-cell lymphoma model G4 Atm-/- mTERTER/ER. The representative data show that telomerase reactivation in the backdrop of genomic instability induced by telomere dysfunction can greatly enhance tumorigenesis. The protocol also describes the procedures used to isolate neural stem cells (NSCs) from mTERT-ER and LSL-mTERT mice and reactivate telomerase activity in NSCs in vitro. The representative data show that reactivation of telomerase can enhance the self-renewal capability and neurogenesis in vitro. Finally, the protocol describes the procedures for performing telomere FISH (Fluorescence In Situ Hybridization) on both mouse FFPE (Formalin Fixed and Paraffin Embedded) brain tissues and metaphase chromosomes of cultured cells.

Telomere and telomerase play essential roles in ageing and tumorigenesis. The goal of this protocol is to show how to generate two murine inducible telomerase knock-in alleles and how to utilize them in the studies of tissue degeneration/regeneration and cancer.

ניצול Murine עין מתנהל הטלומרז אללים במחקרים של ניוון רקמה / התחדשות והסרטן

Telomere dysfunction-induced loss of genome integrity and its associated DNA damage signaling and checkpoint responses are well-established drivers that ...

Induction of Accelerated Atherosclerosis in Mice: The "Wire-Injury" Model

Atherosclerosis is a proliferative fibro-inflammatory disease developing in the arterial wall, inducing a deficient blood flow or a lack of blood flow. Moreover, by rupture of the defective vascular wall, atherosclerosis induces occlusive thrombus formation, which represents the main cause of myocardial infarction or stroke and the most frequent cause of death. Despite the advances in the cardiovascular field, many questions remain unanswered, and additional basic research is essential to improve our understanding of the molecular mechanisms during atherosclerosis and its effects. Due to limited clinical studies, there is a need for representative animal models recreating atherosclerotic conditions such as neointima formation after stent implantation, balloon angioplasty, or endarterectomy. Since the mouse presents many advantages and is the most frequently used model for studying molecular processes, the current study proposes an invasive procedure of endothelial denudation, also known as the wire-injury model, which is representative of the human condition of neointima formation in arteries after revascularization procedures.

Induction of Accelerated Atherosclerosis in Mice: The &quot;Wire-Injury&quot; Model

This study describes an invasive procedure for the induction of accelerated atherosclerosis in mice. In comparison to other methods using electric- or cryo-induced injury, mechanical-induced injury mimics the human condition of restenosis after revascularization therapies and is ideal for the study of the molecular mechanisms involved.

אינדוקציה של טרשת עורקים מואצת בעכברים: מודל "פגיעת חוט"

Atherosclerosis is a proliferative fibro-inflammatory disease developing in the arterial wall, inducing a deficient blood flow or a lack of blood flow. ...

The Pilocarpine Model of Temporal Lobe Epilepsy and EEG Monitoring Using Radiotelemetry System in Mice

Temporal lobe epilepsy (TLE) is a common neurological disorder in adulthood. For translational studies of chronic epilepsy, pilocarpine-induced status epilepticus (SE) is frequently selected to recapitulate spontaneous recurrent seizures (SRS). Here we present a protocol of SE induction by intraperitoneal (i.p.) injection of pilocarpine and monitoring of chronic recurring seizures in live animals using a wireless telemetry video and electroencephalogram (EEG) system. We demonstrated notable behavioral changes that need attention after pilocarpine injection and their correlation with hippocampal neuronal loss at 7 days and 6 weeks post-pilocarpine. We also describe the experimental procedures of electrode implantation for video and EEG recording, and analysis of the frequency and duration of chronic recurrent seizures. Finally, we discuss the possible reasons why the expected results are not achieved in each case. This provides a basic overview of modeling chronic epilepsy in mice and guidelines for troubleshooting. We believe this protocol can serve as a baseline for suitable models of chronic epilepsy and epileptogenesis.

This manuscript describes a method of inducing status epilepticus by systemic pilocarpine injection and of monitoring spontaneous recurrent seizures in live animals using a wireless telemetry video and electroencephalogram system. This protocol can be utilized for studying the pathophysiologic mechanisms of chronic epilepsy, epileptogenesis, and acute seizures.

המודל פילוקרפין של אפילפסיה של אונה רקתית EEG ניטור באמצעות מערכת Radiotelemetry בעכברים

Temporal lobe epilepsy (TLE) is a common neurological disorder in adulthood. For translational studies of chronic epilepsy, pilocarpine-induced status ...

Murine Distal Colostomy, A Novel Model of Diversion Colitis in C57BL/6 Mice

Diversion colitis (DC) is a frequent clinical condition occurring in patients with bowel segments excluded from the fecal stream as a result of a diverting enterostomy. The etiology of this disease remains ill-defined but appears to differ from that of classical inflammatory bowel diseases such as Crohn's disease and ulcerative colitis. Research aimed to decipher the pathophysiological mechanisms leading to the development of this disease has been severely hampered by the lack of an appropriate murine model. This protocol generates a murine model of DC that facilitates the study of the immune system's role and its interaction with the microbiome in the development of DC. In this model using C57BL/6 animals, distal parts of the colon are excluded from the fecal stream by creating a distal colostomy, triggering the development of mild to moderate inflammation in the excluded bowel segments and reproducing the hallmark lesions of human DC with a moderate systemic inflammatory response. In contrast to the rat model, a large number of genetically-modified murine models on the C57BL/6 background are available. The combination of these animals with our model allows the potential roles of individual cytokines, chemokines, or receptors of bioactive molecules (e.g., interleukin (IL)-17; IL-10, chemokine CXCL13, chemokine receptors CXCR5 and CCR7, and the sphingosine-1-phosphate receptor 4) to be assessed in the pathogenesis of DC. The availability of congenic mouse strains on the C57BL/6 background largely facilitates transfer experiments to establish the roles of distinct cell types involved in the etiology of DC. Finally, the model offers the opportunity to assess the influences of local interventions (e.g., modification of the local microbiome or local anti-inflammatory therapy) on mucosal immunity in affected and non-affected bowel segments and the on systemic immune homeostasis.

Murine distal colostomy provides a murine model for human diversion colitis, a predominantly lymphocytic colitis in colon segment excluded from the fecal stream.

מאתר למעי הגס הדיסטלי, מודל הרומן של הסחה קוליטיס בעכברים C57BL/6

Diversion colitis (DC) is a frequent clinical condition occurring in patients with bowel segments excluded from the fecal stream as a result of a ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

Contractility איזומטרי מדידה של עורק מצע המעי העליון העכבר באמצעות חוט Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...