Diese Studie wurde von der Ophthalmic Research Institute of Australia (Oria) unterstützt. Wir danken Prof. Algis Vingrys und Dr. Bang Bui, University of Melbourne, für zunächst uns helfen, die VEP Aufnahmetechnik zu entwickeln.

Ethics Hinweise: Alle Verfahren, die Tiere wurden in Übereinstimmung mit dem australischen Code of Practice für die Pflege und Verwendung von Tieren zu wissenschaftlichen Zwecken und den Richtlinien der ARVO Erklärung für die Nutzung von Tieren in Ophthalmic und Vision Research durchgeführt und wurden durch die genehmigte Tierethikkommission der Macquarie University. 1. VEP Elektrodenimplantation <ol><li> Betäuben das Tier mit einer intraperitonealen Injektion von Ketamin (75 mg / kg) und Medetomidin (0,5 mg / kg). Hinweis: Nach der Einleitung der Narkose, beachten Sie die Entzugs Reflexe (Pinch-Test, der Hornhaut und der Augenlidreflex, etc.) und ihre Abwesenheit als Indikation zur Operation zu beginnen. Tiere ständig in der gesamten Operation zu überwachen und zu verwalten zusätzliche Anästhetika (10% vom Ausgangs Ketamin Dosis pro top-up), wenn die Reflexe vorhanden sind. Adulte Ratten (&gt; 12 Wochen) werden in den Experimenten verwendet. </li><li> Rasur die Haut des OP-Bereich.Legen Sie das Tier auf eine wärmende Unterlage (37 ° C), um die Körpertemperatur während der Operation aufrecht zu erhalten. Bereiten Sie die Haut durch topische Anwendung von PVP-Jod. Bewerben Chirurgisches Tuch. Gelten topische Augensalbe zur Hornhaut Trockenheit unter Vollnarkose zu vermeiden. Pflegen Keimfreiheit, indem sie sterile Instrumente. </li><li> Einen Längshautschnitt auf der Mittellinie der Kopfhaut. Deaktivieren Sie das Bindegewebe um gute Belichtung des Schädels zu erreichen. </li><li> Vorsichtig bohren kleine Bohrlöcher manuell mit einem Mikrohandbohrmaschine auf 7 mm hinter dem Bregma und 3 mm lateral der Mittellinie. </li><li> Implantatschraube Elektroden durch den Schädel in die Rinde (Bereich 17), Durchdringen der Kortikalis zu einer Tiefe von etwa 0,5 mm. Implantat eine Referenzelektrode Schraube auf der Mittellinie 3 mm rostral des Bregma. Bewerben Zahnzement zu umhüllen und fixieren Sie die Schrauben (nicht immer erforderlich). </li><li> Naht der Haut des Kopfes, zu verwalten antibiotische Salbe auf die Haut und lassen Sie das Tier recover aus der Narkose auf einer Erwärmung pad. Hinweis: Alternativ können Elektroden können dem Tageslicht ausgesetzt werden, so dass die Haut muss nicht in jeder Aufzeichnungs wieder geöffnet werden. Unmittelbar nach Beendigung der Operation, aber vor dem Aufwachraum, verwalten ein nicht-steroidale Entzündungshemmer (NSAID) (wenn nicht vor der Operation verabreicht wird) oder ein Opioid-Analgetikum. Überwachen Sie die Tiere ständig bis zur vollständigen Wiederherstellung nach Betäubung und ambulant voll. </li><li> Lassen Sie mindestens 1 Woche für die Tiere von der Operation vor der VEP Aufnahme erholen. </li></ol> 2. Optic Nerve Injection <ol><li> Betäuben das Tier, bereiten die Haut und wenden drapieren wie oben beschrieben (1.1 und 1.2). </li><li> Einen 1- bis 1,5-cm-Einschnitt in der Haut oberhalb der Umlaufbahn von einer zufällig ausgewählten Auge. Öffnen Sie das Unterhautgewebe, um die Augenhöhle mit feinen Iris Schere zu erreichen. Öffnen Sie die Bindehaut und vorderen Tenon-Kapsel unter dem Operationsmikroskop. </li><li> Ziehen Sie die äußeren AugenMuskeln und intraobital Tränendrüsen auf ca. 3 mm Länge des Sehnervs freizulegen. Öffnen Sie die Dura und Arachnoidea Angelegenheit Schichten um den Sehnerv in Längsrichtung mit einer ophthalmischen Klinge. </li><li> Legen Sie die Glaspipette in den Sehnerv in einem Abstand von 2 mm hinter der ganzen Welt. Die Glas-Mikropipette wird in eine Hamilton-Spritze befestigt ist. </li><li> Injizieren 1% Lysolecithin (0,4-1,0 ul, mit 0,02% Evans Blau, das keine Auswirkungen auf die Myelinisierung) langsam in das Nerven ungefähr über einen Zeitraum von 30 sec. </li><li> Naht der Hautschnitt. Gelten antibiotische Salbe um Infektionen zu vermeiden. Partneraugen können als interne Kontrollen für die Elektrophysiologie Aufnahmen serviert. </li><li> Legen Sie die Tiere auf einer Erwärmung pad sich von der Anästhesie zu erholen. </li></ol> 3. VEP Recording <ol><li> Betäuben das Tier und bereiten die Haut wie 1.1 und 1.2. Hinweis: Niederdosis Narkosemittel kann für elektrophysiologische reco verwendet werdenrding (Ketamin 40 mg / kg und Medetomidin 0,25 mg / kg). </li><li> Legen Sie die Ratte in einem dunklen Raum und lassen Sie ihn in die Dunkelheit für 5 anzupassen - 30 min. In einigen Fällen können Ratten jeweils dunkel angepasst O / N für scotopic oder Licht zur photopischen VEP Aufnahmen 8 angepasst werden. </li><li> Aufrechterhaltung der Körpertemperatur auf 37 ± 0,5 ° C durch die homoeothermic Tuchsystem mit einer rektalen Temperaturfühler. </li><li> Erweitern die Schüler mit 1,0% Tropicamid Augentropfen. Öffnen Sie die Haut über den Schädel, um auf die vorge platziert in situ Schraube Elektroden. </li><li> Schließen Sie die Schraube über der kontralateralen visuellen Kortex des stimulierten Auge und dem Referenzschraube an den Verstärker. Legen Sie eine Nadelelektrode in den Schwanz als die Erde. Messen Sie und pflegen die Elektrodenimpedanz unter 5 kOhm. </li><li> Legen Sie eine Mini-Ganzfeld-Stimulator direkt auf der Haut um die Augenlider, um überlegene Augen Isolierung 7 bereitzustellen. Die Beleuchtung der Stimulator muss vorher kalibriert werdenmit einem Photometer. </li><li> Liefern Fotostimulation durch Lichtblitze 100 mal mit einer Frequenz von 1 Hz, bei niedrigen und hohen Bandpassfiltereinstellungen von 1 bis 100 Hz. Das Signal Abtastrate bei 5 kHz. Gilt: Signale sollte mindestens bei ca. 250 abgetastet werden - 300 Hz, um sicherzustellen, dass mehr als zwei Proben werden während eines jeden Zyklus gesammelt. </li><li> Naht der Haut zurück und die Tiere zu halten auf einer Erwärmung pad sich von der Anästhesie zu erholen. Aufzeichnungs kann wiederholt auf einzelne Tiere aufgezeichnet werden, um funktionelle Veränderungen über einen Zeitraum zu überwachen. </li><li> Am Endpunkt, eine Überdosis zu verabreichen Injektion von Natrium-Pentobarbital (100 mg / kg, ip), um das Tier einschläfern. Bestätigen Sie die Euthanasie durch Herzstillstand, Atemstillstand und eine Abnahme der Körpertemperatur. </li></ol> 4. Gewebepräparation und Histologie <ol><li> Entfernen Sie die Sehnerven von eingeschläfert Tiere unter dem Mikroskop und befestigen Sie das Gewebe in 1% Paraformaldehyd O / N. </li> &lt;li&gt; gründlich waschen das Gewebe mit Kochsalzlösung. Behandeln Sie das Gewebe in einem automatischen Gewebeprozessor und betten in Paraffin. Geschnittenen Abschnitte (5-10 &amp; mgr; m) unter Verwendung eines Rotationsmikrotom. Hinweis: Für die Immunhistochemie Studie beheben Gewebe in 1% Paraformaldehyd, wasche mit Kochsalzlösung und Inkubation mit 30% Saccharose O / N. Betten Sie Gewebe in OCT Einbettmedium und machen Gefrierschnitten unter Verwendung eines Kryostaten. </li><li> Abschnitte in 0,1% fast blue Lösung wie Luxol (in 95% Ethanol) O / N Inkubieren bei 56 ° C. Differenzieren Sie die Abschnitte in 0,05% Lithiumcarbonat für 30 Sekunden und dann 70% Ethanol für weitere 30 sec. Schließlich in 0,1% Cresylviolett Lösung Gegenfärbung 30 Sekunden vor der Montage der Teile. Nutzen Sie die schnelle Blau-Färbung, um Myelin in den Sehnerv 5 zu identifizieren. </li></ol>

<ol>
	<li>Balcer, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+practice.+Optic+neuritis.">Clinical practice. Optic neuritis.</a> N Engl J Med. 354 (12), 1273-1280 (2006).</li><li>Lassmann, H., Waxman, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Multiple sclerosis as a neuronal disease. , 153-164 (2005).</li><li>Fahle, M., Bach, M., Heckenlively, J., Arden, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Principles and practice of clinical electrophysiology of vision. , 207-234 (2006).</li><li>Halliday, A. M., McDonald, W. I., Mushin, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Delayed+visual+evoked+response+in+optic+neuritis.">Delayed visual evoked response in optic neuritis.</a> Lancet. 1, 982-985 (1972).</li><li>You, Y., Klistorner, A., Thie, J., Graham, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Latency+delay+of+visual+evoked+potential+is+a+real+measurement+of+demyelination+in+a+rat+model+of+optic+neuritis.">Latency delay of visual evoked potential is a real measurement of demyelination in a rat model of optic neuritis.</a> Invest Ophthalmol Vis Sci. 52 (9), 6911-6918 (2011).</li><li>You, Y., Klistorner, A., Thie, J., Gupta, V. K., Graham, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Axonal+loss+in+a+rat+model+of+optic+neuritis+is+closely+correlated+with+visual+evoked+potential+amplitudes+using+electroencephalogram+based+scaling.">Axonal loss in a rat model of optic neuritis is closely correlated with visual evoked potential amplitudes using electroencephalogram based scaling.</a> Invest Ophthalmol Vis Sci. 53, 3662 (2012).</li><li>You, Y., Klistorner, A., Thie, J., Graham, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improving+reproducibility+of+VEP+recording+in+rats:+electrodes,+stimulus+source+and+peak+analysis.">Improving reproducibility of VEP recording in rats: electrodes, stimulus source and peak analysis.</a> Doc Ophthalmol. 123 (2), 109-119 (2011).</li><li>Heiduschka, P., Schraermeyer, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+visual+function+in+pigmented+and+albino+rats+by+electroretinography+and+visual+evoked+potentials.">Comparison of visual function in pigmented and albino rats by electroretinography and visual evoked potentials.</a> Graefes Arch Clin Exp Ophthalmol. 246 (11), 1559-1573 (2008).</li><li>You, Y., Thie, J., Klistorner, A., Gupta, V. K., Graham, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Normalization+of+visual+evoked+potentials+using+underlying+electroencephalogram+levels+improves+amplitude+reproducibility+in+rats.">Normalization of visual evoked potentials using underlying electroencephalogram levels improves amplitude reproducibility in rats.</a> Invest Ophthalmol Vis Sci. 53 (3), 1473-1478 (2012).</li><li>Levkovitch-Verbin, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Animal+models+of+optic+nerve+diseases.">Animal models of optic nerve diseases.</a> Eye (Lond). 18 (11), 1066-1074 (2004).</li><li>Henry, K. R., Rhoades, R. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Relation+of+albinism+and+drugs+to+the+visual+evoked+potential+of+the+mouse).">Relation of albinism and drugs to the visual evoked potential of the mouse).</a> J Comp Physiol Psychol. 92 (2), 271-279 (1978).</li><li>Murrell, J. C., Waters, D., Johnson, C. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparative+effects+of+halothane,+isoflurane,+sevoflurane+and+desflurane+on+the+electroencephalogram+of+the+rat.">Comparative effects of halothane, isoflurane, sevoflurane and desflurane on the electroencephalogram of the rat.</a> Lab Anim. 42 (2), 161-170 (2008).</li><li>Makela, K., Hartikainen, K., Rorarius, M., Jantti, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Suppression+of+F-VEP+during+isoflurane-induced+EEG+suppression.">Suppression of F-VEP during isoflurane-induced EEG suppression.</a> Electroencephalogr Clin Neurophysiol. 100 (3), 269-272 (1996).</li><li>Boyes, W. K., Padilla, S., Dyer, R. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Body+temperature-dependent+and+independent+actions+of+chlordimeform+on+visual+evoked+potentials+and+axonal+transport+in+optic+system+of+rat.">Body temperature-dependent and independent actions of chlordimeform on visual evoked potentials and axonal transport in optic system of rat.</a> Neuropharmacology. 24 (8), 743-749 (1985).</li><li>Hetzler, B. E., Boyes, W. K., Creason, J. P., Dyer, R. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Temperature-dependent+changes+in+visual+evoked+potentials+of+rats.">Temperature-dependent changes in visual evoked potentials of rats.</a> Electroencephalogr Clin Neurophysiol. 70 (2), 137-154 (1988).</li><li>Mitchell, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effects+of+lysolecithin+on+non-myelinated+axons+in+vitro.">The effects of lysolecithin on non-myelinated axons in vitro.</a> Acta Neuropathol. 58 (4), 243-248 (1982).</li><li>Meyer, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acute+neuronal+apoptosis+in+a+rat+model+of+multiple+sclerosis.">Acute neuronal apoptosis in a rat model of multiple sclerosis.</a> J Neurosci. 21 (16), 6214-6220 (2001).</li><li>Lachapelle, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Failure+of+remyelination+in+the+nonhuman+primate+optic+nerve.">Failure of remyelination in the nonhuman primate optic nerve.</a> Brain Pathol. 15 (3), 198-207 (2005).</li></ol>

None of the authors have competing interests or conflicting interests.

The optic nerve is very susceptible to mechanical damage. Optic nerve crush injury over a duration of 1 s can lead to about 75% loss of RGC over a period of 2 weeks10. Therefore, extreme care is required while performing the surgical procedures. According to the authors&#8217; experience, it is much better to adapt a blunt dissection approach to expose and make way through the tissues around the optic nerve along the orientation of the nerve, rather than penetrating in a perpendicular orientation to the optic ...

Optic neuritis is one of the most common form of optic neuropathy, causing complete or partial loss of vision1. Histologically, it is featured by inflammatory demyelination, retinal ganglion cell axonal loss and varying degrees of remyelination in the optic nerve2. Optic neuritis is usually the manifest onset of multiple sclerosis. The visual evoked potential (VEP) is a non-invasive tool for investigating the function of the visual system. It reflects the post-retinal function from the retina to the primary visual cortex and is affected in many optic nerve disease conditions3. The VEP has been predominantly used in optic neuritis patients to assess the integrity of the visual pathway4. 
The latency of VEP, which reflects the velocity of signal conduction along the visual pathway, is considered to be an accurate measurement of the level of myelin associated changes in the optic nerve5; while the amplitude of VEP is believed to be closely correlated with axonal damage of the retinal ganglion cells (RGC)6. This hypothesis has been fairly well established using the rat model of lysolecithin-induced optic nerve demyelination5. 
Here, we explicate a comprehensive protocol of optic nerve microinjection technique in rodents, which can minimise the surgical manipulation-related damage to the nerve per se as well as to the adjacent tissues such as extraocular muscles and blood vessels. Also, the skull electrode implantation surgery has been described for VEP recording in animals7. The VEP recordings can be repeatedly carried out on animals over a period of time to assess demyelination/remyelination related changes as well as impact on axonal integrity in the optic nerve.

<table><tbody><tr><td>Ketamine 100 mg/ml (Ketamil)</td><td>Troy Laboratories</td><td>AC 116</td><td></td></tr><tr><td>Medetomidine 1 mg/ml (Domitor)</td><td>Pfizer</td><td>sc-204073</td><td></td></tr><tr><td>Tropicamide 1.0% (Mydriacyl)</td><td>Alcon</td><td>sc-202371</td><td></td></tr><tr><td>Homoeothermic blanket system</td><td>Harvard Apparatus</td><td>NC9203819</td><td></td></tr><tr><td>Impedance meter&amp;nbsp;</td><td>Grass</td><td>F-EZM5</td><td></td></tr><tr><td>Screw electrodes&amp;nbsp;</td><td>Micro Fasteners</td><td>M1.0&amp;times;3mm Csk Slot M/T 304 S/S</td><td></td></tr><tr><td>Subdermal needle electrodes&amp;nbsp;</td><td>Grass</td><td>F-E3M-72</td><td></td></tr><tr><td>Rapid Repair&amp;nbsp;</td><td>DeguDent GmbH</td><td></td><td></td></tr><tr><td>Light-emitting diode&amp;nbsp;</td><td>Nichia</td><td>NSPG300A</td><td></td></tr><tr><td>Bioamplifier</td><td>CWE, Inc.</td><td>BMA-400</td><td></td></tr><tr><td>CED system</td><td>Cambridge Electronic Design, Ltd.</td><td>Power1401</td><td></td></tr><tr><td>Hamilton syringe&amp;nbsp;</td><td>Hamilton</td><td>87930</td><td></td></tr><tr><td>Lysolecithin</td><td>Sigma</td><td>L4129</td><td></td></tr><tr><td>Evan&amp;rsquo;s blue&amp;nbsp;</td><td>Sigma</td><td>E2129</td><td></td></tr></tbody></table>

visual evoked potential recording in a rat model of experimental optic nerve demyelination

The visual evoked potential (VEP) recording is widely used in clinical practice to assess the severity of optic neuritis in its acute phase, and to monitor the disease course in the follow-up period. Changes in the VEP parameters closely correlate with pathological damage in the optic nerve. This protocol provides a detailed description about the rodent model of optic nerve microinjection, in which a partial demyelination lesion is produced in the optic nerve. VEP recording techniques are also discussed. Using skull implanted electrodes, we are able to acquire reproducible intra-session and between-session VEP traces. VEPs can be recorded on individual animals over a period of time to assess the functional changes in the optic nerve longitudinally. The optic nerve demyelination model, in conjunction with the VEP recording protocol, provides a tool to investigate the disease processes associated with demyelination and remyelination, and can potentially be employed to evaluate the effects of new remyelinating drugs or neuroprotective therapies.

Reproduzierbare inner sessional VEP Spuren sind in Abbildung 1 dargestellt und eine signifikante Verzögerung in N1 Latenzzeit kann nach dem Sehnerv Injektion zu sehen. Teil Sehnerv Läsionen der Demyelinisierung auf histologischen Schnitten mit Luxol schnelle Blau-Färbung 5 beobachtet werden. Abbildung 2 zeigt einen repräsentativen Querschnitt mit einer kleinen Brenn demyelinisierten Läsion in der Mitte des Sehnerven. Beachten Sie, dass Querschnitt repräsentieren nicht G...

Watch this Scientific Journal Video about Visual Evoked Potential Recording in a Rat Model of Experimental Optic Nerve Demyelination at JoVE.com

Visual Evoked Potential Recording in a Rat Model of Experimental Optic Nerve Demyelination

Focal demyelination is induced in the optic nerve using lysolecithin microinjection. Visual evoked potentials are recorded via skull electrodes implanted over the visual cortex to examine the signal conduction along the visual pathway in vivo. This protocol details the surgical procedures underlying electrode implantation and optic nerve microinjection.

Visuell evozierten Potential-Aufnahme in einem Rattenmodell für Experimentelle Optic Nerve Demyelinisierung

The visual evoked potential (VEP) recording is widely used in clinical practice to assess the severity of optic neuritis in its acute phase, and to ...

visual-evoked-potential-recording-rat-model-experimental-optic-nerve

Universidad de Cartagena UDEC

Research

JoVE Journal

Neuroscience

12.2K Aufrufe.  Macquarie University. Focal demyelination is induced in the optic nerve using lysolecithin microinjection. Visual evoked potentials are recorded via skull electrodes implanted over the visual cortex to examine the signal conduction along the visual pathway in vivo. This protocol details the surgical procedures underlying electrode implantation and optic nerve microinjection. 

Serie: Visual Evoked Potential Recording in a Rat Model of Experimental Optic Nerve Demyelination

Single-unit In vivo Recordings from the Optic Chiasm of Rat

Information about the visual world is transmitted to the brain in sequences of action potentials in retinal ganglion cell axons that make up the optic nerve. In vivo recordings of ganglion cell spike trains in several animal models have revealed much of what is known about how the early visual system processes and encodes visual information. However, such recordings have been rare in one of the most common animal models, the rat, possibly owing to difficulty in detecting spikes fired by small diameter axons. The many retinal disease models involving rats motivate a need for characterizing the functional properties of ganglion cells without disturbing the eye, as with intraocular or in vitro recordings. Here, we demonstrate a method for recording ganglion cell spike trains from the optic chiasm of the anesthetized rat. We first show how to fabricate tungsten-in-glass electrodes that can pick up electrical activity from single ganglion cell axons in rat. The electrodes outperform all commercial ones that we have tried. We then illustrate our custom-designed stereotaxic system for in vivo visual neurophysiology experiments and our procedures for animal preparation and reliable and stable electrode placement in the optic chiasm.

Single-unit In vivo Recordings from the Optic Chiasm of Rat

Retinal ganglion cells transmit visual information from the eye to the brain with sequences of action potentials. Here, we demonstrate how to record the action potentials of single ganglion cells in vivo from anesthetized rats.

Single-Einheit In-vivo- Aufnahmen aus dem Chiasma der Rat

Information about the visual world is transmitted to the brain in sequences of action potentials in retinal ganglion cell axons that make up the optic ...

Forschung

Neurowissenschaften

Implantation and Recording of Wireless Electroretinogram and Visual Evoked Potential in Conscious Rats

The full-field electroretinogram (ERG) and visual evoked potential (VEP) are useful tools to assess retinal and visual pathway integrity in both laboratory and clinical settings. Currently, preclinical ERG and VEP measurements are performed with anesthesia to ensure stable electrode placements. However, the very presence of anesthesia has been shown to contaminate normal physiological responses. To overcome these anesthesia confounds, we develop a novel platform to assay ERG and VEP in conscious rats. Electrodes are surgically implanted sub-conjunctivally on the eye to assay the ERG and epidurally over the visual cortex to measure the VEP. A range of amplitude and sensitivity/timing parameters are assayed for both the ERG and VEP at increasing luminous energies. The ERG and VEP signals are shown to be stable and repeatable for at least 4 weeks post surgical implantation. This ability to record ERG and VEP signals without anesthesia confounds in the preclinical setting should provide superior translation to clinical data.

We show surgical implantation and Recording procedures to measure visual electrophysiological signals from the eye (electroretinogram) and brain (visual evoked potential) in conscious rats, which is more analogous to the human condition where recordings are conducted without anesthesia confounds.

Implantations- und Aufzeichnung von Wireless-Elektroretinogramm und visuell evozierten Potenziale in wachen Ratten

The full-field electroretinogram (ERG) and visual evoked potential (VEP) are useful tools to assess retinal and visual pathway integrity in both ...

Verhalten

Rat Model of Photochemically-Induced Posterior Ischemic Optic Neuropathy

Posterior Ischemic optic neuropathy (PION) is a sight-devastating disease in clinical practice. However, its pathogenesis and natural history have&#160;remained poorly understood. Recently, we developed a reliable, reproducible animal model of PION and tested the treatment effect of some neurotrophic factors in this model1. The purpose of this video is to demonstrate our photochemically induced model of posterior ischemic optic neuropathy, and to evaluate its effects with retrograde labeling of retinal ganglion cells. Following surgical exposure of the posterior optic nerve, a photosensitizing dye, erythrosin B, is intravenously injected and a laser beam is focused onto the optic nerve surface. Photochemical interaction of erythrosin B and the laser during irradiation damages the vascular endothelium, prompting microvascular occlusion mediated by platelet thrombosis and edematous compression. The resulting ischemic injury yields a gradual but pronounced retinal ganglion cell dieback, owing to a loss of axonal input - a remote, injury-induced and clinically relevant outcome. Thus, this model provides a novel platform to study the pathophysiologic course of PION,&#160;and can be further optimized for testing therapeutic approaches for optic neuropathies as well as other CNS ischemic diseases.

The goal of this protocol is to photochemically induce ischemic injury to the posterior optic nerve in rat. This model is critical to studies of the pathophysiology of posterior ischemic optic neuropathy, and therapeutic approaches for this and other optic neuropathies, as well as of other CNS ischemic diseases.

Rattenmodell der photochemisch-induzierte Posterior ischämische Optikusneuropathie

Posterior Ischemic optic neuropathy (PION) is a sight-devastating disease in clinical practice. However, its pathogenesis and natural history ...

Medizin

Simultaneous Recording of Electroretinography and Visual Evoked Potentials in Anesthetized Rats

The electroretinogram (ERG) and visual evoked potential (VEP) are commonly used to assess the integrity of the visual pathway. The ERG measures the electrical responses of the retina to light stimulation, while the VEP measures the corresponding functional integrity of the visual pathways from the retina to the primary visual cortex following the same light event. The ERG waveform can be broken down into components that reflect responses from different retinal neuronal and glial cell classes. The early components of the VEP waveform represent the integrity of the optic nerve and higher cortical centers. These recordings can be conducted in isolation or together, depending on the application. The methodology described in this paper allows simultaneous assessment of retinal and cortical visual evoked electrophysiology from both eyes and both hemispheres. This is a useful way to more comprehensively assess retinal function and the upstream effects that changes in retinal function can have on visual evoked cortical function.

This protocol describes simultaneous measurement of electroretinogram and visual evoked potentials in anesthetized rats.

Gleichzeitige Aufnahme von Elektroretinographie und visuell evozierten Potentialen in narkotisierten Ratten

The electroretinogram (ERG) and visual evoked potential (VEP) are commonly used to assess the integrity of the visual pathway. The ERG measures the ...

Visual Evoked Potential Recordings in Mice Using a Dry Non-invasive Multi-channel Scalp EEG Sensor

For scalp EEG research environments with laboratory mice, we designed a dry-type 16 channel EEG sensor which is non-invasive, deformable, and re-usable because of the plunger-spring-barrel structural facet and mechanical strengths resulting from metal materials. The whole process for acquiring the VEP responses in vivo from a mouse consists of four steps: (1) sensor assembly, (2) animal preparation, (3) VEP measurement, and (4) signal processing. This paper presents representative measurements of VEP responses from multiple mice with a submicro-voltage signal resolution and sub-hundred millisecond temporal resolution. Although the proposed method is safer and more convenient compared to other previously reported animal EEG acquiring methods, there are remaining issues including how to enhance the signal-to-noise ratio and how to apply this technique with freely moving animals. The proposed method utilizes easily available resources and shows a repetitive VEP response with a satisfactory signal quality. Therefore, this method could be utilized for longitudinal experimental studies and reliable translational research exploiting non-invasive paradigms.

We designed a dry-type 16 channel EEG sensor which is non-invasive, deformable, and re-usable. This paper describes the whole process from manufacturing the proposed EEG electrode to signal processing of visual evoked potential (VEP) signals measured on a mouse scalp using a dry non-invasive multi-channel EEG sensor.

Visuelle evozierte möglichen Aufnahmen bei Mäusen mit einer trockenen nichtinvasive Mehrkanal Scalp EEG Sensor

For scalp EEG research environments with laboratory mice, we designed a dry-type 16 channel EEG sensor which is non-invasive, deformable, and re-usable ...

In Vivo Methods to Assess Retinal Ganglion Cell and Optic Nerve Function and Structure in Large Animals

The optic nerve collects axons signals from the retinal ganglion cells and transmits visual signal to the brain. Large animal models of optic nerve injury are essential for translating novel therapeutic strategies from rodent models to clinical application due to their closer similarities to humans in size and anatomy. Here we describe some in vivo methods to evaluate the function and structure of the retinal ganglion cells (RGCs) and optic nerve (ON) in large animals, including visual evoked potential (VEP), pattern electroretinogram (PERG) and optical coherence tomography (OCT). Both goat and non-human primate were employed in this study. By presenting these in vivo methods step by step, we hope to increase experimental reproducibility among different labs and facilitate the usage of large animal models of optic neuropathies.

In Vivo Methods to Assess Retinal Ganglion Cell and Optic Nerve Function and Structure in Large Animals

Here we demostrate several in vivo tests (flash visual evoked potential, pattern electroretinogram and optic coherence tomography) in goat and rhesus macaque to understand the structure and function of the optic nerve and its neurons.

Im lebenden Organismus Methoden zur Beurteilung der Funktion und Struktur retinaler Ganglienzellen und Sehnerven bei Großtieren

The optic nerve collects axons signals from the retinal ganglion cells and transmits visual signal to the brain. Large animal models of optic nerve injury ...

Inducing Local Demyelination in the Optic Nerve Using Lysolecithin

Source: You, Y. et al., <a href="https://appjove.unicartagenaproxy.elogim.com/v/52934">Visual Evoked Potential Recording in a Rat Model of Experimental Optic Nerve Demyelination.</a> J. Vis. Exp. (2015)
This video demonstrates the procedure of surgically accessing the optic nerve in an anesthetized animal and precisely delivering lysolecithin to induce focal demyelination, creating neuropathological conditions for further study.

Induktion der lokalen Demyelinisierung im Sehnerv mit Lysolecithin

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. ...

JoVE Encyclopedia of Experiments

Neuropathology

Epilepsy and Seizure Disorders

Preparing a Rat Model to Assess Visual Pathway Integrity Using an Electrophysiology Setup

Source: Nguyen, C. T. et al., <a href="https://appjove.unicartagenaproxy.elogim.com/v/54158">Simultaneous Recording of Electroretinography and Visual Evoked Potentials in Anesthetized Rats.</a>&#160;J. Vis. Exp. (2016)
This video demonstrates the preparation of a rat model to study retinal health and visual pathway integrity. Visual evoked potential and electroretinography reference and active electrodes are connected to the rat, followed by aligning the platform with the rat in front of the Ganzfeld bowl for even retinal illumination and closing the Faraday cage to avoid external interference.

Vorbereitung eines Rattenmodells zur Beurteilung der Integrität der Sehbahn unter Verwendung eines elektrophysiologischen Aufbaus

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. ...

Neurophysiology

Electrophysiology Techniques

Electron Microscopic Analysis of Myelin Damage in a Rat Model of Optic Nerve Injury

Take fixed optic nerve sections from a control rat and a rat with optic nerve injury.
The control section exhibits axons with tightly packed myelin sheaths containing concentric plasma membrane layers. The injured section exhibits decompacted myelin with large gaps.
Incubate with an osmium-ferrocyanide complex that binds to lipids and enhances the myelin sheath&#39;s contrast.
Treat with thiocarbohydrazide to create additional binding sites for osmium.
Reapply osmium to further enhance lipid contrast.
Stain with heavy metals, which bind to macromolecules and enhance structural contrast.
Dehydrate using increasing alcohol concentrations and rinse with acetone for resin embedding.
Embed the tissue in resin, and coat the resin with gold to improve image quality.
Use serial block-face scanning electron microscopy to simultaneously slice and image. Focus the electron beam on the surface and detect the backscattered electrons to generate images.
Myelin sheaths appear dark, while spaces within damaged sheaths appear lighter, allowing differentiation between normal and damaged myelin.

Elektronenmikroskopische Analyse von Myelinschäden in einem Rattenmodell für Sehnervenverletzungen

Neuromuscular Disorders

Recording of Visual Evoked Potentials via the Skull Electrodes in a Rat Model

Source: You, Y., et al. <a href="https://appjove.unicartagenaproxy.elogim.com/v/52934">Visual Evoked Potential Recording in a Rat Model of Experimental Optic Nerve Demyelination.</a> J. Vis. Exp. (2015).
This video demonstrates the process of measuring visual evoked potentials or VEPs in a dark-adapted, anesthetized rat with pre-implanted skull electrodes, which record the electrical signals from the brain&#39;s visual cortex in response to light stimuli.

Aufzeichnung visuell evozierter Potentiale über die Schädelelektroden in einem Rattenmodell

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. VEP ...