Name: ocular kinematics measured by in vitro stimulation of the cranial nerves in the turtle
Uploaded: 2018-06-02T12:30:00
Description: Watch this Scientific Journal Video about Ocular Kinematics Measured by In Vitro Stimulation of the Cranial Nerves in the Turtle at JoVE.com

Forfatterne takke fru Paulette McKenna og Lisa Pezzino i denne undersøgelse for sekretærservice og Mr. Phil Auerbach for teknisk support. Forfatterne også takke Drs. Michael Ariel og Michael S. Jones (Saint Louis University School of Medicine) for at indføre os til in vitro- isoleret hoved forberedelse. Finansiering til støtte for dette samarbejde blev leveret af Department of Biology (Robert S. Chase fond), den akademiske Forskningsudvalget og neurovidenskab Program på Lafayette College. Endelig, dette arbejde er dedikeret til Mr. Phil Auerbach, som afgik 28 September 2016; han afmonteret en scanning elektron mikroskop og anerkendt nytten af dets 5-aksede fase til brug i denne protokol. Hans venskab og opfindsomhed vil blive savnet meget.

Bemærk: Red-eared skyderen skildpadder, både mandlige og kvindelige, blev købt fra en kreditor. Skildpadder har været opstaldet i en varm animalske suite indeholder to 60-gallon bøtter udstyret med mursten øer for Sole under 250-W infrarødt lys. Miljøet blev opretholdt på et 14/10-h lys/mørke cyklus med vandtemperatur på 22 ° C. Lysene blev tændt kl 6:00 og slukket kl 8:00. Kampvogne udstyret med filtreringssystemer var rengøres ugentligt, og skildpadder blev fodret ad libitum hver anden dag. Pleje...

<ol>
	<li>Kikillus, K. H., Hare, K. M., Hartley, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Minimizing+false-negatives+when+predicting+the+potential+distribution+of+an+invasive+species:+A+bioclimatic+envelope+for+the+red-eared+slider+at+global+and+regional+scales.">Minimizing false-negatives when predicting the potential distribution of an invasive species: A bioclimatic envelope for the red-eared slider at global and regional scales.</a> Anim Conserv. 13, 5-15 (2010).</li><li>Lutz, P. L., Rosenthal, M., Sick, T. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Living+without+oxygen:+turtle+brain+as+a+model+of+anaerobic+metabolism.">Living without oxygen: turtle brain as a model of anaerobic metabolism.</a> Mol Physiol. 8, 411-425 (1985).</li><li>Lutz, P. L., Milton, 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=Negotiating+brain+anoxia+survival+in+the+turtle.">Negotiating brain anoxia survival in the turtle.</a> J Exp Biol. 207, 3141-3147 (2004).</li><li>Storey, K. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anoxia+tolerance+in+turtles:+Metabolic+regulation+and+gene+expression.">Anoxia tolerance in turtles: Metabolic regulation and gene expression.</a> Comp Biochem Physiol A-Mol Integr Physiol. 147 (2), 263-276 (2007).</li><li>Granda, A. M., Dearworth, J. R., Subramaniam, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Balanced+interactions+in+ganglion-cell+receptive+fields.">Balanced interactions in ganglion-cell receptive fields.</a> Vis Neurosci. 16, 319-332 (1999).</li><li>Dearworth, J. R., Granda, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Multiplied+functions+unify+shapes+of+ganglion-cell+receptive+fields+in+retina+of+turtle.">Multiplied functions unify shapes of ganglion-cell receptive fields in retina of turtle.</a> J Vis. 2 (3), 204-217 (2002).</li><li>Nesbit, S. C., Van Hoof, A. G., Le, C. C., Dearworth Jr, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extracellular+recording+of+light+responses+from+optic+nerve+fibers+and+the+caudal+photoreceptor+in+the+crayfish.">Extracellular recording of light responses from optic nerve fibers and the caudal photoreceptor in the crayfish.</a> J Undergrad Neurosci Educ. 14 (1), A29-A38 (2015).</li><li>McMahon, B. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Respiratory+and+circulatory+compensation+to+hypoxia+in+crustaceans.">Respiratory and circulatory compensation to hypoxia in crustaceans.</a> Resp Phsiol. 128 (3), 349-364 (2001).</li><li>Leigh, R. J., Zee, D. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The neurology of eye movements. , (1999).</li><li>Robinson, D. 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+method+of+measuring+eye+movement+using+a+scleral+search+coil+in+a+magnetic+field.">A method of measuring eye movement using a scleral search coil in a magnetic field.</a> IEEE Trans Biomed Eng. 10, 137-145 (1963).</li><li>Judge, S. J., Richmond, B. J., Chu, F. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Implantation+of+magnetic+search+coils+for+measurement+of+eye+position:+an+improved+method.">Implantation of magnetic search coils for measurement of eye position: an improved method.</a> Vis Res. 20, 535-538 (1980).</li><li>Ong, J. K. Y., Halswanter, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measuring+torsional+eye+movements+by+tracking+stable+iris+features.">Measuring torsional eye movements by tracking stable iris features.</a> J Neurosci Meth. 192, 261-267 (2010).</li><li>Kimmel, D. L., Mammo, D., Newsome, W. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tracking+the+eye+non-invasively:+simultaneous+comparison+of+the+scleral+search+coil+and+optical+tracking+techniques+in+the+macaque+monkey.">Tracking the eye non-invasively: simultaneous comparison of the scleral search coil and optical tracking techniques in the macaque monkey.</a> Front Behav Neurosci. 6 (49), 1-17 (2012).</li><li>Otero-Millan, J., Roberts, D. C., Lasker, A., Zee, D. S., Kheradmand, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Knowing+what+the+brain+is+seeing+in+three+dimensions:+A+novel,+noninvasive,+sensitive,+accurate,+and+low-noise+technique+for+measuring+ocular+torsion.">Knowing what the brain is seeing in three dimensions: A novel, noninvasive, sensitive, accurate, and low-noise technique for measuring ocular torsion.</a> J Vis. 15 (14), 1-15 (2015).</li><li>Demski, L. S., Bauer, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Eye+movements+evoked+by+electrical+stimulation+of+the+brain+in+anesthetized+fishes.">Eye movements evoked by electrical stimulation of the brain in anesthetized fishes.</a> Brain Behav Evol. 11, 109-129 (1975).</li><li>Gioanni, H., Bennis, M., Sansonetti, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Visual+and+vestibular+reflexes+that+stabilize+gaze+in+the+chameleon.">Visual and vestibular reflexes that stabilize gaze in the chameleon.</a> Vis Neurosci. 10, 947-956 (1993).</li><li>Straka, H., Dieringer, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Basic+organization+principles+of+the+VOR:+lessons+from+frogs.">Basic organization principles of the VOR: lessons from frogs.</a> Prog Neurobio. 73 (4), 259-309 (2004).</li><li>Voss, J., Bischof, H. -. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Eye+movements+of+laterally+eyed+birds+are+not+independent.">Eye movements of laterally eyed birds are not independent.</a> J Exp Biol. 212 (10), 1568-1575 (2009).</li><li>Ariel, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Independent+eye+movements+in+the+turtle.">Independent eye movements in the turtle.</a> Vis Neurosci. 5, 29-41 (1990).</li><li>Ariel, M., Rosenberg, A. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+synaptic+drugs+on+turtle+optokinetic+nystagmus+and+the+spike+responses+of+the+basal+optic+nucleus.">Effects of synaptic drugs on turtle optokinetic nystagmus and the spike responses of the basal optic nucleus.</a> Vis Neurosci. 7, 431-440 (1991).</li><li>Balaban, C. D., Ariel, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+"beat-to-beat"+interval+generator+for+optokinetic+nystagmus.">A "beat-to-beat" interval generator for optokinetic nystagmus.</a> Biol Cybern. 66, 203-216 (1992).</li><li>Keifer, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+eye-blink+reflex+model:+Role+of+excitatory+amino+acid+receptors+and+labeling+of+network+activity+with+sulforhodamine.">In vitro eye-blink reflex model: Role of excitatory amino acid receptors and labeling of network activity with sulforhodamine.</a> Exp Brain Res. 97, 239-253 (1993).</li><li>Keifer, J., Armstrong, K. E., Houk, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+classical+conditioning+of+abducens+nerve+discharge+in+turtles.">In vitro classical conditioning of abducens nerve discharge in turtles.</a> J Neurosci. 15, 5036-5048 (1995).</li><li>Rosenberg, A. F., Ariel, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+model+for+optokinetic+eye+movements+in+turtles+that+incorporates+properties+of+retinal+slip+neurons.">A model for optokinetic eye movements in turtles that incorporates properties of retinal slip neurons.</a> Vis Neurosci. 13, 375-383 (1996).</li><li>Ariel, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Open-loop+optokinetic+responses+of+the+turtle.">Open-loop optokinetic responses of the turtle.</a> Vis Res. 37, 925-933 (1997).</li><li>Anderson, C. W., Keifer, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Properties+of+conditioned+abducens+nerve+responses+in+a+highly+reduced+in+vitro+brainstem+preparation+from+the+turtle.">Properties of conditioned abducens nerve responses in a highly reduced in vitro brainstem preparation from the turtle.</a> J Neurophysiol. 81, 1242-1250 (1999).</li><li>Keifer, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+classical+conditioning+of+the+turtle+eyeblink+reflex:+Approaching+cellular+mechanisms+of+acquisition.">In vitro classical conditioning of the turtle eyeblink reflex: Approaching cellular mechanisms of acquisition.</a> Cerebell. 2, 55-61 (2003).</li><li>Zhu, D., Keifer, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pathways+controlling+trigeminal+and+auditory+nerve-evoked+abducens+eyeblink+reflexes+in+pond+turtles.">Pathways controlling trigeminal and auditory nerve-evoked abducens eyeblink reflexes in pond turtles.</a> Brain Behav Evol. 64, 207-222 (2004).</li><li>Jones, M. S., Ariel, M. <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+unilateral+eighth+nerve+block+on+fictive+VOR+in+the+turtle.">The effects of unilateral eighth nerve block on fictive VOR in the turtle.</a> Br Res. 1094, 149-162 (2006).</li><li>Jones, M. S., Ariel, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Morphology,+intrinsic+membrane+properties,+and+rotation-evoked+responses+of+trochlear+motoneurons+in+the+turtle.">Morphology, intrinsic membrane properties, and rotation-evoked responses of trochlear motoneurons in the turtle.</a> J Neurophysiol. 99 (3), 1187-1200 (2008).</li><li>Krenz, J. G., Naylor, G. J. P., Shaffer, H. B., Janzen, F. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+phylogenetics+and+evolution+of+turtles.">Molecular phylogenetics and evolution of turtles.</a> Mol Phylogenet Evol. 37 (1), 178-191 (2005).</li><li>Dearworth, J. 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=Role+of+the+trochlear+nerve+in+eye+abduction+and+frontal+vision+of+the+red-eared+slider+turtle+(Trachemys+scripta+elegans).">Role of the trochlear nerve in eye abduction and frontal vision of the red-eared slider turtle (Trachemys scripta elegans).</a> J Comp Neur. 52, 3464-3477 (2013).</li><li>Dearworth, J. 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=Pupil+constriction+evoked+in+vitro+by+stimulation+of+the+oculomotor+nerve+in+the+turtle+(Trachemys+scripta+elegans).">Pupil constriction evoked in vitro by stimulation of the oculomotor nerve in the turtle (Trachemys scripta elegans).</a> Vis Neurosci. 26, 309-318 (2009).</li><li>Mead, K., 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=IFEL+TOUR:+a+description+of+the+introduction+to+FUN+electrophysiology+labs+workshop+at+Bowdoin+College,+July+27-30,+and+the+resultant+faculty+learning+community.">IFEL TOUR: a description of the introduction to FUN electrophysiology labs workshop at Bowdoin College, July 27-30, and the resultant faculty learning community.</a> J Undergrad Neurosci Educ. 5, A42-A48 (2007).</li><li>Jackson, D. C., Ultsch, G. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Physiology+of+hibernation+under+the+ice+by+turtles+and+frogs.">Physiology of hibernation under the ice by turtles and frogs.</a> J Exp Zool A Ecol Genet Physiol. 313 (6), 311-327 (2010).</li><li>Romano, J. M., Dearworth, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pupil+constriction+evoked+by+stimulation+of+the+ciliary+nerve+in+the+red-eared+slider+turtle+(Trachemys+scripta+elegans).">Pupil constriction evoked by stimulation of the ciliary nerve in the red-eared slider turtle (Trachemys scripta elegans).</a> J Penns Acad Sci. 85, 4-8 (2011).</li><li>Miller, J. M., Robins, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extraocular-muscle+forces+in+alert+monkey.">Extraocular-muscle forces in alert monkey.</a> Vis Res. 32, 1099-1113 (1992).</li><li>Gamlin, P. D., Miller, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extraocular+muscle+motor+units+characterized+by+spike-triggered+averaging+in+alert+monkey.">Extraocular muscle motor units characterized by spike-triggered averaging in alert monkey.</a> J Neurosci Meth. 204, 159-167 (2011).</li><li>Quaia, C., Ying, H. S., Optican, L. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Viscoelastic+properties+of+passive+eye+muscle+in+primates.+III:+Force+elicited+by+natural+elongations.">The Viscoelastic properties of passive eye muscle in primates. III: Force elicited by natural elongations.</a> PLOS ONE. 5, A236-A254 (2010).</li><li>Anderson, S. 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=Dynamics+of+primate+oculomotor+plant+revealed+by+effects+of+abducens+microstimulation.">Dynamics of primate oculomotor plant revealed by effects of abducens microstimulation.</a> J Neurophys. 101, 2907-2923 (2009).</li><li>Maxwell, J. H., Harless, M., Morlock, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anesthesia+and+surgery.">Anesthesia and surgery.</a> Turtles: Perspective and Research. , 127-152 (1979).</li><li>AVMA Panel on Euthanasia. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=American+Veterinary+Medical+Association.">American Veterinary Medical Association.</a> J Am Vet Med Assoc. 218 (5), 669-696 (2001).</li><li>Clarke, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Shaping+the+pupil's+response+to+light+in+the+hooded+rat.">Shaping the pupil's response to light in the hooded rat.</a> Exp Br Res. 176, 641-651 (2007).</li><li>Bennett, R. 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+review+of+anesthesia+and+chemical+restraint+in+reptiles.">A review of anesthesia and chemical restraint in reptiles.</a> J Zoo Wild Med. 22 (3), 282-303 (1991).</li><li>Bickler, P. E., Buck, L. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hypoxia+Tolerance+in+Reptiles,+Amphibians,+and+Fishes:+Life+with+Variable+Oxygen+Availability.">Hypoxia Tolerance in Reptiles, Amphibians, and Fishes: Life with Variable Oxygen Availability.</a> Ann Rev Physiol. 69, 145-170 (2007).</li></ol>

Kritiske trin:
De kritiske trin inden for denne protokol er følgende: 1) dissektion og omhu at bevare levedygtigheden af transected nerver; 2) den matchende størrelser af de suge elektroder til hjernenerver reagere konsekvent; og 3) placering af hoved i kardan til at give tilstrækkelig kalibrering af rotationer af øjet.
Fejlfinding:
Dissektion kan være udfordrende, men efter endt det et par gange, skridt bør blive relativt ligetil. Hvis n...

Begrundelse for at anvende Red-eared skyderen skildpadder i elektrofysiologiske eksperimenter:
Rød eared skyderen skildpadder (Trachemys scripta elegans), betragtes som en af verdens værste invasive arter1 og kan angive, at et økosystem er i problemer. Grunden hvorfor Rødøret lysbillede turtles er så vellykket er dårligt forstået, men det kan til dels skyldes deres tolerant fysiologi og besiddelse af nervøs væv, der kan overleve under hypoksiske betingelser2,3,4 . Brug dem til eksperimenter ikke truer deres numre og med minimal indsats, elektrofysiologiske præparater kan forblive levedygtige over udvidede varigheder, så længe 18 timer5,6. Fordelen er lignende til fordel for ved hjælp af hvirvelløse dyr såsom Krebs7, som også har evnen til at modstå lave niveauer af ilt8.
Teknikker til måling af øjenbevægelser:
Tilgange til at måle øjenbevægelser i frontal-eyed dyr ved hjælp af ikke-menneskelige primater har været veludviklet9. Øjet roterer i kredsløb omkring tre akser: vandret, lodret og deformation. Den magnetiske spolen søgemetode er generelt betragtes som den mest pålidelige for måling rotationer, men er invasiv, der kræver små bredbånd skal indsættes i scleras af dyr10,11. Video-baserede systemer kan også måle rotationer og har fordelen at være ikke-invasiv. Udviklingen af bedre kameraer sammen med innovative billedbehandling har forbedret deres funktionalitet at gøre video-baserede systemer et attraktivt alternativ til at overveje12,13,14.
De teknikker, der er udviklet til måling af øjenbevægelser i nonmammals har været langt mindre betydelig. Foranstaltninger er enten lav opløsning eller beskrive kun nogle af rotationer15,16,17,18. Manglende udvikling kan være delvist skylden på vanskelighederne med uddannelse nonmammals at følge visuelle mål. Selvom øjenbevægelser er blevet godt undersøgt i rød eared skyderen skildpadder19,20,21,22,23,24,25 ,26,27,28,29,30, på grund af udfordringen i uddannelse dyr til at spore mål, den præcise kinematik af deres øjenbevægelser er dårligt forstået.
Rød eared skyderen skildpadder er generelt betragtes som lateral-eyed hvirveldyr, men fordi de kan fuldt trække deres hoveder ind i deres shell31, betydelig okklusion af de laterale visuelle felter af rygskjoldet opstår32. Resultatet er, at deres visuelle sigtelinie er tvunget mod fronten, hvilket gør dem opføre sig mere som frontal-eyed pattedyr. Derfor, deres anvendelse som en model for udvikling af metoder til måling af øjenbevægelser tilbyder også en unik evolutionært perspektiv.
Den protokol, der er beskrevet i dette værk bruger en in vitro- isoleret hoved forberedelse til at identificere kinematik af øjenbevægelser i rød eared skyderen skildpadder. Hjerner er dissekeret fra kranier forlader hjernenerver intakt. Hoveder er placeret i en kardan til at kalibrere øjenbevægelser og fremkalde reaktioner ved elektrisk stimulation af kranienerver innerverer øjet muskler. Foranstaltninger af rotationer af øjnene er udført af en video-baseret system, bruger Softwarealgoritmer, som spor den mørke elev og markeringer af iris. Forberedelse giver mulighed for at måle kinematik både ekstraokulær (dvs., vandrette, lodrette og torsions rotationer)32 og intraokulært (dvs., elev ændringer)33 bevægelser.
Modelsystem til analyse af efferente nervebaner:
Mere generelt giver tilgangen inspektørerne mulighed for at studere hvordan efferente neurale signaler generere øjenbevægelser når musklerne starter fra deres afslappet stater og i mangel af integrerede sensoriske oplysninger behandles af hjernen32, 33. Således, øjet kinematik kan undersøges i en modelsystem, hvor de behandles udelukkende af de efferente neurale pathway forlader hjernen og synapsing på musklerne.

<table>
	<tbody>
		<tr>
			<td>Red-eared slider turtles</td>
			<td>Kons Scientific</td>
			<td>Trachemys scripta elegans</td>
			<td>Large size (carapace length 15-20 cm)</td>
		</tr>
		<tr>
			<td>Sodium chloride</td>
			<td>Sigma-Aldrich Co. LLC.</td>
			<td>S5886</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium chloride</td>
			<td>Sigma-Aldrich Co. LLC.</td>
			<td>P5405</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnesium choride</td>
			<td>Sigma-Aldrich Co. LLC.</td>
			<td>M7304</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium bicarbonate</td>
			<td>Sigma-Aldrich Co. LLC.</td>
			<td>S5761</td>
			<td></td>
		</tr>
		<tr>
			<td>Dextrose</td>
			<td>Sigma-Aldrich Co. LLC.</td>
			<td>C5767</td>
			<td></td>
		</tr>
		<tr>
			<td>Concentrated hydrochloric acid</td>
			<td>Sigma-Aldrich Co. LLC.</td>
			<td>H7020</td>
			<td></td>
		</tr>
		<tr>
			<td>Calcium chloride</td>
			<td>Sigma-Aldrich Co. LLC.</td>
			<td>C7902</td>
			<td></td>
		</tr>
		<tr>
			<td>pH meter</td>
			<td>Oakton</td>
			<td>pH 6+</td>
			<td></td>
		</tr>
		<tr>
			<td>Suction stimulation electrode</td>
			<td>A-M Systems</td>
			<td>573000</td>
			<td>Bipolar suction electrode. Note that 573000 has been replaced with 573050.</td>
		</tr>
		<tr>
			<td>Capillary glass</td>
			<td>A-M systems</td>
			<td>626000</td>
			<td>Single-barrel borosilicate capillary glass without microfilament, length 10 cm, outside diameter 1.0 mm, inner diameter 0.50 mm</td>
		</tr>
		<tr>
			<td>Alternative suction stimulation electrode</td>
			<td>A-M Systems</td>
			<td>573050</td>
			<td>Bipolar suction electrode. Requires larger diameter capillary glass: 627000, outside diameter 1.2 mm, inner diameter 0.68 mm</td>
		</tr>
		<tr>
			<td>Stereoscope</td>
			<td>Lieca</td>
			<td>GZ7</td>
			<td>Magnification range, 10x &#8211; 70x</td>
		</tr>
		<tr>
			<td>Fiber optic light source</td>
			<td>Amscope</td>
			<td>HL250-A</td>
			<td>150W Fiber optical microscope illuminator light box</td>
		</tr>
		<tr>
			<td>Rongeurs</td>
			<td>Carolina Biological Supply Company</td>
			<td>625654</td>
			<td>stainless steel, straight spring, 5.25&#34;</td>
		</tr>
		<tr>
			<td>Blunt dissection probe</td>
			<td>Carolina Biological Supply Company</td>
			<td>627405</td>
			<td>Huber mall probe, double-ended probe and seeker, 6&#34;</td>
		</tr>
		<tr>
			<td>Microscissors</td>
			<td>Carolina Biological Supply Company</td>
			<td>623555</td>
			<td>Iris microdissecting scissors, stainless steel, 0.5&#34; blades, 4.75&#34; long</td>
		</tr>
		<tr>
			<td>Fine forceps</td>
			<td>Sigma-Aldrich Co. LLC.</td>
			<td>F6521</td>
			<td>Jewelers forceps, dumont No. 5, inox alloy, 4.25&#34;</td>
		</tr>
		<tr>
			<td>Curved forceps</td>
			<td>Sigma-Aldrich Co. LLC.</td>
			<td>Z168696</td>
			<td>Medium tip, curved forceps, stainless steel, 4&#34;</td>
		</tr>
		<tr>
			<td>Scalpel handle</td>
			<td>Sigma-Aldrich Co. LLC.</td>
			<td>S2896</td>
			<td>Scalpel handles, No. 3, stainless steel</td>
		</tr>
		<tr>
			<td>Scalpel blade</td>
			<td>Sigma-Aldrich Co. LLC.</td>
			<td>S2771</td>
			<td>Scalpel blades, No. 11, steel</td>
		</tr>
		<tr>
			<td>Guillotine</td>
			<td>Harvard Apparatus</td>
			<td>73-1918</td>
			<td>Kleine guillotine type 7575</td>
		</tr>
		<tr>
			<td>Spatula</td>
			<td>Sigma</td>
			<td>Z648299</td>
			<td>Micro spoon and spatula weighing set. Use small spatula: 5.9&#8221; long x 0.07&#8221; diameter handle with square end: 0.17&#8221; x 1.3&#8221; long, other end round: 0.17&#8221; x 1.27&#8221; long</td>
		</tr>
		<tr>
			<td>Hook</td>
			<td>Autozone</td>
			<td>98069</td>
			<td>SureBilt hook and pick set. Use grinder to dull sharp points of hook to prevent injury to animals mouth.</td>
		</tr>
		<tr>
			<td>95/5% O2/CO2</td>
			<td>Airgas, Inc.</td>
			<td>X02OX95C2003102</td>
			<td>5% Carbon dioxide balance oxygen certified standard gas mixture, size 200 Cylinder, CGA-296</td>
		</tr>
		<tr>
			<td>Regulator</td>
			<td>Airgas, Inc.</td>
			<td>Y11244D296-AG</td>
			<td>Single stage brass 0-100 psi analytical cylinder regulator CGA-296 with needle outlet. Use brass adjustable airline pipe valve to go from 3/8&#34;, inner diameter, vinyl airline tubing connected to regulator to a 3/16&#34;, inner diameter, airline connection going to airstone or glass pasteur pipette.</td>
		</tr>
		<tr>
			<td>Adjustable airline pipe valve</td>
			<td>Doctors Foster and Smith</td>
			<td>CD-12061</td>
			<td>Brass valve</td>
		</tr>
		<tr>
			<td>Rigid table</td>
			<td>Unknown</td>
			<td>Unknown</td>
			<td>Auto-clave door laid on top of a sturdy table. Nine 5&#34; diameter tennis balls isolate vibrations from the top surface of the table.</td>
		</tr>
		<tr>
			<td>5&#34; tennis ball</td>
			<td>Petco Animal Supplies, Inc.</td>
			<td>712868</td>
			<td>Petco Jumbo Pet Tennis Ball: balls are unsliced and held within an integrated frame on the underside part of the autoclave door.</td>
		</tr>
		<tr>
			<td>Alternative vibration isolation table</td>
			<td>Newport Corporation</td>
			<td>INT1-36-6-N</td>
			<td>Rigid vibration control system, integrity 1: Surface dimensions, 3&#39; x 6&#39;</td>
		</tr>
		<tr>
			<td>Gimbal</td>
			<td>ISI, International Scientific Instruments, Inc.</td>
			<td>Stage from SUPER III-A Scanning EM</td>
			<td>5-axis eucentric stage: X, Y, and Z linear movements, &#177;20 mm, 0.1 mm precision; Rotations, vertical, &#177;10&#176;, and horizontal, &#177;12.5&#176;, with 1.25&#176; precision. Note: from decommission instrument.</td>
		</tr>
		<tr>
			<td>Chuck for gimbal</td>
			<td>Unknown</td>
			<td>Unknown</td>
			<td>Chuck from an old microtome of unknown manufacture was machined to fit the shaft of the specimen holder of the Scanning EM stage</td>
		</tr>
		<tr>
			<td>Alternative gimbal</td>
			<td>ThorLabs, Inc.</td>
			<td>GN2/M with MBT602/M</td>
			<td>Dual-axis goniometer (GN2/M) mounted on 3-axis microblock stage with thumbscrew adjusters (MBT602/M): design a chuck to hold turtle head with eye at 12.7 mm above top surface of goniometer (distance to point of rotation)</td>
		</tr>
		<tr>
			<td>Video-based eye tracking system</td>
			<td>Arrington Research, Inc.</td>
			<td>ViewPoint EyeTracker, PC-60</td>
			<td>Tracking method: Infrared video by dark pupil; Black and white camera (Item BC02): 30 Hz, 640 x 480; System requirements: Windows 2000, XP, 7, 8, 8.1, 10; Visual range: Horizontal +/- 44&#176;; vertical +/- 20&#176;; Accuracy ~0.5&#176;; Spatial resolution ~0.15&#176;; Pupil size resolution ~0.03 mm; Eye data: X, Y position of gaze, pupil height and width, torsion, delta time, total time, and regions of interest (ROI); Real-time communication (Item 0022): 4-Channel AnalogOut with eight TTL input channels to mark codes into the data file</td>
		</tr>
		<tr>
			<td>Multi-position magnetic base</td>
			<td>Harbor Freight Tools</td>
			<td>Pittsburg, item #5645</td>
			<td>Magnetic holder reaches up to 12&#34; and produces 45 lbs. of magnetic pull. Use to position camera. Machine thread holes onto the end of the rod to mount cameras.</td>
		</tr>
		<tr>
			<td>Micromanipulator</td>
			<td>Kopf</td>
			<td>900</td>
			<td>5 axis manipulation for mount of suction electrode: X, Y, Z linear travel, 2 axis of rotation</td>
		</tr>
		<tr>
			<td>Dissection scope on boom</td>
			<td>Lieca</td>
			<td>GZ6</td>
			<td>Magnification range, 6.7x &#8211; 40x</td>
		</tr>
		<tr>
			<td>Nerve/muscle stimulator</td>
			<td>Astro-Med Grass Telefactor</td>
			<td>Grass S88</td>
			<td>Dual pulse voltage stimulator: two output channels that can be operated independently or synchronized to generate non-isolated constant voltage pulses (10 mv to 150 V). Pulses can be single (10 &#956;sec to 10 sec), repetitive (0.01 Hz to 1 KHz), and trains (1 ms to 10 s) and synchronized with TTL inputs and output. Send TTL outputs via the output channels of a DB25 connector to the TTL input channels of the ViewPoint EyeTracker. Note: Astro-Med Grass Telefactor is no longer in business.</td>
		</tr>
		<tr>
			<td>Current isolation device</td>
			<td>Astro-Med Grass Telefactor</td>
			<td>PSIU6</td>
			<td>Current stimulus isolation unit: enables safe delivery of constant currents by the S88 to the preparation. The PSIU6 connects by a BNC cable to one of the output channels of the S88. Multiplier switches on the PSIU6 allow the S88 to generate a wide array of current amplitudes ranging from 0.1 &#181;A to 15 mA.</td>
		</tr>
		<tr>
			<td>Alternative nerve/muscle stimulator with isolation</td>
			<td>A-M Systems</td>
			<td>2100</td>
			<td>Isolated Pulse Stimulator: Unit has built-in isolator to produce constant currents.</td>
		</tr>
	</tbody>
</table>

ocular kinematics measured by in vitro stimulation of the cranial nerves in the turtle

Når dyrene er aflivet, begynder deres væv at dø. Skildpadder tilbyder en fordel på grund af en længere overlevelsestid af deres væv, især sammenlignet med varmblodede hvirveldyr. På grund af dette, kan in vitro- eksperimenter i skildpadder udføres for længere tid til at undersøge de neurale signaler og kontrol af deres målrettede aktioner. Ved hjælp af en isoleret hoved forberedelse, vi målte kinematik øje bevægelser i skildpadder, og deres graduering af elektriske signaler transporteres af hjernenerver. Efter at hjernen var fjernet fra kraniet, blev forlader hjernenerver intakt, dissekeret hovedet placeret i en kardan til at kalibrere øjenbevægelser. Glas elektroderne var knyttet til hjernenerver (oculomotor, trochlea, og abducens) og stimuleres med strøm til at fremmane øjenbevægelser. Vi overvågede øjenbevægelser med en infrarød video tracking system og kvantificerede rotationer af øjnene. Nuværende pulser med en vifte af amplituder, frekvenser, og tog varigheder blev brugt til at observere effekter på svar. Fordi præparatet er adskilt fra hjernen, kan den efferente pathway gonna muskel mål undersøges isoleret at undersøge neurale signaler i mangel af centralt forarbejdede sensorisk information.

Figur 1 viser stills af billeder taget fra en video, der beskriver dissektion. Billeder give typiske steder af nerver før skæring fra hjernen.
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/56864/56864fig1.jpg"> Figur 1: Stills af billeder taget fra video af dissektion at vise placeringen af synsnerven (nII), oculomotor nerve (nIII), trochlea nerve (nIV), og abducens nerve (...

Watch this Scientific Journal Video about Ocular Kinematics Measured by In Vitro Stimulation of the Cranial Nerves in the Turtle at JoVE.com

Ocular Kinematics Measured by In Vitro Stimulation of the Cranial Nerves in the Turtle

Denne protokol beskriver hvordan man bruger en in vitro- isoleret turtle head forberedelse til måler kinematikken af deres øjenbevægelser. Efter fjernelse af hjernen fra kraniet, kan hjernenerver stimuleres med strøm til at kvantificere rotationer af øjet og ændringer i elev størrelser.

Okulær kinematik målt ved In Vitro Stimulation af hjernenerver i skildpadden

After animals are euthanized, their tissues begin to die. Turtles offer an advantage because of a longer survival time of their tissues, especially when ...

Research

JoVE Journal

Neuroscience

Ocular Kinematics Measured by In Vitro Stimulation of the Cranial Nerves in the Turtle

TMS: Using the Theta-Burst Protocol to Explore Mechanism of Plasticity in Individuals with Fragile X Syndrome and Autism

Fragile X Syndrome (FXS), also known as Martin-Bell Syndrome, is a genetic abnormality found on the X chromosome.1,2 Individuals suffering from FXS ...

The NeuroStar TMS Device: Conducting the FDA Approved Protocol for Treatment of Depression

The Neuronetics NeuroStar Transcranial Magnetic Stimulation (TMS) System is a class II medical device that produces brief duration, pulsed magnetic ...

Visualization of Proprioceptors in Drosophila Larvae and Pupae

Visualization of Proprioceptors in Drosophila Larvae and Pupae

Visualisering af proprioceptorer i<em&gt; Drosophila</em&gt; Larver og pupper

Proprioception is the ability to sense the motion, or position, of body parts by responding to stimuli arising within the body. In fruitflies and other ...

Direct Intraventricular Delivery of Drugs to the Rodent Central Nervous System

Direkte Intraventrikulær Delivery of Drugs til Rodent centrale nervesystem

Due to an inability to cross the blood brain barrier, certain drugs need to be directly delivered into the central nervous system (CNS). Our lab focuses ...

Fast Micro-iontophoresis of Glutamate and GABA: A Useful Tool to Investigate Synaptic Integration

Hurtig Mikro-iontophoresis af glutamat og GABA: et nyttigt redskab til Undersøge Synaptic Integration

One of the fundamental interests in neuroscience is to understand the integration of excitatory and inhibitory inputs along the very complex structure of ...

Generation of Local CA1 &#947; Oscillations by Tetanic Stimulation

Generation of Local CA1 ÃƒÅ½Ã‚Â³ Oscillations by Tetanic Stimulation

Generation af Lokale CA1 γ Svingninger ved tetanisk Stimulation

Neuronal network oscillations are important features of brain activity in health and disease and can be modulated by a range of clinically used drugs. A ...

Caspase-3 Activity in the Rat Amygdala Measured by Spectrofluorometry After Myocardial Infarction

Caspase-3 Aktivitet i Rat Amygdala Målt spektrofluorometri efter myokardieinfarkt

Myocardial infarction (MI) has dramatic mid- and long-term consequences at the physiological and behavioral levels, but the mechanisms involved are still ...

Utilizing Combined Methodologies to Define the Role of Plasma Membrane Delivery During Axon Branching and Neuronal Morphogenesis

Udnytte Kombineret Metoder til Definer rolle plasmamembranen Delivery Under Axon Forgrening og neuronal Morfogenese

During neural development, growing axons extend to multiple synaptic partners by elaborating axonal branches. Axon branching is promoted by extracellular ...

The Power of Interstimulus Interval for the Assessment of Temporal Processing in Rodents

Kraften i Interstimulus Interval for vurdering af tidsmæssige forarbejdes i Rodents

Temporal processing deficits have been implicated as a potential elemental dimension of higher-level cognitive processes, commonly observed in ...

Assessing Pupil-linked Changes in Locus Coeruleus-mediated Arousal Elicited by Trigeminal Stimulation

Vurdering af elev-linkede ændringer i locus Coeruleus-medieret ophidselse, fremkaldt af trigeminal stimulation

Current scientific literature provides evidence that trigeminal sensorimotor activity associated with chewing may affect arousal, attention, and cognitive ...