Columnar neurons support saccadic bar tracking in Drosophila
Curation statements for this article:-
Curated by eLife
eLife assessment
This paper provides valuable new insight into the neural encoding and behavioral tracking of visual objects in the Drosophila. It provides solid evidence that a specific type of neuron in the fly visual system (T3 neuron) is involved in the tracking of moving objects during flight. With additional experimental evidence to resolve whether T3 neurons function as local object detectors, this paper would be of broad interest to visual neuroscientists.
This article has been Reviewed by the following groups
Listed in
- Evaluated articles (eLife)
- Preprints for generating images with AI (BlueReZZ)
Abstract
Tracking visual objects while maintaining stable gaze is complicated by the different computational requirements for figure-ground discrimination, and the distinct behaviors that these computations coordinate. Drosophila melanogaster uses smooth optomotor head and body movements to stabilize gaze, and impulsive saccades to pursue elongated vertical bars. Directionally selective motion detectors T4 and T5 cells provide inputs to large-field neurons in the lobula plate, which control optomotor gaze stabilization behavior. Here, we hypothesized that an anatomically parallel pathway represented by T3 cells, which provide inputs to the lobula, drives bar tracking body saccades. We combined physiological and behavioral experiments to show that T3 neurons respond omnidirectionally to the same visual stimuli that elicit bar tracking saccades, silencing T3 reduced the frequency of tracking saccades, and optogenetic manipulation of T3 acted on the saccade rate in a push–pull manner. Manipulating T3 did not affect smooth optomotor responses to large-field motion. Our results show that parallel neural pathways coordinate smooth gaze stabilization and saccadic bar tracking behavior during flight.
Article activity feed
-
-
Author Response
Reviewer #1 (Public Review):
While the circuits underlying the computation of directional motion information in the fly brain are very well described, much less is known about the neurons serving the detection of objects. In a previous publication from the same lab, it has been shown that flies perform body saccades to track a moving object during flight. In the current paper, Frighetto and Frye provide evidence that T3 cells, a population of neurons within the optic lobes, are involved in this task. First, they performed 2-photon Calcium imaging from T3 cells to show that these cells respond to moving bars, which they later use in behavioural experiments. They then silenced T3 cells using genetic tools and tested the behavior of these flies in response to a rotating bar using two different setups. In one, the flies …
Author Response
Reviewer #1 (Public Review):
While the circuits underlying the computation of directional motion information in the fly brain are very well described, much less is known about the neurons serving the detection of objects. In a previous publication from the same lab, it has been shown that flies perform body saccades to track a moving object during flight. In the current paper, Frighetto and Frye provide evidence that T3 cells, a population of neurons within the optic lobes, are involved in this task. First, they performed 2-photon Calcium imaging from T3 cells to show that these cells respond to moving bars, which they later use in behavioural experiments. They then silenced T3 cells using genetic tools and tested the behavior of these flies in response to a rotating bar using two different setups. In one, the flies are fixed and bilateral changes in wing stroke amplitude are used as a measure for turning, in the other, flies are magnetically tethered such that they can rotate around the vertical body axis. Silencing T3 cells leads to the abolishment of the steering response induced by object position using a bar that is defined by its motion relative to the surround, but leaves the response to object motion intact. In the magnetically tethered flies, it reduces the number of saccades and thus leads to an impairment of bar-tracking behavior. In another set of experiments they optogenetically activated the whole population of T3 neurons (which supposedly impairs their normal function), which leads to an increase in the number of saccades after the activation (when the light stimulus used to activate the cells is turned off). Silencing the neurons necessary for detection of local motion, T4 and T5 cells, in contrast reduces responses elicited by object motion rather than position, but also has an impact on object tracking saccades. The authors provide a simple model, where speed-dependent signals from multiple T3 cells are integrated and trigger a saccade, when a threshold is reached.
The data generally support the conclusion that T3 cells play a role in detecting bar position and in controlling saccades in response to rotating bars. However, there are some inconsistencies in the data that are not sufficiently explored and discussed.
- In a previous paper from the lab (Keleş et al., 2020), it was shown that T3 cells respond preferentially to small objects, whereas here they robustly respond to elongated bars and even large-field gratings. This discrepancy is not discussed.
The most likely explanation is that Keleş et al. (2020) work used stimuli of half-contrast (or lower) to probe contrast polarity effects, whereas our stimuli here match the behavior experiments using maximum contrast broadband stimuli. Keleş et al. (2020) work also provided visual stimuli over the full display, >200-degrees in azimuth, whereas here we only provide stimuli unilaterally over <100 degrees; perhaps there was some effect of contralateral stimulation. Finally, different Gal4 drivers; here we use a split-Gal4 that is highly specific for T3. Keleş et al. (2020) work used a normal Gal4 driver less clean than the split. We shall discuss these discrepancies in revision.
- In a previous paper, the authors showed that integrated positional error rather than bar position is used to elicit bar-tracking saccades and that saccade amplitude is relatively stereotyped. However, here they show, that T3 cells respond much more strongly to a slowly moving stimulus (18{degree sign}/s) rather than to the fast moving stimuli used for the behavioral experiments (> 90{degree sign}/s). This response property plays an important role for the model they propose. My general concern here is that the findings might not be generalizable to slower moving bars, where more precise, position-dependent responses could play a larger role, and that these fast moving bar stimuli represent an extreme situation, where the flies cannot accurately track bar position any more.
We agree that flies will not accurately track purely positional cues at higher bar speeds, since responses to positional signals are inherently sluggish. In free-flight, files execute orientation saccades when a stationary post subtends ~30 degrees (bar width used here), at which point the leading edge of the post is moving ~250°/s (van Breugel and Dickinson 2012). Thus, higher bar speeds are the norm for flies, and our behavioral stimuli (90°/s) was chosen to robustly trigger tracking saccades and to compare with previously published behavioral data sets. Bar velocity of 18°/s is far below the range that robustly triggers orientation saccades. We image at 90°/s and 180°/s to show that T3 responses to behaviorally relevant bar speeds could reasonably act as inputs to an integrate-and-fire behavioral controller. These points shall be clarified in revision.
- The claim that T3 cells are tuned to stimulus velocity is not supported by the data in my view. For the bar stimuli, the authors only tested speeds of 18{degree sign}/s and above 90{degree sign}/s, but nothing in between. For the grating motion there seems to be an influence of temporal frequency for the same stimulus velocity (see e.g. Fig.1_1), but this is not quantified.
We shall add a full spatiotemporal response profile in revision. One note: we presented T3 responses to different grating speeds in Supplemental material because our goal was merely to indicate speed sensitivity by T3, rather than to present a comprehensive speed tuning curve. T3 is distinct from T4 and T5 in that it is not directionally selective, is full-wave rectified for contrast, and shows similar responses to bars of differing temporal frequencies moving at the same speed. These properties are also likely accompanied by a broad spatial frequency sensitivity (which would bestow speed tuning), but in revision shall either demonstrate this or remove claim to it.
- The results from the optogenetic activation experiments are hard to interpret, as it is unclear how a prolonged activation of all T3 cells would affect the downstream circuitry. It is not clear that this experiment is equivalent to a "loss-of-function perturbation" of T3 cells as the authors claim in the text.
We are making an assumption, which we shall clarify in revision, that downstream circuitry requires a spatiotemporal progression of columnar activity, as would be generated by the projection of a discrete bar-type-object moving across the eye, and that activation of all columnar inputs together, as would occur with CsChrimson stimulation, would disrupt this discrimination. Although it is a supposition, we feel that it is parsimonious. We compared the effect of CsChrimson stimulation under two different LED intensities but found no effect on bar tracking behavior.
Reviewer #2 (Public Review):
In their manuscript titled "Feature detecting columnar neurons mediate object tracking saccades in Drosophila", Frighetto & Frye study the effect manipulating T3 neurons has on tethered flight saccades. The authors first characterize the responses of T3 neurons to simple visual stimuli, and then manipulate T3 cells (with both Kir2.1 and CsCrimson) and study the effects on the fly's tethered flight behavior, focusing on different types of sharp turns (saccades). Finally, the authors suggest an integrate and fire model to explain how an array of T3-like neurons can produce some of the recorded behavior.
The authors study the elementary, yet challenging, computation of object discrimination. They hone in on a cell type that most likely plays an important role in the circuit. However, the authors do not sufficiently clarify the framework in which they conceptualize T3's role in object discrimination, neither when discussing it in the introduction/discussion nor when explaining experimental results. The authors present the work in comparison to T4/T5 cells. However, T4/T5 cells have been shown to be both local motion detectors and the main cell types to compute motion in the fly's eye. Downstream neurons integrate over these local units to detect different patterns of global and local motion (Authors should cite Krapp 1996 Nature). Are the authors suggesting that T3 neurons perform a similar function only as local object detectors? That is a bold claim that will need to be supported with more experimental results and reconciled with previous results. We already know of other Lobula Columnar neurons (LCs) that respond to different sizes, some even smaller than the optimal T3 stimulus (e.g. Klapoetke 2022 Neuron) and we know of LCs that respond to small objects that do not receive major inputs from T3 cells (e.g. Hindmarsh 2021 Nature).
We are attempting to posit a simple and parsimonious framework for T3 action. Are T3 neurons “local object detectors”? T3 is clearly not “selective” for local objects, since we show that they respond to elongated bars and wide-field gratings (at least when projected over the ipsilateral visual hemisphere). T3 is, however, “sensitive” to objects: vertical bars yielded a mean response peak ~1 ΔF/F whereas a small square object elicited a peak of ~4 ΔF/F (Keleş et al., 2020). This amplitude differential likely indicates surround inhibition, but does not preclude a downstream integrating neuron from pooling columnar inputs to assemble a spatial receptive field for either an elongated bar or a small object. Individual T4/T5 neurons show roughly double the response amplitude to a small object than a long vertical bar (Keleş et al., 2020), which is consistent with other reports, but one would not classify T4/T5 as “small object detectors” as they play a fundamental role in detecting wide-field motion stimuli. We intend to posit that (i) columnar T3 neurons are small-field (local) detectors of the features contained within stimuli that flies readily track, (ii) that the integration of these local signals could support the integrated error computations that flies make to track bars, which (iii) explains why T3 blockade compromises bar tracking saccades. We do not mean to claim that T3 are the first, last, or only inputs to object detection circuitry in deeper neuropiles. We shall endeavor to clarify these issues in revision.
These differences between T4/T5 cells and T3s also make interpreting the experimental manipulations more challenging. When hyperpolarizing T4/T5 or 'blinding' them with CsCrimson activation, the visual motion circuit is severely disrupted. However, the same cannot be said about inactivating/blinding T3 neurons and the object detection circuit (if it is indeed a single circuit). The authors are justified in deducing a connection between blocking T3 neurons and a reduction in bar tracking, but generalizing the results to object detection requires more experiments and clarifications.
We consider “bar tracking” to be one form of object detection, but not the only form. A bar is an “object” (albeit a tall object) in the sense that it is optically disparate from the visual surround. Thus, inactivating/blinding T3 indeed severely disrupts the detection of bar-type-objects. We shall clarify the language to remove any confusion between “object” and “bar”. We do not mean to generalize T3 function to all object vision in the same way that T4/T5 function is generalized to all motion vision, and this shall be clarified in revision.
When framing the manuscript in the object detection framework, previous results regarding the definition of an object should also be addressed. Maimon Curr. Biol. 2008 and work from their own lab (Mongeau, 2019) have already shown that tethered flies respond differently to bars and small objects (fixating on the former while anti-fixating on the latter). Previous work has also shown that T3 neurons respond strongly to small objects and suppress responses to long bars (Tanaka Curr. Biol. 2020). Since all the behavioral experiments in the current manuscript and all the visual stimuli are full arena-length bars, it is impossible to tell whether the T3 results generalize to small objects and even how to reconcile the stronger response to small objects with the role ascribed to T3 cells in generating behavioral responses to long bars.
This amplitude differential between small object and elongated bar responses by T3 likely indicates surround inhibition, but does not preclude a downstream integrating neuron from pooling columnar inputs to assemble a spatial receptive field for either an elongated bar or a small object. Consider that T4/T5 neurons show roughly double the response amplitude to a small object than a long vertical bar (Keleş et al., 2020 and consistent with other reports), but one would not classify T4/T5 as “object detectors” as their small-field columnar signals are integrated by downstream wide-field neurons that assemble spatial filters for specific patterns of optic flow that are generated during flight maneuvers (Krapp et al., 1996 Nature). One downstream integrator of T3 inputs, LC11, is more selective for small objects than T3. We shall clarify these points in revision.
Finally, the authors propose a model for a hypothetical neuron downstream of T3 that would integrate over several T3s and generate saccades. However, given the current knowledge level in the fly vision field, the model should either be grounded more in actual circuit connectivity or produce testable predictions that would guide further research.
We are currently working on the putative downstream partners of T3, and testing for the integration of T3 signals. Preliminary data show that silencing a specific LC class postsynaptic to T3 recapitulates the effects of silencing T3 on saccadic bar pursuit. In the revised version of the manuscript we will provide additional discussion.
The authors should decide whether they would like to address these concerns with more specific experiments that would shed light on the role T3 has to play under different conditions and different definitions of a visual object, or whether they would prefer to limit the scope of their claims.
We shall endeavor to do both!
Reviewer #3 (Public Review):
In free flight, flies largely change their course direction through rapid body turns termed saccades. Given how important these turns are in determining their overall behavior and navigation, it is important to understand the neural circuits that drive the timing of triggering these saccades, as well as their amplitude. In this paper the authors leverage the powerful genetic tools available in the fruit fly, Drosophila, to address this question by performing physiology experiments as well as behavioral experiments with inactivation and activation perturbations.
The authors make three primary conclusions based on their experiments: (1) the feature detecting visual pathway (T3) is responsible for triggering saccades in response to moving objects, but not widefield motion, (2) the pathway primarily responsible for wide field motion encoding (T4/T5) is responsible for triggering saccades in response to widefield motion, and (3) the T4/T5 pathways is responsible for controlling the amplitude of both object and widefield motion triggered saccades.
The authors go on to show that using calcium imaging data of T3 activity it is possible to predict under what conditions flies will initiate a saccade when presented with objects moving at different speeds, resulting in a parsimonious model for how saccades are triggered.
Together, the imaging, behavior, and modeling provide compelling evidence for claims 1 and 2, however, the evidence and modeling for point 3 - the amplitude of the saccades - is lacking. The statistical analysis does not go into sufficient detail in comparing across different cases, and in particular, there is little mention of the effect sizes, which appear to be quite small (this is primarily in reference to 3F and 4E). The data suggest that both the T3 and T4/T5 pathways contribute to saccade amplitude, instead of T4/T5 being the only or primary drivers.
We agree that the evidence suggests that both T3 and T4/T5 pathways contribute to saccade amplitude for bar tracking behavior, and shall clarify this conclusion in revision. However, we also note that the effect of silencing T4/T5 is more prominent (e.g., peak angular velocity) and more consistent across visual conditions. We will dig deeper into the data to substantiate this point. The effect sizes might be small because the silencing approach (i.e., inward rectifying Kir2.1 channels) maintains a hyperpolarized state but does not completely block neuron function; consider that the wide-field optomotor responses of T4/T5>Kir2.1 flies is reduced but not eradicated (Fig. 3A_1).
-
eLife assessment
This paper provides valuable new insight into the neural encoding and behavioral tracking of visual objects in the Drosophila. It provides solid evidence that a specific type of neuron in the fly visual system (T3 neuron) is involved in the tracking of moving objects during flight. With additional experimental evidence to resolve whether T3 neurons function as local object detectors, this paper would be of broad interest to visual neuroscientists.
-
Reviewer #1 (Public Review):
While the circuits underlying the computation of directional motion information in the fly brain are very well described, much less is known about the neurons serving the detection of objects. In a previous publication from the same lab, it has been shown that flies perform body saccades to track a moving object during flight. In the current paper, Frighetto and Frye provide evidence that T3 cells, a population of neurons within the optic lobes, are involved in this task. First, they performed 2-photon Calcium imaging from T3 cells to show that these cells respond to moving bars, which they later use in behavioural experiments. They then silenced T3 cells using genetic tools and tested the behavior of these flies in response to a rotating bar using two different setups. In one, the flies are fixed and …
Reviewer #1 (Public Review):
While the circuits underlying the computation of directional motion information in the fly brain are very well described, much less is known about the neurons serving the detection of objects. In a previous publication from the same lab, it has been shown that flies perform body saccades to track a moving object during flight. In the current paper, Frighetto and Frye provide evidence that T3 cells, a population of neurons within the optic lobes, are involved in this task. First, they performed 2-photon Calcium imaging from T3 cells to show that these cells respond to moving bars, which they later use in behavioural experiments. They then silenced T3 cells using genetic tools and tested the behavior of these flies in response to a rotating bar using two different setups. In one, the flies are fixed and bilateral changes in wing stroke amplitude are used as a measure for turning, in the other, flies are magnetically tethered such that they can rotate around the vertical body axis. Silencing T3 cells leads to the abolishment of the steering response induced by object position using a bar that is defined by its motion relative to the surround, but leaves the response to object motion intact. In the magnetically tethered flies, it reduces the number of saccades and thus leads to an impairment of bar-tracking behavior. In another set of experiments they optogenetically activated the whole population of T3 neurons (which supposedly impairs their normal function), which leads to an increase in the number of saccades after the activation (when the light stimulus used to activate the cells is turned off). Silencing the neurons necessary for detection of local motion, T4 and T5 cells, in contrast reduces responses elicited by object motion rather than position, but also has an impact on object tracking saccades. The authors provide a simple model, where speed-dependent signals from multiple T3 cells are integrated and trigger a saccade, when a threshold is reached.
The data generally support the conclusion that T3 cells play a role in detecting bar position and in controlling saccades in response to rotating bars. However, there are some inconsistencies in the data that are not sufficiently explored and discussed.
1. In a previous paper from the lab (Keleş et al., 2020), it was shown that T3 cells respond preferentially to small objects, whereas here they robustly respond to elongated bars and even large-field gratings. This discrepancy is not discussed.
2. In a previous paper, the authors showed that integrated positional error rather than bar position is used to elicit bar-tracking saccades and that saccade amplitude is relatively stereotyped. However, here they show, that T3 cells respond much more strongly to a slowly moving stimulus (18{degree sign}/s) rather than to the fast moving stimuli used for the behavioral experiments (> 90{degree sign}/s). This response property plays an important role for the model they propose. My general concern here is that the findings might not be generalizable to slower moving bars, where more precise, position-dependent responses could play a larger role, and that these fast moving bar stimuli represent an extreme situation, where the flies cannot accurately track bar position any more.
3. The claim that T3 cells are tuned to stimulus velocity is not supported by the data in my view. For the bar stimuli, the authors only tested speeds of 18{degree sign}/s and above 90{degree sign}/s, but nothing in between. For the grating motion there seems to be an influence of temporal frequency for the same stimulus velocity (see e.g. Fig.1_1), but this is not quantified.
4. The results from the optogenetic activation experiments are hard to interpret, as it is unclear how a prolonged activation of all T3 cells would affect the downstream circuitry. It is not clear that this experiment is equivalent to a "loss-of-function perturbation" of T3 cells as the authors claim in the text.
-
Reviewer #2 (Public Review):
In their manuscript titled "Feature detecting columnar neurons mediate object tracking saccades in Drosophila", Frighetto & Frye study the effect manipulating T3 neurons has on tethered flight saccades. The authors first characterize the responses of T3 neurons to simple visual stimuli, and then manipulate T3 cells (with both Kir2.1 and CsCrimson) and study the effects on the fly's tethered flight behavior, focusing on different types of sharp turns (saccades). Finally, the authors suggest an integrate and fire model to explain how an array of T3-like neurons can produce some of the recorded behavior.
The authors study the elementary, yet challenging, computation of object discrimination. They hone in on a cell type that most likely plays an important role in the circuit. However, the authors do not …
Reviewer #2 (Public Review):
In their manuscript titled "Feature detecting columnar neurons mediate object tracking saccades in Drosophila", Frighetto & Frye study the effect manipulating T3 neurons has on tethered flight saccades. The authors first characterize the responses of T3 neurons to simple visual stimuli, and then manipulate T3 cells (with both Kir2.1 and CsCrimson) and study the effects on the fly's tethered flight behavior, focusing on different types of sharp turns (saccades). Finally, the authors suggest an integrate and fire model to explain how an array of T3-like neurons can produce some of the recorded behavior.
The authors study the elementary, yet challenging, computation of object discrimination. They hone in on a cell type that most likely plays an important role in the circuit. However, the authors do not sufficiently clarify the framework in which they conceptualize T3's role in object discrimination, neither when discussing it in the introduction/discussion nor when explaining experimental results. The authors present the work in comparison to T4/T5 cells. However, T4/T5 cells have been shown to be both local motion detectors and the main cell types to compute motion in the fly's eye. Downstream neurons integrate over these local units to detect different patterns of global and local motion (Authors should cite Krapp 1996 Nature). Are the authors suggesting that T3 neurons perform a similar function only as local object detectors? That is a bold claim that will need to be supported with more experimental results and reconciled with previous results. We already know of other Lobula Columnar neurons (LCs) that respond to different sizes, some even smaller than the optimal T3 stimulus (e.g. Klapoetke 2022 Neuron) and we know of LCs that respond to small objects that do not receive major inputs from T3 cells (e.g. Hindmarsh 2021 Nature).
These differences between T4/T5 cells and T3s also make interpreting the experimental manipulations more challenging. When hyperpolarizing T4/T5 or 'blinding' them with CsCrimson activation, the visual motion circuit is severely disrupted. However, the same cannot be said about inactivating/blinding T3 neurons and the object detection circuit (if it is indeed a single circuit). The authors are justified in deducing a connection between blocking T3 neurons and a reduction in bar tracking, but generalizing the results to object detection requires more experiments and clarifications.
When framing the manuscript in the object detection framework, previous results regarding the definition of an object should also be addressed. Maimon Curr. Biol. 2008 and work from their own lab (Mongeau, 2019) have already shown that tethered flies respond differently to bars and small objects (fixating on the former while anti-fixating on the latter). Previous work has also shown that T3 neurons respond strongly to small objects and suppress responses to long bars (Tanaka Curr. Biol. 2020). Since all the behavioral experiments in the current manuscript and all the visual stimuli are full arena-length bars, it is impossible to tell whether the T3 results generalize to small objects and even how to reconcile the stronger response to small objects with the role ascribed to T3 cells in generating behavioral responses to long bars.
Finally, the authors propose a model for a hypothetical neuron downstream of T3 that would integrate over several T3s and generate saccades. However, given the current knowledge level in the fly vision field, the model should either be grounded more in actual circuit connectivity or produce testable predictions that would guide further research.
The authors should decide whether they would like to address these concerns with more specific experiments that would shed light on the role T3 has to play under different conditions and different definitions of a visual object, or whether they would prefer to limit the scope of their claims.
-
Reviewer #3 (Public Review):
In free flight, flies largely change their course direction through rapid body turns termed saccades. Given how important these turns are in determining their overall behavior and navigation, it is important to understand the neural circuits that drive the timing of triggering these saccades, as well as their amplitude. In this paper the authors leverage the powerful genetic tools available in the fruit fly, Drosophila, to address this question by performing physiology experiments as well as behavioral experiments with inactivation and activation perturbations.
The authors make three primary conclusions based on their experiments: (1) the feature detecting visual pathway (T3) is responsible for triggering saccades in response to moving objects, but not widefield motion, (2) the pathway primarily responsible …
Reviewer #3 (Public Review):
In free flight, flies largely change their course direction through rapid body turns termed saccades. Given how important these turns are in determining their overall behavior and navigation, it is important to understand the neural circuits that drive the timing of triggering these saccades, as well as their amplitude. In this paper the authors leverage the powerful genetic tools available in the fruit fly, Drosophila, to address this question by performing physiology experiments as well as behavioral experiments with inactivation and activation perturbations.
The authors make three primary conclusions based on their experiments: (1) the feature detecting visual pathway (T3) is responsible for triggering saccades in response to moving objects, but not widefield motion, (2) the pathway primarily responsible for wide field motion encoding (T4/T5) is responsible for triggering saccades in response to widefield motion, and (3) the T4/T5 pathways is responsible for controlling the amplitude of both object and widefield motion triggered saccades.
The authors go on to show that using calcium imaging data of T3 activity it is possible to predict under what conditions flies will initiate a saccade when presented with objects moving at different speeds, resulting in a parsimonious model for how saccades are triggered.
Together, the imaging, behavior, and modeling provide compelling evidence for claims 1 and 2, however, the evidence and modeling for point 3 - the amplitude of the saccades - is lacking. The statistical analysis does not go into sufficient detail in comparing across different cases, and in particular, there is little mention of the effect sizes, which appear to be quite small (this is primarily in reference to 3F and 4E). The data suggest that both the T3 and T4/T5 pathways contribute to saccade amplitude, instead of T4/T5 being the only or primary drivers.
-
-