Name: methods for intravenous self administration in a mouse model
Uploaded: 2012-12-08T14:00:00
Duration: 12 min 9 s
Description: Watch this Scientific Journal Video about Methods for Intravenous Self Administration in a Mouse Model at JoVE.com

जानवरों पर प्रयोग पशु की देखभाल और मैकगिल विश्वविद्यालय पशु देखभाल समिति के कनाडा परिषद द्वारा निर्धारित दिशा निर्देशों और नियमों के अनुसार में प्रदर्शन किया गया. इस शोध स्वास्थ्य अनुसंधान (CIHR) की कनाडा के संस्थानों से केजी के लिए सम्मानित किया गया धन के द्वारा समर्थित किया गया. ब्याज की कोई संघर्ष की घोषणा की.

<ol>
	<li>Rocha, B. A., Ator, R., Emmett-Oglesby, M. W., Hen, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Intravenous Cocaine Self-Administration in Mice Lacking 5-HT1B Receptors. Pharmacology Biochemistry and Behavior. 57 (3), 407-412 (1997).</li><li>Zapata, A., Chefer, V. I., Ator, R., Shippenberg, T. S., Rocha, B. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioural+sensitization+and+enhanced+dopamine+response+in+the+nucleus+accumbens+after+intravenous+cocaine+self-administration+in+mice.">Behavioural sensitization and enhanced dopamine response in the nucleus accumbens after intravenous cocaine self-administration in mice.</a> European Journal of Neuroscience. 17 (3), 590-596 (2003).</li><li>Pritchett-Corning, K. R., Mulder, G. B., Luo, Y., White, W. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Principles+of+Rodent+Surgery+for+the+New+Surgeon.">Principles of Rodent Surgery for the New Surgeon.</a> J. Vis. Exp. (47), e2586 (2011).</li><li>Hoogstraten-Miller, S. L., Brown, P. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Techniques in Aseptic Rodent Surgery Current Protocols in Immunology. , 1.12-1.14 (2001).</li><li>Thomsen, M., Caine, S. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+Intravenous+Drug+Self-Administration+in+Rats+and+Mice.">Chronic Intravenous Drug Self-Administration in Rats and Mice.</a> Current Protocols in Neuroscience. . 9, 20.1-20.40 (2005).</li><li>Gill, K. J., Boyle, A. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Confirmation+of+quantitative+trait+loci+for+cocaine+induced+activation+in+the+AcB/BcA+series+of+recombinant+congenic+strains.">Confirmation of quantitative trait loci for cocaine induced activation in the AcB/BcA series of recombinant congenic strains.</a> Pharmacogenetics. 13, 329-338 (2003).</li><li>Gill, K. J., Boyle, A. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+influences+on+drug-induced+psychomotor+activation+in+mice.">Genetic influences on drug-induced psychomotor activation in mice.</a> Genes, Brain &#38; Behavior. 7, 859-868 (2008).</li><li>Boyle, A. E., Gill, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sensitivity+of+AXB/BXA+recombinant+inbred+lines+of+mice+to+the+locomotor+activating+effects+of+cocaine:+A+QTL+analysis.">Sensitivity of AXB/BXA recombinant inbred lines of mice to the locomotor activating effects of cocaine: A QTL analysis.</a> Pharmacogenetics. 11 (3), 255-264 (2001).</li><li>Boyle, A. E., Gill, K. 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+verification+of+previously+identified+QTLs+for+cocaine-induced+activation+using+a+panel+of+B6.A+chromosome+substitution+strains+(CSS)+and+A/JxC57Bl/6J+F2+mice.">A verification of previously identified QTLs for cocaine-induced activation using a panel of B6.A chromosome substitution strains (CSS) and A/JxC57Bl/6J F2 mice.</a> Psychopharmacology. 207 (2), 325-3234 (2009).</li><li>Thomsen, M., Caine, S. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Psychomotor+stimulant+effects+of+cocaine+in+rats+and+15+mouse+strains.">Psychomotor stimulant effects of cocaine in rats and 15 mouse strains.</a> Experimental and Clinical Psychopharmacology. 19 (5), 321-341 (2011).</li><li>Thomsen, M., Caine, S. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intravenous+drug+self-administration+in+mice:+practical+considerations.">Intravenous drug self-administration in mice: practical considerations.</a> Behavior genetics. 37 (1), 101-118 (2007).</li></ol>

ब्याज की कोई संघर्ष की घोषणा की.

मादक पदार्थों के सेवन के पशु मॉडल मादक पदार्थों से संबंधित व्यवहार के आनुवंशिक आधार को समझने में विशेष रूप से उपयोगी हैं. उदाहरण के लिए, विभिन्न आनुवंशिक प्रोफाइल के साथ चूहों उनके 6-8 कोकीन के लिए संवेदनशीलता में मतभेद दिखाने पैतृक और पहचान करने की क्षमता जीन उम्मीदवारों mediating प्ररूपी परिवर्तनशीलता मनाया 9 मदद. अंतःशिरा कैथीटेराइजेशन इस पत्र में वर्णित प्रक्रियाओं काफी सफलता के साथ इस्तेमाल किया गया है के रूप में के रूप में अच्छी तरह से चूहों विभिन्न आनुवंशिक पृष्ठभूमि 10 के चूहों के विभिन्न प्रकारों में दवा IVSA की जांच. इस वीडियो में दिखाया प्रक्रियाओं के दौरान पर ध्यान केंद्रित करने के लिए महत्वपूर्ण कारकों पर प्रकाश डाला और catherization सर्जरी के बाद के क्रम में विश्वसनीय नसों में आत्म - प्रशासन डेटा प्राप्त. सबसे पहले, सही atrium के भीतर कैथेटर टयूबिंग की नियुक्ति के लिए महत्वपूर्ण है, क्रम में दाग के थक्कों से कैथेटर विफलता को रोकने के लिए. शल्य चिकित्सा के दौरान यह सुनिश्चित करने के लिए महत्वपूर्ण है कि कैथेटरअंत बेरोक है, और या तो दिल या नस ऊतक द्वारा occluded नहीं. दूसरा, दैनिक कैथेटर निस्तब्धता दोनों के पहले और बाद में चालू सत्र के क्रम में रुकावटों को रोकने की आवश्यकता है. अंत में, कैथेटर प्रवेशनी दोनों प्रवेशनी कैप और क्रिस्टल को शामिल किया गया है जब पशुओं के लिए उनके घर के पिंजरों में हैं के साथ लगातार कवर चाहिए, मलबे के प्रवेश को रोकने के लिए. कैथेटर प्रवेशनी के मामूली रुकावट एक ठीक 26 गेज सुई का उपयोग करते हुए, लेकिन दैनिक निस्तब्धता दिनों जब जानवरों IVSA के लिए परीक्षण नहीं कर रहे पर के साथ heparinized एंटीबायोटिक समाधान की आवश्यकता है, विशेष रूप से उखाड़ फेंकना जा सकता है. पशु अस्तित्व सर्जरी, सड़न रोकनेवाला तकनीक का अच्छा ज्ञान प्रदर्शन, analgesia और संज्ञाहरण आवश्यक है. जबकि इस वीडियो उचित शल्य चिकित्सा प्रशिक्षण की जगह नहीं है, यह इस प्रतिमान के लिए आवश्यक तकनीक हासिल करने के इच्छुक शोधकर्ताओं के लिए एक गाइड के रूप में इस्तेमाल किया जा सकता है.

<table border="1"><tbody><tr><td> अभिकर्मक के नाम </td><td> कंपनी </td><td> सूचीपत्र संख्या </td><td> टिप्पणियां (वैकल्पिक) </td></tr><tr><td> Ticarcillin Disodium नमक </td><td> सिग्मा Aldrich </td><td> T5639-1G </td><td> 1 ग्राम की बोतल </td></tr><tr><td> आँसू Naturale PM (नेत्र स्नेहक) </td><td> Alcon </td><td> N / A </td><td> 3.5 ग्राम ट्यूब यह आंख स्नेहक सबसे फार्मेसियों में पाया जा सकता है. कोई नेत्र आँख स्नेहक इस्तेमाल किया जा सकता है. </td></tr><tr><td> कोकीन </td><td> Medisca Pharmaceutique </td><td> 0723-06 </td><td> 1 ग्राम की बोतल </td></tr><tr><td> Ketamine </td><td> Medisca Pharmaceutique </td><td> 1754-04 </td><td> 25 ग्राम की बोतल </td></tr><tr><td> Midazolam </td><td> Medisca Pharmaceutique </td><td> 2519-06 </td><td> 1 ग्राम की बोतल </td></tr><tr><td> Heparin सोडियम नमक </td><td> सिग्मा Aldrich </td><td> H4784-250mg </td><td> 250 ग्राम की बोतल </td></tr><tr><td> Ketoprofen </td><td> सिग्मा Aldrich </td><td> K1751 - 1G </td><td> 1 ग्राम की बोतल </td></tr><tr><td> Amikicin सल्फेट नमक </td><td> सिग्मा Aldrich </td><td> A2324-5G </td><td> 5 ग्राम की बोतल </td></tr><tr><td> सुक्रोज ACS अभिकर्मक </td><td> सिग्मा Aldrich </td><td> S5016-500G </td><td> 500 ग्राम की बोतल </td></tr><tr><td> नरम जाल के साथ मानक माउस कैथेटर </td><td> CamCaths </td><td> MIVSA/20/26/BC-1S </td><td> टयूबिंग विनिर्देशों: 26SWG, 0.451, 0.254 </td></tr><tr><td> क्रिस्टल टोपी applicator </td><td> वैज्ञानिक HRS </td><td> 313CAC </td><td> यह सफेद कैथेटर टोपी है. </td></tr><tr><td> Tygon माइक्रो टयूबिंग बोर </td><td> थॉमस वैज्ञानिक </td><td> 9561S41 </td><td> टयूबिंग विनिर्देशों (बोर x आयुध डिपो इंच में x दीवार मोटाई): 0.01 x 0.03 x .01 </td></tr>
 </tbody>
</table>

methods for intravenous self administration in a mouse model

पशु मॉडल के लिए दवाओं के मजबूत प्रभाव नसों में आत्म - प्रशासन प्रतिमान (IVSA) सहित, का अध्ययन करने के लिए विकसित किया गया है. एक IVSA प्रतिमान का उपयोग करने के लिए कोकीन के रूप में इस तरह के दुरुपयोग की दवाओं के मजबूत गुणों का अध्ययन का लाभ तथ्य यह है कि दवा के बजाय आत्म प्रशासित experimenter प्रशासित सुदृढीकरण की अनुसूची बदल किया जा सकता है, और की मात्रा का सही माप शामिल दवा के रूप में के रूप में अच्छी तरह से भस्म समय और चतुर्थ इंजेक्शन के पैटर्न को प्राप्त किया जा सकता है. इसके अलावा, प्रशासन की नसों मार्ग पहले पारित चयापचय या स्वाद के लिए संबंधित संभावित घालमेल से बचा जाता है, और खून और दिमाग दवा के स्तर में तेजी से वृद्धि का उत्पादन. इस वीडियो के रूप में बताया गया है, नसों में आत्म - प्रशासन सर्जरी और सावधानीपूर्वक दैनिक कैथेटर के दौरान सावधान कैथेटर प्लेसमेंट निस्तब्धता और रखरखाव के बाद पूर्व खाद्य प्रतिबंध या पहले दवा प्रशिक्षण के बिना प्राप्त किया जा सकता है. प्रायोगिक इस में उल्लिखित प्रक्रियाओंकागज पशु आवास के विवरण और दशानुकूलन विधियों, स्फूर्त प्रशिक्षण मीठा दूध समाधान का उपयोग, और कैथिटर आरोपण सर्जरी शामिल हैं.

<img src="/files/ftp_upload/3739/3739fig1.jpg" alt="चित्रा 1" /> चित्रा 1 नसों में नशीली दवाओं के आत्म - प्रशासन के लिए जवाब देने के पैटर्न दवा, खुराक सीमा और माउस तनाव कार्यरत द्वारा अलग अलग होंगे. प्रस्तुत आंकड़ा कोकीन सफल catherization वीडियो में वर्णित प्रक्रिया का उपयोग सर्जरी के बाद आत्म - प्रशासन के आंकड़ों से पता चलता है. आंकड़ा (SEM ±) मतलब अर्जित कोकीन सुई लेनी और मतलब (SEM ±) कोकीन की खपत 4 कोकीन सुदृढीकरण के एक FR1 समय पर एक यादृच्छिक क्रम में प्रस्तुत खुराक की एक सीमा के पार (मिलीग्राम / किग्रा शरीर के वजन) से पता चलता है. भुज: जलसेक प्रति आत्म प्रशासित दवा की खुराक. वाम तालमेल: सुई लेनी की कुल संख्या 2 घंटा operant सत्र के दौरान अर्जित किए हैं. सही तालमेल: परीक्षण 2 घंटा सत्र के दौरान मिलीग्राम / किलो में कुल कोकीन सेवन. सभी 13 कैथेटर अध्ययन की अवधि (4 सप्ताह) के लिए पेटेंट बने रहे. एक एक तरह खुराक पर आयोजित की एनोवा से पता चला है कि micई एक खुराक पर निर्भर फैशन में कोकीन प्रशासन थे [(1,12) एफ = 42.8, पी &lt;0.05]. खुराक - प्रतिक्रिया वक्र (3,36) एफ = 29.6, पी 0.05 &lt;] लीवर अधिक मात्रा में दबाव में कमी के बावजूद अधिक कोकीन की खपत में वृद्धि हुई है. प्रत्येक डेटा बिंदु प्रत्येक कोकीन (SEM ±) खुराक सीडी 1 चूहों (n = 13 / खुराक, पुरुषों और महिलाओं संयुक्त) में एकत्र 3 परीक्षण सत्र के औसत का प्रतिनिधित्व करता है. सक्रिय (दवा प्रबलित) बनाम निष्क्रिय खुराक प्रतिक्रिया वक्र भर में जवाब लीवर की तुलना में दो तरह से एनोवा का उपयोग करने के लिए सुनिश्चित करें कि चूहों दो levers के बीच भेदभाव करने के लिए किए गए थे. CD1 चूहों के लिए सक्रिय लीवर के लिए एक प्राथमिकता [(1,12) एफ = 10.255, पी 0.05 &lt;] पूरी खुराक - प्रतिक्रिया वक्र (डेटा यहाँ दिखाया नहीं) पर विश्लेषण से पता चला

Watch this Scientific Journal Video about Methods for Intravenous Self Administration in a Mouse Model at JoVE.com

Methods for Intravenous Self Administration in a Mouse Model

नसों में आत्म - प्रशासन प्रतिमान (IVSA) कृन्तकों में दुरुपयोग की दवाओं के मजबूत गुणों की जांच में सोने के मानक माना जाता है. इस पांडुलिपि प्रयोगात्मक प्रक्रियाओं और शल्य चिकित्सा तकनीक के लिए विश्वसनीय IVSA डेटा प्राप्त करने के लिए आवश्यक रूपरेखा. विशेष रूप से, सावधानीपूर्वक कैथेटर आरोपण और रखरखाव डाला जाता है.

नसों में स्वयं प्रशासन के लिए एक माउस मॉडल में तरीके

Animal models have been developed to study the reinforcing effects of drugs, including the intravenous self-administration (IVSA) paradigm. The advantages ...

methods-for-intravenous-self-administration-in-a-mouse-model

Research

JoVE Journal

Neuroscience

Assessment of Ultrasonic Vocalizations During Drug Self-administration in Rats

Drug self-administration procedures are commonly used to study behavioral and neurochemical changes associated with human drug abuse, addiction and relapse. Various types of behavioral activity are commonly utilized as measures of drug motivation in animals. However, a crucial component of drug abuse relapse in abstinent cocaine users is "drug craving", which is difficult to model in animals, as it often occurs in the absence of overt behaviors. Yet, it is possible that a class of ultrasonic vocalizations (USVs) in rats may be a useful marker for affective responses to drug administration, drug anticipation and even drug craving. Rats vocalize in ultrasonic frequencies that serve as a communicatory function and express subjective emotional states. Several studies have shown that different call frequency ranges are associated with negative and positive emotional states. For instance, high frequency calls ("50-kHz") are associated with positive affect, whereas low frequency calls ("22-kHz") represent a negative emotional state. This article describes a procedure to assess rat USVs associated with daily cocaine self-administration. For this procedure, we utilized standard single-lever operant chambers housed within sound-attenuating boxes for cocaine self-administration sessions and utilized ultrasonic microphones, multi-channel recording hardware and specialized software programs to detect and analyze USVs. USVs measurements reflect emotionality of rats before, during and after drug availability and can be correlated with commonly assessed drug self-administration behavioral data such lever responses, inter-response intervals and locomotor activity. Since USVs can be assessed during intervals prior to drug availability (e.g., anticipatory USVs) and during drug extinction trials, changes in affect associated with drug anticipation and drug abstinence can also be determined. In addition, determining USV changes over the course of short- and long-term drug exposure can provide a more detailed interpretation of drug exposure effects on affective functioning.

Drug self-administration and ultrasonic vocalizations (USV) are used as behavioral assessments in animal research, but rarely in combination. The purpose of this article is to describe the advantages of recording USVs during drug self-administration procedures to assess affective responses to drug experience.

अल्ट्रासोनिक vocalizations चूहे में स्व - प्रशासन औषधि के दौरान मूल्यांकन

Drug self-administration procedures are commonly used to study behavioral and neurochemical changes associated with human drug abuse, addiction and ...

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.

एकाधिक माउस Neuroanatomical चुंबकीय अनुनाद इमेजिंग

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

Microdialysis of Ethanol During Operant Ethanol Self-administration and Ethanol Determination by Gas Chromatography

Operant self-administration methods are commonly used to study the behavioral and pharmacological effects of many drugs of abuse, including ethanol. However, ethanol is typically self-administered orally, rather than intravenously like many other drugs of abuse. The pharmacokinetics of orally administered drugs are more complex than intravenously administered drugs. Because understanding the relationship between the pharmacological and behavioral effects of ethanol requires knowledge of the time course of ethanol reaching the brain during and after drinking, we use in vivo microdialysis and gas chromatography with flame ionization detection to monitor brain dialysate ethanol concentrations over time. 
Combined microdialysis-behavioral experiments involve the use of several techniques. In this article, stereotaxic surgery, behavioral training and microdialysis, which can be adapted to test a multitude of self-administration and neurochemical centered hypotheses, are included only to illustrate how they relate to the subsequent phases of sample collection and dialysate ethanol analysis. Dialysate ethanol concentration analysis via gas chromatography with flame-ionization detection, which is specific to ethanol studies, is described in detail. Data produced by these methods reveal the pattern of ethanol reaching the brain during the self-administration procedure, and when paired with neurochemical analysis of the same dialysate samples, allows conclusions to be made regarding the pharmacological and behavioral effects of ethanol.

A method to determine the time course of ethanol concentration in the brains of rats during operant ethanol self-administration is described. Gas chromatography with flame ionization detection is used to quantify ethanol in the dialysate samples, because it has the sensitivity required for the small volumes that are generated.

इथेनॉल के गैस क्रोमैटोग्राफी द्वारा operant इथेनॉल आत्म - प्रशासन और इथेनॉल के दृढ़ संकल्प के दौरान microdialysis

Operant self-administration methods are commonly used to study the behavioral and pharmacological effects of many drugs of abuse, including ethanol.  ...

Methods for Studying the Mechanisms of Action of Antipsychotic Drugs in Caenorhabditis elegans

Caenorhabditis elegans is a simple genetic organism amenable to large-scale forward and reverse genetic screens and chemical genetic screens. The C. elegans genome includes potential antipsychotic drug (APD) targets conserved in humans, including genes encoding proteins required for neurotransmitter synthesis and for synaptic structure and function. APD exposure produces developmental delay and/or lethality in nematodes in a concentration-dependent manner. These phenotypes are caused, in part, by APD-induced inhibition of pharyngeal pumping1,2. Thus, the developmental phenotype has a neuromuscular basis, making it useful for pharmacogenetic studies of neuroleptics. Here we demonstrate detailed procedures for testing APD effects on nematode development and pharyngeal pumping. For the developmental assay, synchronized embryos are placed on nematode growth medium (NGM) plates containing APDs, and the stages of developing animals are then scored daily. For the pharyngeal pumping rate assay, staged young adult animals are tested on NGM plates containing APDs. The number of pharyngeal pumps per unit time is recorded, and the pumping rate is calculated. These assays can be used for studying many other types of small molecules or even large molecules.

Methods for Studying the Mechanisms of Action of Antipsychotic Drugs in Caenorhabditis elegans

Approaches for testing the effects of antipsychotic drugs (APDs) in Caenorhabditis elegans are demonstrated. Assays are described for testing drug effects on development and viability and on pharyngeal pumping rate. These methods are also applicable for pharmacogenetic experiments with drug classes other than APDs.

में antipsychotic दवाओं की कार्रवाई के तंत्र के अध्ययन के लिए तरीके<em&gt; Caenorhabditis एलिगेंस</em

Caenorhabditis elegans is a simple genetic organism amenable to  large-scale forward and reverse genetic screens and chemical genetic screens. The C.  ...

The Corneal Micropocket Assay: A Model of Angiogenesis in the Mouse Eye

The mouse corneal micropocket assay is a robust and quantitative in vivo assay for evaluating angiogenesis. By using standardized slow-release pellets containing specific growth factors that trigger blood vessel growth throughout the naturally avascular cornea, angiogenesis can be measured and quantified. In this assay the angiogenic response is generated over the course of several days, depending on the type and dose of growth factor used. The induction of neovascularization is commonly triggered by either basic fibroblast growth factor (bFGF) or vascular endothelial growth factor (VEGF). By combining these growth factors with sucralfate and hydron (poly-HEMA (poly(2-hydroxyethyl methacrylate))) and casting the mixture into pellets, they can be surgically implanted in the mouse eye. These uniform pellets slowly-release the growth factors over five or six days (bFGF or VEGF respectively) enabling sufficient angiogenic response required for vessel area quantification using a slit lamp. This assay can be used for different applications, including the evaluation of angiogenic modulator drugs or treatments as well as comparison between different genetic backgrounds affecting angiogenesis. A skilled investigator after practicing this assay can implant a pellet in less than 5 min per eye.

The protocol describes the corneal micropocket assay as developed in mice.

कॉर्नियल Micropocket परख: माउस आँखों में angiogenesis की एक मॉडल

The mouse corneal micropocket assay is a robust and quantitative in vivo assay for evaluating angiogenesis. By using standardized slow-release pellets ...

Immunohistochemical and Calcium Imaging Methods in Wholemount Rat Retina

In this paper we describe the tools, reagents, and the practical steps that are needed for: 1) successful preparation of wholemount retinas for immunohistochemistry and, 2) calcium imaging for the study of voltage gated calcium channel (VGCC) mediated calcium signaling in retinal ganglion cells. The calcium imaging method we describe circumvents issues concerning non-specific loading of displaced amacrine cells in the ganglion cell layer.

Immunohistochemistry protocols are used to study the localization of a specific protein in the retina. Calcium imaging techniques are employed to study calcium dynamics in retinal ganglion cells and their axons.

Wholemount चूहा रेटिना में immunohistochemical और कैल्शियम इमेजिंग तरीकों

In this paper we describe the tools, reagents, and the practical steps that are needed for: 1) successful preparation of wholemount retinas for ...

Methods for Cell-attached Capacitance Measurements in Mouse Adrenal Chromaffin Cell

Neuronal transmission is an integral part of cellular communication within the brain. Depolarization of the presynaptic membrane leads to vesicle fusion known as exocytosis that mediates synaptic transmission. Subsequent retrieval of synaptic vesicles is necessary to generate new neurotransmitter-filled vesicles in a process identified as endocytosis. During exocytosis, fusing vesicle membranes will result in an increase in surface area and subsequent endocytosis results in a decrease in the surface area. Here, our lab demonstrates a basic introduction to cell-attached capacitance recordings of single endocytic events in the mouse adrenal chromaffin cell. This type of electrical recording is useful for high-resolution recordings of exocytosis and endocytosis at the single vesicle level. While this technique can detect both vesicle exocytosis and endocytosis, the focus of our lab is vesicle endocytosis. Moreover, this technique allows us to analyze the kinetics of single endocytic events. Here the methods for mouse adrenal gland tissue dissection, chromaffin cell culture, basic cell-attached techniques, and subsequent examples of individual traces measuring singular endocytic event are described.

After exocytosis, fused plasma membrane is retrieved through a process known as endocytosis. This mechanism reforms new synaptic vesicles for the next round of release. Individual endocytic events are captured and analyzed through the use of the cell-attached capacitance recordings in mouse adrenal chromaffin cells.

माउस अधिवृक्क क्रोमाफिन सेल में सेल संलग्न समाई माप के लिए तरीके

Neuronal transmission is an integral part of cellular communication within the brain. Depolarization of the presynaptic membrane leads to vesicle fusion ...

Neurobehavioral Assessments in a Mouse Model of Neonatal Hypoxic-ischemic Brain Injury

We performed unilateral carotid artery occlusion on CD-1 mice to create a neonatal hypoxic-ischemic (HI) model and investigated the effects of neonatal HI brain injury by studying neurobehavioral functions in these mice compared to non-operated (i.e., normal) mice. During the study, Rice-Vannucci&#39;s method was used to induce neonatal HI brain damage in postnatal day 7-10 (P7-10) mice. The HI operation was performed on the pups by unilateral carotid artery ligation and exposure to hypoxia (8% O2 and 92% N2 for 90 min). One week after the operation, the damaged brains were evaluated with the naked eye through the semi-transparent skull and were categorized into subgroups based on the absence (&#34;no cortical injury&#34; group) or presence (&#34;cortical injury&#34; group) of cortical injury, such as a lesion in the right hemisphere. On week 6, the following neurobehavioral tests were performed to evaluate the cognitive and motor functions: passive avoidance task (PAT), ladder walking test, and grip strength test. These behavioral tests are helpful in determining the effects of neonatal HI brain injury and are used in other mouse models of neurodegenerative diseases. In this study, neonatal HI brain injury mice showed motor deficits that corresponded to right hemisphere damage. The behavioral test results are relevant to the deficits observed in human neonatal HI patients, such as cerebral palsy or neonatal stroke patients. In this study, a mouse model of neonatal HI brain injury was established and showed different degrees of motor deficits and cognitive impairment compared to non-operated mice. This work provides basic information on the HI mouse model. MRI images demonstrate the different phenotypes, separated according to the severity of brain damage by motor and cognitive tests.

We performed unilateral carotid artery occlusion on postnatal day 7-10 CD-1 mouse pups to create a neonatal hypoxic-ischemic (HI) model and investigated the effects of HI brain injury. We studied neurobehavioral functions in these mice compared to non-operated normal mice.

नवजात Hypoxic के एक माउस मॉडल में Neurobehavioral आकलन-कोरोनरी मस्तिष्क चोट

We performed unilateral carotid artery occlusion on CD-1 mice to create a neonatal hypoxic-ischemic (HI) model and investigated the effects of neonatal HI ...

Methods and Tips for Intravenous Administration of Adeno-associated Virus to Rats and Evaluation of Central Nervous System Transduction

Adeno-associated virus (AAV) vectors are a key reagent in the neurosciences for clustered regularly interspaced short palindromic repeats (CRISPR), optogenetics, cre-lox targeting, etc. The purpose of this manuscript is to aid the investigator attempting expansive central nervous system (CNS) gene transfer in the rat via tail vein injection of AAV. Wide-scale expression is relevant for conditions with widespread pathology, and a rat model is significant due to its greater size and physiologic similarities to humans compared to mice. In this example application, a wide-scale neuronal transduction is used to mimic a neurodegenerative disease that affects the entire spinal cord, amyotrophic lateral sclerosis (ALS). The efficient wide-scale CNS transduction can also be used to deliver therapeutic protein factors in pre-clinical studies. After a post-injection expression interval of several weeks, the effects of the transduction are evaluated. For a green fluorescent protein (GFP) control vector, the amount of GFP in the cerebellum is estimated quickly and reliably by a basic imaging program. For motor disease phenotypes that are induced by the ALS related protein transactive response DNA-binding protein of 43 kDa (TDP-43), the deficits are scored by escape reflex and rotarod. Beyond disease modeling and gene therapy, there are diverse potential applications for the wide-scale gene targeting described here. The expanded use of this method will aid in expediting hypothesis testing in the neurosciences and neurogenetics.

Methods for a wide-scale central nervous system gene delivery in the rat are covered. In this example, the purpose is to mimic a disease that affects the entire spinal cord. The widespread transduction can be used to deliver a therapeutic protein to the CNS from a one-time, peripheral administration.

तरीकों और Adeno के नसों में प्रशासन के लिए युक्तियां-चूहों और केंद्रीय तंत्रिका तंत्र Transduction के मूल्यांकन से जुड़े वायरस

Adeno-associated virus (AAV) vectors are a key reagent in the neurosciences for clustered regularly interspaced short palindromic repeats (CRISPR), ...

Assessment of Blood-brain Barrier Permeability by Intravenous Infusion of FITC-labeled Albumin in a Mouse Model of Neurodegenerative Disease

Disruption of blood-brain barrier (BBB) integrity is a common feature for different neurological and neurodegenerative diseases. Although the interplay between perturbed BBB homeostasis and the pathogenesis of brain disorders needs further investigation, the development and validation of a reliable procedure to accurately detect BBB alterations may be crucial and represent a useful tool for potentially predicting disease progression and developing targeted therapeutic strategies.
Here, we present an easy and efficient procedure for evaluating BBB leakage in a neurodegenerative condition like that occurring in a preclinical mouse model of Huntington disease, in which defects in the permeability of BBB are clearly detectable precociously in the disease. Specifically, the high molecular weight fluorescein isothiocyanate labelled (FITC)-albumin, which is able to cross the BBB only when the latter is impaired, is acutely infused into a mouse jugular vein and its distribution in the vascular or parenchymal districts is then determined by fluorescence microscopy.
Accumulation of green fluorescent-albumin in the brain parenchyma functions as an index of aberrant BBB permeability and, when quantitated by using Image J processing software, is reported as Green Fluorescence Intensity.

In this study, we present an easy and efficient procedure for evaluating the disruption of the blood-brain barrier under neurodegenerative conditions. To achieve our objective, we infused high molecular weight fluorescein isothiocyanate labelled (FITC)-albumin into mouse jugular vein and evaluated its leakage into the brain parenchyma by fluorescence microscopy.

Neurodegenerative रोग के एक माउस मॉडल में FITC-लेबल एल्ब्युमिन की नसों में अर्क से रक्त मस्तिष्क बाधा पारगम्यता का आकलन

Disruption of blood-brain barrier (BBB) integrity is a common feature for different neurological and neurodegenerative diseases. Although the interplay ...