Name: using looming visual stimuli to evaluate mouse vision
Uploaded: 2019-06-13T14:30:00
Description: Watch this Scientific Journal Video about Using Looming Visual Stimuli to Evaluate Mouse Vision at JoVE.com

This work was supported by NIH R01 EY028915 (TI) and RPB grants.

All experiments and animal care were conducted in accordance with protocol approved by the Institutional Animal Care and Use Committees at Wayne State University (protocol no. 17-11-0399).
1. Preparation for the experiment
<ol>
	<li>Build a rectangular open-lid enclosure to house the mouse during looming visual stimuli presentation. We constructed a 40 cm x 50 cm x 33 cm enclosure using aluminum framing and PVC panels (Figure 1A,B). Lay a sheet ...

<ol>
	<li>Enroth-Cugell, C., Robson, J. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+contrast+sensitivity+of+retinal+ganglion+cells+of+the+cat.">The contrast sensitivity of retinal ganglion cells of the cat.</a> The Journal of Physiology. 187 (3), 517-552 (1966).</li><li>Boycott, B. B., W&#228;ssle, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+morphological+types+of+ganglion+cells+of+the+domestic+cat's+retina.">The morphological types of ganglion cells of the domestic cat's retina.</a> The Journal of Physiology. 240 (2), 397-419 (1974).</li><li>Livingstone, M. S., Hubel, 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=Segregation+of+form,+color,+movement,+and+depth:+anatomy,+physiology,+and+perception.">Segregation of form, color, movement, and depth: anatomy, physiology, and perception.</a> Science. 240 (4853), 740-749 (1988).</li><li>Livingstone, M. S., Hubel, 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=Psychophysical+evidence+for+separate+channels+for+the+perception+of+form,+color,+movement,+and+depth.">Psychophysical evidence for separate channels for the perception of form, color, movement, and depth.</a> The Journal of Neuroscience. 7 (11), 3416-3468 (1987).</li><li>W&#228;ssle, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Parallel+processing+in+the+mammalian+retina.">Parallel processing in the mammalian retina.</a> Nature Reviews Neuroscience. 5 (10), 747-757 (2004).</li><li>Awatramani, G. B., Slaughter, M. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Origin+of+transient+and+sustained+responses+in+ganglion+cells+of+the+retina.">Origin of transient and sustained responses in ganglion cells of the retina.</a> The Journal of Neuroscience. 20 (18), 7087-7095 (2000).</li><li>Ghosh, K. K., Bujan, S., Haverkamp, S., Feigenspan, A., W&#228;ssle, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Types+of+bipolar+cells+in+the+mouse+retina.">Types of bipolar cells in the mouse retina.</a> The Journal of Comparative Neuroscience. 469 (1), 70-82 (2004).</li><li>W&#228;ssle, H., Puller, C., Muller, F., Haverkamp, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cone+contacts,+mosaics,+and+territories+of+bipolar+cells+in+the+mouse+retina.">Cone contacts, mosaics, and territories of bipolar cells in the mouse retina.</a> The Journal of Neuroscience. 29 (1), 106-117 (2009).</li><li>Helmstaedter, M., 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=Connectomic+reconstruction+of+the+inner+plexiform+layer+in+the+mouse+retina.">Connectomic reconstruction of the inner plexiform layer in the mouse retina.</a> Nature. 500 (7461), 168-174 (2013).</li><li>Shekhar, 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=Comprehensive+Classification+of+Retinal+Bipolar+Neurons+by+Single-Cell+Transcriptomics.">Comprehensive Classification of Retinal Bipolar Neurons by Single-Cell Transcriptomics.</a> Cell. 166 (5), 1308-1323 (2016).</li><li>Wu, S. M., Gao, F., Maple, 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=Functional+architecture+of+synapses+in+the+inner+retina:+segregation+of+visual+signals+by+stratification+of+bipolar+cell+axon+terminals.">Functional architecture of synapses in the inner retina: segregation of visual signals by stratification of bipolar cell axon terminals.</a> The Journal of Neuroscience. 20 (12), 4462-4470 (2000).</li><li>Sun, W., Li, N., He, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Large-scale+morphological+survey+of+mouse+retinal+ganglion+cells.">Large-scale morphological survey of mouse retinal ganglion cells.</a> The Journal of Comparative Neuroscience. 451 (2), 115-126 (2002).</li><li>Volgyi, B., Chheda, S., Bloomfield, S. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tracer+coupling+patterns+of+the+ganglion+cell+subtypes+in+the+mouse+retina.">Tracer coupling patterns of the ganglion cell subtypes in the mouse retina.</a> The Journal of Comparative Neuroscience. 512 (5), 664-687 (2009).</li><li>Kong, J. H., Fish, D. R., Rockhill, R. L., Masland, R. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diversity+of+ganglion+cells+in+the+mouse+retina:+Unsupervised+morphological+classification+and+its+limits.">Diversity of ganglion cells in the mouse retina: Unsupervised morphological classification and its limits.</a> The Journal of Comparative Neuroscience. 489 (3), 293-310 (2005).</li><li>Sumbul, U., 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=A+genetic+and+computational+approach+to+structurally+classify+neuronal+types.">A genetic and computational approach to structurally classify neuronal types.</a> Nature Communications. 5, 3512 (2014).</li><li>Baden, T., 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=The+functional+diversity+of+retinal+ganglion+cells+in+the+mouse.">The functional diversity of retinal ganglion cells in the mouse.</a> Nature. 529 (7586), 345-350 (2016).</li><li>Lindstrom, S. H., Ryan, D. G., Shi, J., DeVries, S. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Kainate+receptor+subunit+diversity+underlying+response+diversity+in+retinal+Off+bipolar+cells.">Kainate receptor subunit diversity underlying response diversity in retinal Off bipolar cells.</a> The Journal of Physiology. 592, 1457-1477 (2014).</li><li>Euler, T., Haverkamp, S., Schubert, T., Baden, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Retinal+bipolar+cells:+elementary+building+blocks+of+vision.">Retinal bipolar cells: elementary building blocks of vision.</a> Nature Reviews Neuroscience. 15 (8), 507-519 (2014).</li><li>Yoshida, 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=A+key+role+of+starburst+amacrine+cells+in+originating+retinal+directional+selectivity+and+optokinetic+eye+movement.">A key role of starburst amacrine cells in originating retinal directional selectivity and optokinetic eye movement.</a> Neuron. 30 (3), 771-780 (2001).</li><li>Pearson, R. A., 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=Restoration+of+vision+after+transplantation+of+photoreceptors.">Restoration of vision after transplantation of photoreceptors.</a> Nature. 485 (7396), 99-103 (2012).</li><li>Saszik, S. M., Robson, J. G., Frishman, 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=The+scotopic+threshold+response+of+the+dark-adapted+electroretinogram+of+the+mouse.">The scotopic threshold response of the dark-adapted electroretinogram of the mouse.</a> The Journal of Physiology. 543, 899-916 (2002).</li><li>Reuter, J. H., Sanyal, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+and+degeneration+of+retina+in+rds+mutant+mice:+the+electroretinogram.">Development and degeneration of retina in rds mutant mice: the electroretinogram.</a> Neuroscience Letters. 48 (2), 231-237 (1984).</li><li>Woodward, W. 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=Isoflurane+is+an+effective+alternative+to+ketamine/xylazine/acepromazine+as+an+anesthetic+agent+for+the+mouse+electroretinogram.">Isoflurane is an effective alternative to ketamine/xylazine/acepromazine as an anesthetic agent for the mouse electroretinogram.</a> Documenta Ophthalmologica. 115 (3), 187-201 (2007).</li><li>Cahill, H., Nathans, 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+optokinetic+reflex+as+a+tool+for+quantitative+analyses+of+nervous+system+function+in+mice:+application+to+genetic+and+drug-induced+variation.">The optokinetic reflex as a tool for quantitative analyses of nervous system function in mice: application to genetic and drug-induced variation.</a> PLoS One. 3 (4), 2055 (2008).</li><li>Prusky, G. T., Alam, N. M., Beekman, S., Douglas, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+quantification+of+adult+and+developing+mouse+spatial+vision+using+a+virtual+optomotor+system.">Rapid quantification of adult and developing mouse spatial vision using a virtual optomotor system.</a> Investigative Ophthalmology &amp; Visual Science. 45 (12), 4611-4616 (2004).</li><li>Lu, Q., Ganjawala, T. H., Hattar, S., Abrams, G. W., Pan, Z. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Robust+Optomotor+Assay+for+Assessing+the+Efficacy+of+Optogenetic+Tools+for+Vision+Restoration.">A Robust Optomotor Assay for Assessing the Efficacy of Optogenetic Tools for Vision Restoration.</a> Investigative Ophthalmology &amp; Visual Science. 59 (3), 1288-1294 (2018).</li><li>Xue, T., 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=Melanopsin+signalling+in+mammalian+iris+and+retina.">Melanopsin signalling in mammalian iris and retina.</a> Nature. 479 (7371), 67-73 (2011).</li><li>Yilmaz, M., Meister, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+innate+defensive+responses+of+mice+to+looming+visual+stimuli.">Rapid innate defensive responses of mice to looming visual stimuli.</a> Current Biology. 23 (20), 2011-2015 (2013).</li><li>De Franceschi, G., Vivattanasarn, T., Saleem, A. B., Solomon, 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=Vision+Guides+Selection+of+Freeze+or+Flight+Defense+Strategies+in+Mice.">Vision Guides Selection of Freeze or Flight Defense Strategies in Mice.</a> Current Biology. 26 (16), 2150-2154 (2016).</li><li>Temizer, I., Donovan, J. C., Baier, H., Semmelhack, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Visual+Pathway+for+Looming-Evoked+Escape+in+Larval+Zebrafish.">A Visual Pathway for Looming-Evoked Escape in Larval Zebrafish.</a> Current Biology. 25 (14), 1823-1834 (2015).</li><li>Guest, B. B., Gray, 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=Responses+of+a+looming-sensitive+neuron+to+compound+and+paired+object+approaches.">Responses of a looming-sensitive neuron to compound and paired object approaches.</a> Journal of Neurophysiology. 95 (3), 1428-1441 (2006).</li><li>McMillan, G. A., Gray, 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=A+looming-sensitive+pathway+responds+to+changes+in+the+trajectory+of+object+motion.">A looming-sensitive pathway responds to changes in the trajectory of object motion.</a> Journal of Neurophysiology. 108 (4), 1052-1068 (2012).</li><li>Vagnoni, E., Lourenco, S. F., Longo, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Threat+modulates+neural+responses+to+looming+visual+stimuli.">Threat modulates neural responses to looming visual stimuli.</a> Eur The Journal of Neuroscience. 42 (5), 2190-2202 (2015).</li><li>Coker-Appiah, D. S., 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=Looming+animate+and+inanimate+threats:+the+response+of+the+amygdala+and+periaqueductal+gray.">Looming animate and inanimate threats: the response of the amygdala and periaqueductal gray.</a> Social Neuroscience. 8 (6), 621-630 (2013).</li><li>Tyll, S., 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=Neural+basis+of+multisensory+looming+signals.">Neural basis of multisensory looming signals.</a> Neuroimage. 65, 13-22 (2013).</li><li>Wei, P., 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=Processing+of+visually+evoked+innate+fear+by+a+non-canonical+thalamic+pathway.">Processing of visually evoked innate fear by a non-canonical thalamic pathway.</a> Nature Communications. 6, 6756 (2015).</li><li>Shang, C., 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=Divergent+midbrain+circuits+orchestrate+escape+and+freezing+responses+to+looming+stimuli+in+mice.">Divergent midbrain circuits orchestrate escape and freezing responses to looming stimuli in mice.</a> Nature Communications. 9 (1), 1232 (2018).</li><li>Salay, L. D., Ishiko, N., Huberman, A. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+midline+thalamic+circuit+determines+reactions+to+visual+threat.">A midline thalamic circuit determines reactions to visual threat.</a> Nature. 557 (7704), 183-189 (2018).</li><li>Vale, R., Evans, D., Branco, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Behavioral+Assay+for+Investigating+the+Role+of+Spatial+Memory+During+Instinctive+Defense+in+Mice.">A Behavioral Assay for Investigating the Role of Spatial Memory During Instinctive Defense in Mice.</a> Journal of Visualized Experiments. (137), 56988 (2018).</li><li>Tungtur, S. K., Nishimune, N., Radel, J., Nishimune, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+Behavior+Tracker:+An+economical+method+for+tracking+behavior+in+home+cages.">Mouse Behavior Tracker: An economical method for tracking behavior in home cages.</a> Biotechniques. 63 (5), 215-220 (2017).</li></ol>

With the looming visual stimuli system, a majority (97%) of healthy eye-mice showed flight response. One of 29 mice did not show an obvious flight response. However, the mouse walked toward the dome and remained near it until looming disappeared, indicating that the mouse was at least cautious when the looming stimuli occurred. Therefore, the looming stimuli consistently elicited innate fear responses in healthy-eyed mice. On the other hand, three blind mice did not show any responses to the looming (preliminary results)...

The visual system starts at the retina, where visual signals are captured by photoreceptors, channeled to bipolar cells (2nd-order neurons), and finally passed to ganglion cells (3rd-order neurons). Retinal 2nd- and 3rd-order neurons are thought to form multiple neural pathways that convey particular aspects of visual signaling such as color, motion, or shape. These diverse visual features are relayed to the lateral geniculate nucleus and the visual cortex. In contrast, visual signals leading to eye movement are sent to the superior colliculus. Classically, two retino-cortical pathways have been identified: the magnocellular and the parvocellular pathways. These pathways encode moving and stationary objects, respectively, and their existence embodies the basic concept of parallel processing1,2,3,4,5,6. Recently, more than 15 types of bipolar cells7,8,9,10,11 and ganglion cells12,13,14,15,16 have been reported in the retina of many species, including the primate retina. These cells are distinguished not only by morphological aspects, but also by the expression of distinct markers and genes8,10,17,18, suggesting that various features of visual signals are processed in parallel, which is more complicated than originally anticipated.
Cellular and molecular technologies have contributed to our understanding of visual processing and potential disease mechanisms that may arise from aberrant visual processing. Such an understanding may contribute to the development of artificial eyes. Although cellular examinations and analysis offer in-depth knowledge at a cellular level, a combination of behavioral experiments and cellular experiments would significantly augment our current understanding of minute visual processes. For example, Yoshida et al.19 found that starburst amacrine cells are the key neurons for motion detection in the mouse retina. Following cellular experiments, they performed the optokinetic nystagmus (OKN) behavioral experiment to show that mutant mice in which starburst amacrine cells were dysfunctional did not respond to moving objects, thereby confirming their cellular investigations. In addition, Pearson et al.20 conducted photoreceptor transplantation in the mouse retina to restore vision in diseased mice. They conducted not only cellular experiments, but also measured mouse behavior through the use of optomotor response recordings and water-maze tasks thus allowing Pearson et al. to verify that transplanted photoreceptors restored vision in the formerly blind mice. Taken together, behavioral experiments are strong tools to assess mouse vision.
Multiple methods are available for measuring mouse vision. These methods have advantages and limitations. In vivo ERG provides information on whether the mouse retina, particularly photoreceptors and ON bipolar cells, appropriately responds to light stimuli. ERG can be tested either under scotopic or photopic conditions21,22. However, ERG requires anesthesia, which might affect the output measurement23. The optokinetic reflex (OKR) or optomotor response (OMR) is a robust method to assess contrast sensitivity and spatial resolution, both functional components of mouse vision. However, OKR requires surgery to attach a fixation device to the mouse skull24. OMR requires neither surgery nor mouse training; however, it requires training to allow an experimenter to subjectively detect subtle mouse head movements in response to a moving grating in an optic drum 25,26. Pupil light reflex measures pupil constriction in response to light stimuli, which does not require anesthesia and exhibits objective and robust responses 19. Although the pupil reflex simulates retinal light response in vivo, the reflex is mediated mainly by the intrinsically photosensitive retinal ganglion cells (ipRGCs) 27. Because ipRGCs represent a small minority of RGCs and do not serve as conventional image-forming ganglion cells, this measurement does not provide information pertaining to the majority of ganglion cells.
The looming light experiment has not previously been considered a major test for measuring mouse vision. However, it is also a robust and reliable vision test across various species, such as mouse28,29, zebrafish30, locust31,32, and human33,34,35. Importantly, the looming experiment is one of only a few methods to test the image-forming pathway - it is not a reflex pathway - given the visual and the limbic systems in the central nervous system are involved in this circuit36,37,38. We have established a looming visual stimulus system and have demonstrated its ability to elicit motion detection in the mouse, which we use as a proxy to assess the intactness of the mouse visual system.

<table border="0" cellpadding="0" cellspacing="0" style="width:1349px;" width="1350">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:299px;">10.1&#34; monitor (2&#176; display)</td>
			<td style="width:148px;">Elecrow</td>
			<td style="width:903px;">Elecrow 10.1 Inch Raspberry Pi 1920x1080p Resolution Display</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">14&#34; Business Class Laptop 5490</td>
			<td>Dell</td>
			<td>84 / rcrc961481-4860836</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">20&#34; x 50&#34; Absorbant Liners</td>
			<td>Fisher Scientific</td>
			<td>AL2050</td>
			<td>works well to protect floor of arena, could use any type of liner</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">21.5&#34; monitor (1&#176; display)</td>
			<td>Acer</td>
			<td>Acer R221Q bid 21.5-inch IPS Full HD Display</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">CCD Camera</td>
			<td>Lumenera Corporation</td>
			<td>Infiniyy3S-1UR</td>
			<td>excellent for behavioral studies due to high fps rate (60 fps)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Enclosure (alminum frames and PVC panels)</td>
			<td>80/20 Inc.</td>
			<td>4x cat.#9010, 4x cat.#9005, 1x cat.#9000, 5x cat.#65-2616</td>
			<td>excellent, used quick build tab to find PVC, joints, and frame</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Ethanol</td>
			<td>Fisher Scientific</td>
			<td>22-032-601</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Excel Spreadsheet Software</td>
			<td>Microsoft Office</td>
			<td></td>
			<td>user friendly and widespread knowledge of Microsoft Office software</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Freearm</td>
			<td>Amazon</td>
			<td></td>
			<td>used to mount camera to the table, could use any mountable extendable arm</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">ImagePro Premiere 3D</td>
			<td>Media Cybernetics</td>
			<td>version 9.3</td>
			<td>good program, could use some updating with the automated tracking feature</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Matlab software (Psychotoolbox 3)</td>
			<td>MathWorks</td>
			<td>Matlab R2018b 64-bit (9.5.0.944444)</td>
			<td>excellent software to generate pattern stimuli of any conditions</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">SteamPix sorftware</td>
			<td>Norpix</td>
			<td>StreamPix 7 64-bit Single Camera</td>
			<td>works well, a few problems with frame dropping but good customer service</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">WD My Book External Hard Drive</td>
			<td>Western Digital</td>
			<td>WDBBGB0080HBK hard drive 8 TB USB 3.0</td>
			<td>necessary if using .avi files with no compression codec due to large size of files</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Wide angle lens</td>
			<td>Navitar</td>
			<td>NMV-5M23</td>
			<td>excellent and necessary to capture entire arena</td>
		</tr>
	</tbody>
</table>

using looming visual stimuli to evaluate mouse vision

The visual system in the central nervous system processes diverse visual signals. Although the overall structure has been characterized from the retina through the lateral geniculate nucleus to the visual cortex, the system is complex. Cellular and molecular studies have been conducted to elucidate the mechanisms underpinning visual processing and, by extension, disease mechanisms. These studies may contribute to the development of artificial visual systems. To validate the results of these studies, behavioral vision testing is necessary. Here, we show that the looming stimulation experiment is a reliable mouse vision test that requires a relatively simple setup. The looming experiment was conducted in a large enclosure with a shelter in one corner and a computer monitor located on the ceiling. A CCD camera positioned next to the computer monitor served to observe mouse behavior. A mouse was placed in the enclosure for 10 minutes and allowed to acclimate to and explore the surroundings. Then, the monitor projected a program-derived looming stimulus 10 times. The mouse responded to the stimuli either by freezing or by fleeing to the hiding place. The mouse&#39;s behavior before and after the looming stimuli was recorded, and the video was analyzed using motion tracking software. The velocity of the mouse movement significantly changed after the looming stimuli. In contrast, no reaction was observed in blind mice. Our results demonstrate that the simple looming experiment is a reliable test of mouse vision.

A mouse with healthy eyes was placed in the enclosure and allowed to acclimate for 10 min. The arena with the monitor on the ceiling was kept under mesopic light conditions (7 x 105 photons/&#956;m2/s). During the acclimation period, the mouse explored the space and found the opaque dome as a refuge. When the mouse moved away from the refuge, video capturing started, followed by initiation of the visual stimulus. In response to the looming stimulus, most mice ran into the dome (flight response), whi...

Watch this Scientific Journal Video about Using Looming Visual Stimuli to Evaluate Mouse Vision at JoVE.com

Using Looming Visual Stimuli to Evaluate Mouse Vision

To examine mouse vision, we conducted a looming test. Mice were placed in a large square arena with a monitor on its ceiling. The looming visual stimulus consistently evoked freezing or flight reactions in the mice.

The visual system in the central nervous system processes diverse visual signals. Although the overall structure has been characterized from the retina ...

Our protocol is a vision test for mice. It is both robust and easy to replicate. While many mouse vision tests are available, the looming experiment is one of a few methods to test the image forming pathway, which is not a visual reflex pathway.While many other visual behavior assay require extensive training in animal surgery, the looming technique requires little experimental training. Begin by constructing a rectangular open lid enclosure using aluminum framing and PVC panels to house the mouse during looming visual stimuli presentation. Lay a sheet of paper to cover the entire floor of the enclosure to ensure easy cleanup between trials.Then, add an opaque shelter in a corner of the enclosure with an entrance facing the center of the arena for easy entrance and exit. Secure a camera with a wide-angle lens to a table mounted stand adjacent to the enclosure for capturing the mouse's behavior. Next, set up a computer monitor on top of the enclosure.Prepare a looming pattern using experimental software to code for an expanding black circle. Set the stimulus to begin at the visual angle of two degrees and expand to 50 degrees over 250 milliseconds and set the code to repeat the stimulus 10 times with an interval of one second. Begin by selecting a mouse for the experiment as well as three blind mice to ensure response to looming stimulus was truly a visually guided behavior.Place the healthy mouse in the enclosure and let it freely explore its surroundings for seven to 15 minutes prior to stimulus onset. 10 seconds prior to the stimulation, start the video capture on the camera. Once the mouse is away from the shelter and moving freely in the open arena, start the looming visual stimuli.Finally, transfer the mouse back to its original cage. Clean the enclosure for the next mouse by spraying the walls and refuge with 70%ethanol and wiping it down and replace the paper floor liner, if soiled. Reposition the refuge to the same initial location if moved during animal transfer and enclosure cleaning.At the computer, save the video clip for each mouse in AVI format without file compression in order to ensure no data loss during transfer to the analysis software. Then, use analysis software to track the animal's motion around the arena prior to, during, and after stimulus presentation. Calculate the velocity and the distance of the mouse from the refuge.If the image of the arena is distorted due to the video angle, correct the x and y coordination prior to the calculation. Finally, compare the parameters before and after looming stimulus onset to determine how the mouse responded to the stimuli. Whether by freezing, fleeing, or demonstrating no change in behavior.Captured video clips were analyzed using a commercial analytics software with a manual tracking function. Using the tracking feature, the mouse's position was identified in each frame of the video before, during, and after the looming stimuli. When flight occurred, the velocity abruptly increased and the distance to the shelter reduced accordingly.In contrast, the velocity was near zero millimeters per second when mice froze. The velocity was reduced compared to the control before looming and the distance to the dome did not change during this period. It is important to remember to get a baseline of movement prior to the stimulus onset or a quantitative reaction to the stimulus cannot be determined.It is important to conduct multiple tests for accurate vision evaluation. Pupil light reflex is one commonly used method, which ensures that the mouse has healthy eyes. The mechanism of looming response is not fully understood.This technique may provide opportunities to explore mechanisms of moving object recognition and fear responses for neuroscientists.

Research

JoVE Journal

Behavior

Using Eye Movements to Evaluate the Cognitive Processes Involved in Text Comprehension

The present article describes how to use eye  tracking methodologies to study the cognitive processes involved in text  comprehension. Measuring eye ...

Irrelevant Stimuli and Action Control: Analyzing the Influence of Ignored Stimuli via the Distractor-Response Binding Paradigm

Selection tasks in which simple stimuli (e.g. letters) are presented and a target stimulus has to be selected against one or more distractor stimuli are ...

Methods to Test Visual Attention Online

Online data collection methods have particular appeal to behavioral scientists because they offer the promise of much larger and much more representative ...

Recording Mouse Ultrasonic Vocalizations to Evaluate Social Communication

Mice emit ultrasonic vocalizations in different contexts throughout development and in adulthood. These vocal signals are now currently used as proxies ...

A Method to Quantify Visual Information Processing in Children Using Eye Tracking

Visual problems that occur early in life can have major impact on a child's development. Without verbal communication and only based on observational ...

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 ...

Using a Split-belt Treadmill to Evaluate Generalization of Human Locomotor Adaptation

Understanding the mechanisms underlying locomotor learning helps researchers and clinicians optimize gait retraining as part of motor rehabilitation. ...

Using Rapid Serial Visual Presentation to Measure Set-Specific Capture, a Consequence of Distraction While Multitasking

This method uses a rapid serial visual presentation (RSVP) paradigm to measure the cost of distraction when participants maintain multiple search goals. ...

Using the Visual World Paradigm to Study Sentence Comprehension in Mandarin-Speaking Children with Autism

Sentence comprehension relies on the ability to rapidly integrate different types of linguistic and non-linguistic information. However, there is ...

Multi-Modal Signals for Analyzing Pain Responses to Thermal and Electrical Stimuli

The assessment of pain relies mostly on methods that require a person to communicate. However, for people with cognitive and verbal impairments, existing ...