 Λοιπόν, βλέπουμε ότι είναι εξαιρετικά. Καλήντες, καλήντες, καλήντες, ευχαριστώ και καλήντες για έναν άλλο σύστημα της Σασεξ-Βυζιον-Σέμινας, όπως πάνω στην πρόκληση του World Wide Neuro. Είμαι Γεώργος Καφέτζις, κύριε του Σεφαγγύρου, από το Μαστος Ωηλίου και τώρα ένας ΠΕΑΤΚΟΤΗΤΗΣ, με τον Μπαντεν. Είναι η πρόσφυγη σας να θέλω να ξεκινήσουμε πίγου, να σημαίνει ότι η Ελλάδα και η Παντοσποσδελία θα απαίνουν να αναφέρει αυτή την ευρωπακτική ενίσμενη φορά από την Ευρώπη και πολύ πιο ασυσίωσης της Σεμμιναρκού. Από την ανθρώπιση αυτή, παρακληθούσαμε, βρισκόμαστε, να δήσουμε για την που έγινε εδώ, και να δημιουργήσουμε το Θεό από τη Βασίντον στον Σέντι Λουίσ, Προφέστος και κυρίως για τη δημιουργία της Φαλμολογίας και της Βασικασίας, Προφέστος Νερκίστας Κερσανστίνερ. Μετά από τη δημιουργία του Βασικού, νερόλογη και νεροσάινση από τη Δημιουργία Γετγέν, Νερκίστας κοινόταν με τον Μαρτιν Στοκκέρ, στο UCL, που διεθνούσε μεταλλασπέτρας από τη Ποτασιογκλωτική Βασικασία, και πώς they influenced signal processing. Στο 2005, κυρίως για τη Σιάτολ και τη Δημιουργία Γετγέν, για κάποιες εξεστές χρόνια στην λάμπα της Ρέτσου Βόγκ. Στο 2009, κυρίως στη Δημιουργία Γετγέν στο Σαντ-λουίστας, ως προσέντης πρόσφεστος, και πέρανται από όλοι, τώρα κυρίως για το τίπορ της Φαλμολογίας και της Βασικασίας, στους δημιουργίες της Ποτασίας, και στους δημιουργίες της Βασικασίας. Στο Σαντ-λουίστας, they employ an interdisciplinary approach and investigate all things visual, from development of circuits and circuit-specific patterning, to identifying novel cell types and their behaviorally relevant functions, where they send projections in subcortical regions, and how to restore the physiologically understood function when that is disrupted by disease. Reading directly from his lab's website and aptly summing it up, the goal is to understand, preserve, and restore vision, and today I believe we will have the pleasure of hearing about their former and their latest findings and understanding vision in his talk entitled vision for predation. So without any further ado from my side, please all welcome Professor Kersensteiner. Daniel, the stage is officially all yours. Okay, thanks a lot. Let me share my screen. Okay, I'm guessing that still all works, right? Okay. So thanks so much George for that very kind introduction and for the invitation to present in this series from which I've learned a lot as a viewer. And so thanks also to you and to Tom for organizing the series and to the other participants for sharing their work. So today I'll talk about vision for predation and the reason we study this topic is that predation is one of the key drivers of animal evolution and a highly complex visual task. And because it is both essential for survival and visually demanding, we expect that predation shape many specializations in visual systems including our own and that understanding how visual systems guide predation will help us understand these specializations which of course determine visual performance in other contexts as well. So the overarching conclusions of my talk are that we find specializations across multiple levels of neural and behavioral organizations that interact in intricate ways to guide predation. So first at the level of neural diversity we identify specific neuron types in the retina and its targets which assemble into cell type specific pathways that guide predation. Second at the level of feature preferences we find neurons with complex feature preferences in these pathways that use only a subset of their features to guide predation. Third at the level of topography we find multiple regional specializations in the retina that align to guide predation. And finally at the level of behavior we find that the viewing strategies during predation focus prey onto these regional specializations of cell type specific pathways to guide predation. So now before I get into the data that support these conclusions let me briefly introduce you to the visual system the early visual system of mammals which is where the stories that I'll tell you take place. So vision begins in the retina which is a part of the central nervous system that sits at the back of the eye looking out onto the world and I'll use this simplified schematic to outline briefly the flow of information through this circuit. So light falls onto the retina here and traverses the circuit before it's absorbed by the photoreceptors which translate changes in photon flux into changes in glutamate release. This synaptic signal is picked up by second order neurons called bipolar cells which relay photoreceptor signals from the outer to the inner retina where they innovate retinal ganglion cells the ganglion cells integrate those inputs and if a threshold is crossed fire action potentials that travel down their axons and these axons form the optic nerve and are the sole source of visual information to the brain. In addition to this vertical pathway there are two classes of interneurons horizontal cells and amocrine cells which contribute to visual processing in the outer and inner retina respectively. Now importantly within each of these five neuron classes there are many distinct neuron types such that the retina overall comprises more than 120 cell types that assemble into about 45 circuits that each extract specific features of the visual environment. Now diversity is maintained beyond the retina in that the 45 or so retinal ganglion cell types that encode the output of retinal computations innovate more than 50 brain areas. And the work I'll tell you about today is part of a larger effort to try and understand what features particular retinal circuits extract and how and where in the brain they sent this information to guide behavior and generate perception. And this matching of visual computations to behavior is also called visual neurorathology. And so today's talk is very much the sequel to a talk I gave previously in this worldwide neuro forum in which I discussed both predatory evasion and prey capture in mice. And today I'll focus on predation and I'll tell you two unpublished stories about the visual pathways that guide predation in mice and then I'll end with a comparative analysis of the hunting behavior of mice and an evolutionarily distant specialized predator hunting the same prey. So let me start briefly by summarizing the main findings of the study on predation that I previously discussed because these findings raised the questions that the first story I'll tell you about addresses. So several years ago Jennifer Hoy and Chris Neill discovered that mice use vision to hunt insects in particular crickets and we followed up on this work by tracking the interactions of mice and crickets in a common 3D reference frame. This allowed us to distinguish three phases of the hunting behavior in mice. The first of these is exploration during which mice are neither approaching nor in contact with the cricket. The second is approach which is when mice are running at speed toward the cricket. And the third is contact which is when the mice are within striking distance of the cricket and frequently bite or grab it. Now one thing we did was to map the distribution of the cricket within the head-centric visual field during these phases. And we found that while during exploration the distribution of the cricket is rather broad and diffuse during both approach and contact mice keep the crickets precisely within their binocular visual field which encompasses about 40 degrees in asymus. So mice pursue prey with their binocular visual field. Now binocular processing in mammalian brains relies on the presence of ipsilaterally projecting retinal ganglion cells which occupy this temporal zone of the binocular area. And we discovered that in mice only a subset of the 45 or so retinal ganglion cells about 9 of the 45 have ipsilateral projections and can mediate binocular vision downstream. Moreover, when we removed these ipsilaterally projecting retinal ganglion cells we found that this disrupted predation. But what we didn't know was which of these 9 ipsi-RGC types matter and which of the many retinal recipient targets and the many different neuron types within they send signals to to guide predation. And so these questions were addressed by a group of people in my lab Carmela Vitale, Jenna Croissan and Keith Johnson. Now a near universal sign of functional binocular vision is the presence of acute zones in the retina. So areas of the retina where the density of retinal ganglion cells is increased and the dendritic and receptive fields are proportionally smaller allowing for higher spatial acuity vision. Now if you look at the distribution of all ganglion cells in the mouse eye there isn't an obvious acute zone but several years ago Adam Blaker when he was a graduate student with Rachel Wong discovered that the sustained on and off alpha ganglion cells which we also identified in the ipsilaterally projecting set have a cell type specific acute zone in the temporal retina that's bisected here you can see by the ipsilateral decosition line. Moreover home granite all were able to more precisely map the position of the cricket in the retina during predation and found that this area which they call the functional focus precisely aligns with this sustained on alpha acute zone. So we therefore hypothesized that the sustained on and off alpha ganglion cells in particular their binocular acute zone might be important for predation. So to test this hypothesis we wanted to specifically target the ipsilaterally projecting sustained on and off alpha cells to be able to ultimately eliminate the binocular coverage of this acute zone. And so we use the following intersectional strategy to do this. So we had previously shown that this serotonin creeline originally described by Eric Williams lab labeled these 9 ipsilaterally projecting ganglion cell types. We then confirmed that a flip line targets the same cells and this was important because it allows us to intersect it with a KCNG for creeline originally characterized by Zhinduan when he was a postdoc with Josh Sains, which labels all the alpha cells. And you can see that in the intersection this should label only this ipsilaterally projecting sustained on and off alpha cells. And so we crossed these two orthogonal recombinases to an intersectional reporter line. And you can see that this labels a specific set of cells in this temporal ipsilaterally projecting retinal ganglion cell zone about 260 cells in each eye. And if I scroll through this confocal movie, you can see that the dendrites of these cells stratify in two bands and seem to have a homogeneous morphology. You can also see that a number of these ganglion cells have their cell bodies displaced into the inner nuclear layer, which is a well-known feature of ipsilaterally projecting cells. Okay, so we wanted to confirm the selectivity and specificity of this labeling. So first we asked whether indeed all the genetically labeled cells in this intersection project ipsilaterally. So to test this, we injected retrograde retrobeats into LGN and superior colliculus on one side of the brain and then looked at the retinas. And what you can see here is that all the genetically labeled cells have the retrograde tracer in the ipsilateral eye, but not in the contralateral eye. So this argues that the intersection labels exclusively ipsilaterally projecting ganglion cells. Next, we wanted to confirm that it labels specifically alpha cells. So we stained for two alpha specific markers, SMI32 and SPP1. And again, we found that these overlap nearly perfectly with the genetic labeling. So the intersection labels ipsilateral cells, it labels alpha cells. There are both transient and sustained versions of alpha cells. And so to distinguish between these, we patched the labeled cells under two photon guidance and found that they are either sustained on alpha cells, which I'm showing you an example of here. So these cells have high baseline firings and then two spots of increasing size at light on. They increase their firing rate with little surround and they suppress it at light off. And they're also showing this chirp stimulus here. They're able to follow high temporal frequencies and are highly contrast sensitive. And the other cell type we find are the sustained off cells and here are their responses. So we established that this intersection label specifically this very small set of cells 260 per eye that are the ipsilaterally projecting sustained on and sustained off alpha cells. So next, we wanted to silence these cells specifically to remove the functional binocular coverage of this acute zone and test the effect on predation. And so to silence the cell, we use the line made by Susan De Mecchi, which in the intersection of Cree and Flip, expresses tetanus toxin. Tetanus toxin is a bacterial protease that cleaves the vesicle associated membrane protein 2 and thus blocks vesicle fusion and neurotransmitter release. And so we can stain for BAM2 to confirm that this is working. Here this is done. Here you're looking at superior colliculus in an animal where we injected the enterograte tracer CTB in one eye and you can see in the contralateral side where we shouldn't have a manipulation, the VAMP is still present in the retinal ganglion cell axons while on the ipsilateral size, the retinal ganglion cell axon terminals are devoid of VAMP arguing that this works at silencing these cells specifically. So then we looked at the effect of this on predation. So on the left here is a control mouse and on the right is a mouse in which we have silenced the specific group of cells. You can see the control mouse rather efficiently catches the cricket while the mouse in which we've silenced these ipsilatorally projecting sustained on and off alpha cell seems to struggle. It struggles in detecting the cricket and it also seems to struggle in making successful approaches and contacts. So this is quantified in the next slide here and I'll use this sort of representation repeatedly as I'll explain it on the top will always be the cumulative probability of all the trials. So each animal is given three to four trials and the data on the bottom are summaries where each data point is the average of individual animal. And so you can see overall the capture time is greatly increased in these ipsi tetanus toxin mice and this is because they have a deficit in detecting the cricket which we measure here by the interval between approaches. So from the end of one approach to the beginning of the next and then also they seem to have deficits in converting approaches into contacts and contacts into captures. So this is a deficit in pursuit whereas this is a deficit in detection. Okay. So it seems that this binocular coverage of the cell type specific acute zone functionally is important and that these cells likely guide predation. The second question I raised was which retinal recipient targets and which cells in these targets do they talk to to guide predation? So here I'm showing you again this intersectional labeling and using it to show that these ipsilaterally projecting sustained on and off alpha cells project strongly both to the dorsolateral geniculate nucleus and superior colliculus the two major image forming retinal recipient targets. So this raises the question of whether the ipsi sustained alpha ganglion cells guide predation through the retinal genicular cortical pathway or through the retinal collicular pathway. So to distinguish between these alternatives we injected pharmac viruses expressing pharmacogenetic silencers either into primary visual cortex so silencing the retinal genicular cortical pathway or superior colliculus. And what you can see here is that silencing visual cortex does not affect predation. So neither the overall capture time nor apparently the detection measured again by the approach interval or the pursuit measured by the number of approaches here. And so this in itself is an interesting finding that visual cortex does not seem to matter for predation under the conditions in which we study it. Importantly we were able to use a visual cliff test of depth perception to confirm the efficacy of our manipulation. By contrast, pharmacogenetic silencing of superior colliculus greatly deteriorates performance in predation. It both lengthens the overall capture time and increases the approach interval and makes the mice need more approaches to ultimately catch the cricket while this does not have any effect on this issue on this assay of depth perception. Okay, so it seems this is stained on and off alpha cells talk to superior colliculus to guide predation. There are a number of different cell types in superior colliculus that can be genetically distinguished. And several studies had indicated that maybe narrow field cells which are targeted by this GRP creeline may receive input from sustained on and off alpha ganglion cells. And so we tested this using this retrograde rabies virus tracing strategy where we use the creeline to limit the expression of a helper virus here. And then about four weeks later, we inject a rabies tracer that can jump one synapse upstream and then we patch the rental ganglion cells labeled by this approach to identify their type both morphologically and functionally. Here I'm showing you an example of a sustained on alpha cell that was labeled in this way. And indeed we find that about half the cells that are labeled retrograde as innervating the narrow field cells are sustained on or off alpha cells. So arguing that these provide strong input to this target cell type. So we wanted to therefore then silence the narrow field cells to see if they are the conduit of the information from the sustained on and off alpha cells for predation. So we did this with this credependent pharmacogenetic silencer again in this GRP creeline which targets the narrow field cells. And so here are movies of the same mouse on the left and on the right on the left it's injected with PBS and on the right with the pharmacogenetic silencing tool. Sorry, the movie on the left is not playing for me so let me briefly get out of my. See if that rectifies it. Okay, yeah, okay. So the mouse on the left that was injected with PBS you will see catches the cricket somewhat quickly whereas the mouse on the right struggles. I mean, it's the same mouse again just injected with CNO. And similar to what we saw with the silencing of the epsilon really projecting sustained on and off alpha cells it struggles both apparently in detecting the cricket as well as in the approach phase of the behavior but eventually you will see that it does succeed in catching and consuming the cricket. Okay, so this is for the quantified here. So again, the overall capture time is increased. Again, there seems to be a deficit in detecting the cricket as measured by the approach interval. There also is a deficit in converting contacts into approaches into contacts and contacts into captures. So in the pursuit phase of the behavior and these data are consistent with previous findings from Jennifer Hollande. So overall, what I've shown you in this part is that binocular vision mediated by the sustained on and off alpha cells in their cell type specific acute zone guides predation likely through the narrow field cells in superior colliculus. Now I've been thought for a while that acute zones and binocularity are important for predation and in fact, it had been hypothesized that both of these specializations arose in evolution because of predation. But of course, it had been difficult to test the behavioral significance of the confluence of these specializations and that's what we were able to do here to selectively eliminate the binocularity or the functional binocular coverage of this cell type specific acute zone and the fact that although this is a very small manipulation in terms of the fraction of ganglion cells manipulated that it has a very high effect or a very big effect on predation I think supports this evolutionary argument. Okay, so we have this cell type specific binocular acute zone which guides predation and this only works if mice keep the crickets precisely focused on this area which raises the question how what drives the what maintains this what drives the head and eye movements that maintain this functional focus while the mice are running quickly and while they are pre-targets are quickly changing directions. Now as mice run forward this functional focus also aligns with the optic flow field that so it emanates from the same point and several years ago Shai Sabah and David Burson had shown that the direction preference axes of direction selective ganglion cells bent toward the singularity of this optic flow field in alignment with the optic flow field and so we were hypothesizing that maybe the direction selective signals from the retina might help in maintaining the functional focus and aligning it with the running direction and this hypothesis was tested by Jenna Krozan and Ning Shen in my lab. So direction selective signals in the retina rely on asymmetric inhibition from starburst amocrine cells to direction selective ganglion cells and here are just a few of the groups that have shown this causally. So we chose to eliminate retinal direction selectivity by ablating starburst amocrine cells using again a strategy in which we express the diphtheriotoxin receptor under control of ultimately chat crease so align that label starburst cell specifically here and then we can use we can inject diphtheriotoxin into adult mice and eliminate these cells. This works quite effectively as you can see here evidenced by the lack of chat staining in the lower panels. We can also confirm that this eliminates direction selectivity in the retinal ganglion cells by doing this in the background of a line in which the retinal ganglion cells specifically that prefer motion in the direction generated when mice are running forward are labeled, so the DRDEGFP line you can see normally these cells respond to drifting grating in their preferred direction strongly but in the opposite direction very little or not at all and when we do the same recordings in the chat DTR mice now these cells respond equally in both directions this is true for a large number of cells so this index of direction selectivity is greatly reduced. We can also confirm that this worked at a behavioral level so it's well known that the optokinetic gaze stabilizing eye movements are reliant on direction selective signals from the retina and so here we're testing the optokinetic nystagmus and you can see that it's abolished in these chat DTR mice. Okay so we functionally removed direction selective signals from the retina the question is what does this do to predation and the answer is very little so both the overall capture time seems to be unchanged they don't seem to have a deficit in detecting the crickets we didn't really expect this but they also seem to have not any changes in the pursuit phase of the behaviors and either again any deficits in converting approaches into context or context into captures and then even they don't seem to have a difference in maintaining the cricket within the functional focus shown here by the distribution of the cricket as a function of ademas in the visual field of mice so this was surprising to us and surprising to a degree that we started to worry about whether it might be a consequence of the specific experimental timeline we used so let me explain that a little bit so in these experiments we inject diphtheria toxin into adult chat DTR mice binocularly and then we wait for about a week before we introduce crickets into the home cages and then for three days and then for the next four days we start to food deprive mice and adapt them to the hunting arena before we then test them so in this experiment likely mice are experiencing crickets and learning to hunt them while direction selectivity from the retina is already removed and so we were worried that they might use alternative strategies to what they normally use and so we were wondering whether we can look a little bit more closely at the time course of starburst amocrine cell deletion and whether that might allow us to convert this into an experiment where we could train mice and adapt mice in the presence of retinal direction selectivity and then test them specifically when they don't have it okay so we looked at the time course of starburst ablation after diphtheria toxin injection at day zero and you can see up to day eight there's really not much of a change in the number of starburst cells either the on starburst or the off starburst but then over the next two days the starburst cells drop off precipitously as shown here so this is really quantified here across several animals you can see that everything is pretty normal up to around day seven or eight and then there's this precipitous drop in particular from day nine to day ten now of course it could be that the the functional deficit precedes the the overt anatomical removal of these cells and so we also head played at mice at day zero when we injected them and then measured the optokinetic nystagmus again as a functional readout of retinal direction selectivity over the subsequent days and you can see that this similarly goes through a very stable phase until about maybe day eight or nine when it drops off precipitously proceeding the time course of the anatomical removal but in a very narrow window and so we modified our behavioral protocol as follows we injected at day zero we then adapted mice to crickets in their home cages from day two to four then trained them over the next five days before we test them and this should mean that during all this period the mice should have normal direction selectivity and learn to hunt the cricket but on the test day all the starboard cells are gone and the optokinetic nystagmus is gone indicating that on the test day they should have no retinal direction selectivity so then we looked at predation at this test day and again there was very little change between the chat DTR and the control mice both in the overall capture time as well as in the detection phase of the behavior measured again by the approach interval or the pursuit phase of the behavior measured by these conversion probabilities and then again these mice also do not have any deficits in maintaining the cricket within the functional focus so one of the reasons these findings are surprising is that the narrow field cells which I've shown you guide predatory behavior in mice do not are well known to be direction selectivity and they're thought to be direction selective in part because they're thought to get input from on-off direction selective ganglion cells so we first wanted to confirm this connectivity and so we used again the same retrograde rabies virus tracing strategy that I've described to you and indeed I told you already that about half the ganglion cells we see in this way are sustained on and off alpha cells but we also find that a quarter of the ganglion cells around our on-off direction selective ganglion cells so this raised the question to what extent retinal direction selectivity in superior colliculus in these NF cells and in other cells is inherited from the retina and is therefore changed when we ablate the starburst amocrine cells and this is a question that's recently been debated somewhat and we thought we had something to contribute to this debate because we are able to remove starburst amocrine cells and eliminate retinal direction selectivity acutely bypassing some of the developmental effects that have been an issue of discussion here so this question was addressed by Zhaiing Fengfeng Song and Mike Fitzpatrick in my lab and Zhaiing Feng recorded the activity of neurons in superior colliculus in a wake head fixed mice running on a cylindrical treadmill in this visual immersive visual stimulation dome we showed two types of stimuli first a sparse noise stimulus to map receptive field and then this drifting grid stimulus which was originally described by JC Kang's group in which we modify the direction of the drift as well as the speed and then we compared these response parameters between the control and the chat DTR mice so here I'm showing you some example receptive field maps of neurons in superior colliculus both the on and the off receptive field of a cell in control and chat DTR and you can see that as was already known most neurons in superior colliculus respond to both on and off stimuli but have a preference for dark objects so this is calculated here by a polarity index which is minus one if the responses were purely off plus one if they were purely on and you can see this distribution is usually not minus one but if they were purely off the receptive fields come in a wide range of sizes which in part I thought to reflect underlying differences in the dendritic architecture of the different cell types that contribute to this data set and importantly neither of these are changed in the chat DTR relative to the control next we looked at the speed tuning and we found again consistent with other results that most neurons in superior colliculus are not moving objects over slower moving objects this is calculated here by a speed preference index where index greater than zero indicates fast preferences and you can again also see that this speed tuning is unaffected by the removal of chat DTR by the removal of the starburst cells importantly however the direction selective cells appear to be or the direction selectivity of superior colliculus in our experiments appears to be eliminated when we eliminate the starburst cells so here I'm showing you an example neuron in superior colliculus and the control that's clearly direction selective compared to a cell that is not and so a small minority of the cells about maybe 10% or so and control our direction selective consistent with other people's results but these cells appear to be or this direction selectivity appears to be eliminated in the chat DTR so here on the right maybe this is the easiest is where we're varying increasing the threshold of direction selectivity starting from a pretty low threshold to increasingly higher levels and then asking what proportion of cells cross this threshold and you can see in control about 10% of the cells maybe are direction selective but in chat DTR really hardly any are we can then take advantage of the fact that the narrow field cells have the smallest dendritic arbors and are known to have the smallest receptive fields usually if you look at cells that have a receptive field side of less than 200°2 you can be sure that you're looking at narrow field cells so we restricted this analysis to only cells whose receptive field area was less than 200° you can see that this increases the proportion of direction selective cells in controls to about 20% but again this direction selectivity appears to be eliminated and the chat DTR so it does suggest that the direction selectivity in superior calculus including of the NF cells is inherited from the RAT9 is eliminated when we ablate the starware cells in this acute fashion in adult mice so we next just confirmed that even when we use a cell elimination strategy for the NF cells but indeed this also not just the pharmacogenetic silencing affects predation so we injected this cre-dependent virus expressing caspase 3 into the GRP line which targets the NF cells you can see again this very reliably generates a big deficit in predation increasing the overall capture times increasing the interval between approaches so introducing deficits in prey detection as well as deficits in prey pursuit as again indicated by these conversion probability although this also does not have any deficit in maintaining the functional focus so what I've shown you in this part is that the narrow field cells in superior calculus do seem to guide predation that's evidenced both by the pharmacogenic silencing and by the ablation but they appear to do so independent of their direction selectivity which we show they inherit from the retina so I think this is an interesting example of what I might call behavioral multiplexing of feature preferences so the NF cells have complex feature preferences they prefer small things over big things dark things over bright things fast things over slow things and are direction selective and it seems that their direction selectivity although often this is considered their cardinal feature is expendable for their role in predation so of course the question is kind of what other features matter and what is the direction selectivity of NF cells for but that will have to remain for another day okay so in this study so far we used a variety of genetic manipulations and viral manipulations to establish causal links between neural specializations and visual behavior now an orthogonal approach to trying to establish such links is to study how different species that may have diverged in evolution in the neural specializations perform in the same visual behavior and so with that in mind I want to introduce you to the evolutionarily distanced specialized cricket hunter that is the fat-tailed dunnardos and subsos crassical data and the study I'll tell you about is a collaboration between Linda Richards-Splat who brought these animals to us so Linda Willmott and Saranja Paul in Linda's lab and Jenna Croissant in my lab so it's worth remembering or learning for the first time that marsupials which these animals are diverged from eutherian mammals like mice about 160 million years ago allowing for significant time for divergence just for scale the last common ancestors of mice and humans lived about 87 million years ago so interestingly one of the areas of divergence and specializations between these animals is binocular vision so here I'm just recapping a couple of the parameters of mice so mice overall have a visual field about 320 degrees of which a fairly small region is binocular and they also have an acute zone which occupies about the same space in terms of degrees of visual angle and they seem to have a pretty meager lateral projection as I illustrated where only about 20% of the ganglion cells temporal of this dequisition line have ipsilateral projections by contrast the dunnards are real extremists in terms of visual field so they both have very large overall visual field 360 degrees with the exception of some blind spots but they also have 140 degrees of binocular vision meaning that actually each eye encompasses 250 degrees of visual space so they have a much larger binocular visual field they also have a more robust ipsilateral projection where about 70% of the ganglion cells temporal of the dequisition line have ipsilateral projection and then they have a smaller acute zone but also a much more significant acute zone in terms of the enrichment of ganglion cells that occupies about 25 degrees of visual space so significant divergence in these specializations which I've previously discussed with you might play a role in predation so we wanted to compare the hunting behavior of dunnards and mice and so here I'm just going to show you two movies illustrating that dunnards are very efficient predators so in this first movie you will see the dunnard catch a cricket sort of out of mid air and I should mention that the movies I've shown you for mice were sped up about one and a half while these movies are in real time this is the second one where you'll just see it make a very efficient approach okay so these movies are indeed representative so overall the dunnards are much faster at catching the cricket this is the overall capture time here again are all the trials and then this is on a per animal basis so of course this could be trivially due to differences in sort of the running speed of these animals but it could be that the dunnards are much faster runners or they turn much more quickly and that this is what mediates this improved performance rather than any differences in their visual systems but this does not seem to be the case so here I'm showing you the distribution of the running speed of dunnards and mice during the approach phase of the behavior and you can see that this is not different and in fact if we look at the speed with which the animals turn their heads in both cases mice have the upper hand so mice are able to turn their heads more quickly and turn their bodies more quickly so clearly this cannot account for the differences or the better performance we see in the dunnards instead we suspect that this might be due to their visual systems now again I distinguished in terms of this visual task two phases of the behavior one detection and two pursuit and so first we looked at detection which again we previously already I explained we measure by either the latency to the first approach or the interval between subsequent approaches and there's a small difference here but not at the level of the animals so it does seem like there isn't much of a difference in the prey detection between mouse and dunnards by contrast when dunnards start making an approach so in the pursuit phase of the behavior they seem to be much more efficient and as a result they need many fewer approaches to catch the cricket and have a much higher probability of catching the cricket once they initiate an approach so this indicates that the visual pursuit is much more effective in dunnards and so the next question which we thought we could answer here was whether it is the overall size of the visual field that matters this much larger binocular visual field than the acute zone and this is interesting because in mice like I mentioned both of these areas are roughly the same size and if we look at the distribution of the cricket within the head centric visual field you can see that it aligns of course with both but in dunnards the binocular region is much larger than the acute zone and so here when we now look at the distribution of the cricket we can see that even though the dunnards have much larger binocular visual field they still keep the cricket precisely within their acute zone so this also highlights the importance specifically of the acute zone for predation behavior we think and here this is quantified as the full width with a half max of these curves across all the different animals a final difference in the behavior of mice and dunnards is that mice have this behavior which is called tigmataxis which means that they stick close to the wall even during the approach so here I'm showing you the distribution of all the mice during the hunting behavior during the approach phase of the hunting behavior in the arena and you can see that they really stick to the wall whereas the dunnards are much more willing to attack crickets across open spaces this is further quantified here as the distance to the boundary during the approach phase and you can see this is much right shifted in the dunnards so to conclude this comparative analysis I've shown you that the fat tailed dunnards are more efficient predators than mice but this increased efficiency cannot be accounted for by differences in the running speed or in the speed with which they turn nor in the ability to detect prey but rather seems to be due to a more efficient visual pursuit which they seem to mediate by focusing the cricket on their acute zones and the final difference was that dunnards are more willing to cross open spaces to attack of course we are now also studying the visual systems of these animals to correlate these behavioral differences with the neural specializations further and with that I want to thank everyone who did the work I think I mentioned everyone involved as I went along it's a really fun group and I'm very happy to be able to work with them and grateful I mentioned our collaborations with Linda Richards Linda Wilmot and Siranjapal on the dunnards then finally I want to thank the funding sources primarily the National Eye Institute as well as the Hope Center and the Grace Nelson Lacey Fund and I thank you for watching Thank you very much Daniel for this fascinating presentation of very interesting findings very meticulous approach maybe even exhaustive I would say given the richness of tools that you are employing and trying to tackle it probably exhausting to listen to but not at all so there are already some questions appearing in the chat and I would like to remind to our audience that after this first round of moderation for me that I will try to go by chronological order in the questions that you are posing we will continue in a more informal fashion in the zoom room that we are currently using and I'm posting the zoom room link already there so the first question appearing in the chat is from democratis Karamanlis is the remaining vision outside the Ipsi sustained alpha superior colloculus pathway used to guide the occasional capture of crickets or is it more like olfactory do occasional captures also happen in darkness yeah so I mean I think the very good question so I don't we don't think olfaction plays a role essentially in our hands this behavior is purely visual so Chris Neal and Jenna Hoy when they initially described this behavior showed that in their arena which was sound isolated if they did it in the darkness there was still a reasonable performance and then when they earplugged the mice performance essentially became chance right sort of in our hands if we do it in the dark the performance is already chance because it's not sound isolated so we've also earplugged the mice and this doesn't make the behavior any worse in the dark okay and olfaction we don't think given that already removing vision makes a chance we don't think olfaction plays a role I mean I think that the performance we're seeing with the ablation of the Ipsi sustained alpha is still above chance by chance I mean sort of chance encounters where if the crickets get very close to the mouse you know through their they will sense them and then they're able to catch them right so the performance we're seeing is still better than what it would be in the dark and you know everyone is equally open to speculate about what that might be due to I mean I think it's worth remembering that we're not really eliminating even the acute zone right we're just eliminating the binocular coverage of it so there certainly is still a lot of information from this area and even from these cell types in this area coming to the brain right so I would assume that that accounts for the residual performance that we're seeing and you know I'm more surprised that we see such a strong effect at all from silence then you know 260 or 50,000 retinal gang themselves right next one appearing in the chat is from Βατσάννα Σωμαία and Apologies if I'm pronouncing the name for the first part of your talk how about the role of ύψι in VLGN in predation VLGN in mouse doesn't project to V1 so the AAV approach doesn't rule out the role of VLGN Yeah that's correct I mean so we have not tested VLGN specifically and there is a now it's not a the alphas as I showed in the sort of enter rate labeling the projection to VLGN is pretty weak there are some axons there I mean yeah we can try that but it seems sort of unlikely given what we know about the function of VLGN in general but it's certainly worth testing we've tested some other brain areas like you know one thing we were worried about in the context of the direction selective manipulation was the role of the accessory optic system so we've silenced the accessory optic system and surprisingly that didn't have any effect on predation even though I would have imagined that removing gaze stability would affect it but it didn't but we haven't tried VLGN yet we could certainly do that Next one is from Ana Vlasic I'm curious about on versus off sustained alpha cells contribution to the predation behaviors the cricket is usually dark on the white background in your task can the animal hunt a white insect on a gray background Yeah so we have not tried that that's a very interesting question so we've now gone down to these two cell types I don't think there is a way manipulation wise for us to distinguish it but I think it's making a good point and trying to change the parameters of the visual task in general I think this is an interesting area of further exploration because I also suspect that maybe for example we see that visual cortex I said does not guide predation or is not important for predation now that might change when we make the task a lot harder visually so I think it's sort of that's certainly something where we can do more and where we haven't really done manipulations of visually what the task is I think sort of Jennifer Hoy has done some work on like virtual version of the task where my seem to approach circles of a certain size that move along a screen I forget right now I'm sorry whether this was only true for dark things I sort of think it was but I'm not sure anyway reach her paper it's excellent maybe that answers the question otherwise we could try to paint the cricket which is sort of would probably be my more analog approach yeah great thank you very much for the responses thus far before I move on to the last question that appears in the chat which is from Tom who naturally asks about you know your new favorite species potentially I have one when it comes to direction selectivity abolishment so could it be that the parameters that you are quantifying behaviorally speaking do not capture a certain change for example like instead of measuring how often not how often sorry how fast the mice turn their head or their body to measure how often they would change it yeah I mean I think again we so far we have still the 3D data for these animals which we haven't analyzed completely yet so we will do that more precise mapping of whether the functional focus changes I've only shown it sort of along as a most so far from the over at camera yeah I've played around a little bit with analyzing you know just around points when the cricket has jumped and they need to change direction and these kinds of things so far none of this has really come out that come back positive but I mean I think there's almost an infinite number of things you can of course yeah but I think we'll we'll look some more but ultimately it won't obscure the fact that you know overall the performance even if you do it sort of in this acute way is really not affected right sort of so like I don't know it's a negative result but it's interesting to me and that these NF cells which are clearly important right in our direction selective are direct are important for the behavior independent of this feature that they encode right I'm not aware of a similar kind of study where you can clearly link some of the future preferences to a behavior but others or kind of so I think that to me is an interesting area to think about in terms of multiplex you know most neurons have sort of complex future preferences and anyway absolutely and like down this line so could it be that for example motion anticipation in the sense of you know how marcosmeister and others have described could be playing a role in guiding the predation itself and not necessarily like direction selectivity but rather like direction anticipation yeah I mean yeah I'm sure motion anticipation in the sense that so you know my barrier marcosmeister that's essentially kind of a form of contrast adaptation if I think I understand correctly right which means that then where the stimulus is perceived is more in line with where it might be I certainly think that could play a role here and obviously you don't need direction selective signals to know which direction something has moved so yeah there's lots of ways to imagine the system working without this intact right great thank you very much for this one question appeared from maria and it gets a thumbs up from marla feller as well maria kozan asks she says very fascinating talk thank you I was wondering could the preference of narrow field cells for faster stimuli be the key feature that is important for predation yeah I mean I think I think the three that I meant right they like small things over big things dark things over bright things and fast things over slow things those are all kind of things that align with a cricket I would say so I think those probably are what matters again it's still interesting that the other one doesn't hopefully it's interesting yeah no absolutely I mean this multiplexing and the last question appearing is as I said from Tom who asks do donuts have alpha cells is there anipsi or contra projection bias bias and how do they respond to light stimuli yeah so I mean this is the part that I decided in the end not to include in the talk so we hopefully soon we'll have we've now recorded maybe 10 of these animals on MEAs and are classifying their ganglion cells they seem to have some cells that are a bit alpha like they have DS cells there are other things but a lot remains to be characterized in terms of ipsilateral projection we've not read this yet but what people have shown is that the proportion of cells that have ipsilateral projections in this area is higher I think I had it on a slide there it's 70% rather than 20% in mice right and a 20% in mice correlates with not enough the 45 types so I assume that the 70% also correlates with a higher proportion of cell types that have ipsilateral projections right and it's more in line with what it would be in cat for example right and cat also not all that in primates all the cells cross on the temporal side of the decossation line and cats it's more of that 70% and probably DS cells don't so we'll have to get more into that so we're going to try to wrap up our initial characterization which is based on MEA I think we will soon also start to try to maybe patch specifically in this acute zone if we think that this really matters right for this behavior but yeah there's lots to be done as you guys well know when you have a species for which not a lot has been done it's sort of like difficult to pick a direction yeah and I'm sure like more questions will follow from Tom's side once we stop broadcasting so given that there are no more questions appearing in the chat I would like to thank you once again for this fascinating talk and thanks of course to the audience for being here for yet another seminar of ours so I'm going ahead posting the zoom room link once again and ending the stream so we can continue okay thanks everyone thanks George