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PONE-D-21-09940

Collective action or individual choice: Spontaneity and individuality contribute to decision-making in Drosophila

PLOS ONE

Dear Dr. Gorostiza,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

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PLOS ONE

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Partly

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2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: I Don't Know

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3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

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4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: No

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5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: In this manuscript, Steymans, et al. examined the consistency of fly to choose to move into a lighted chamber in two different, but related, phototaxis experimental designs. Wild type Drosophila melanogaster are strongly positively phototactic. The authors reexamined the issue of consistency in choice of moving to a lighted chamber by resampling the choices of flies after they have moved into a lighted chamber or remained in a darkened one. The question is interesting as it directly relates to the interpretations of a fly’s behavior in a T-Maze or counter current apparatus, and it sheds light on why some behaviors are found to be more consistent when assayed in groups of flies rather than individually.

Previous work by Seymore Benzer found that the distribution of flies in a phototaxis experiment appeared to be probabilistic; if 70% of flies chose the lighted chamber, a resampling of the flies would show the same distribution regardless of whether the initially choose the lighted chamber or the darkened chamber. However, the conservation of this distribution was not found by work reported in Kain et al, 2012, in which individual flies were repeatedly scored for movement towards light in a very different apparatus. In Kain, et al., the choice to move towards the light varied between individuals.

In Steymans, et al., the authors initial findings from resampling in the T-maze and counter-current apparatus more closely resembled that of Benzer et al. However, after increasing the number of behavior choices in the counter-current apparatus, the authors found that the previous choice strongly correlated with subsequent choices, similar to Kain et al’s finding. The authors offer the hypothesis that this emergence of consistency in choice that emerged in this retesting and in Kain’s et al. experiments is due in part to the inability to assess individual choices in group behavior in the initial counter-current experiments. The difficulty is assessing the consistency (referred to be the authors as Necessity or the need to behavior in a certain way), is also complicated by the presence of spontaneity in an animal’s behavioral choices. These are reasonable inferences.

Major Comments:

When the fly is in the counter-current apparatus or the T-maze, the flies are normally free to move back and forth. The selection they “make” is dictated by their position when the trial ends (30 seconds for T-maze and 15 seconds for the counter-current Apparatus). These are relatively short periods of time for the animals to explore and make a stable choice. The authors should discuss the choice of these time points and how they may impact the measurements of individual choice in phototactic preference.

The Buridan’s paradigm controls are an important part of this paper as they provide a mechanistic understanding for why some flies show lower phototactic behavior in the counter-current apparatus. In these experiments, it would be good if the authors reported Principle Component Analysis, to see how differences in phototaxis load with the same activity measures.

Minor Comments:

In the abstract, the authors refer to flies with an “extremely negative first choice”. Since the choices are binary, I do not understand how a negative choice cab be extremely, moderately or otherwise.

The first sentence of the Introduction mentions a Hallmark of success neuroscience is the ability to predict behavior. Although I understand this helps frame the author’s question, I know of few neuroscientists that would agree with this criterion. Perhaps this can be rephrased so as not to appear to be a straw man.

Reviewer #2: The paper nicely addresses two sets of seemingly canonical and conceptually most influential experiments on the consistency/ variability of behavior that are talked about a lot, but hardly ever substantiated by actual data.

I refrain from reporting in my own words what the authors did. I just report my main concerns to be i) the Methods are not explained with sufficient clarity; to some extent, good info graphics may help; ii) the possible contribution of learning processes is not properly considered.

The “or” in the title implies a dichotomy of action vs choice that the authors may not actually want to be implied. Please reconsider the title.

The Introduction features a number of (words for) behavioural paradigms that are not properly defined or explained. Such as “phototaxis, the paradigm mentioned in the Benzer-quote, the one used by Quinn et al, light-dark T-maze, classic countercurrent phototaxis experiment, Benzer phototaxis experiments”.

Please clearly define these terms and do not use synonyms, and apply the defined terms consistently throughout the manuscript.

Were animals from both sexes used? If not, why not? and: which sex was used? If both sexes were used: could you separate the data by sex?

Not all terms of the CI equation are defined, and FE is defined in the text but not used in the equation. Please check.

Line 172: Better something like “We added the flies from the elevator to the ones in the dark for retesting…?

Line 174: I do not quite understand what you did (and hence the rational) of the procedure described as “Second, …”.

Line 223, after “Therefore…”: I do not get exactly what was done here. Partially, this might be because of uncertainty as to what is the meaning of “repetition, session, choice, experiment, replicate, test” in the preceding and following sections.

Line 289: Do you imply the flies indeed jumped, or do you consider these to be cases of artefact from tracking?

In general, carefully designed info-graphics for the behavioural paradigms would seem very useful.

The Benzer paradigm is explained much better in the Results than in the Methods section…

Line 237: Please explain why what appears to be a very conservative P-threshold was used (P< 0.005(!)).

Line 373: the wording “… we tested whether a previous light/dark choice would influence a subsequent choice” implies some sort of learning, and indeed it would seem important to consider such learning throughout the manuscript as a possibility (also @Introduction). One could view such learning as a “personalizing” of chance/ of action probability fields, really. This might be what happens when, as the authors put it in a subheading “Fly individuality appears with more choices”.

In other words, the flies may behave according to a given probability of choosing dark, and that probability can be changed by the experience the flies make based on this very choice (or multiple such choices). This would call into question the dichotomy the authors put up: for a given fly the behavior is according to its own, self-tailored and adjustable probability. In yet other words, each fly throws a dice as to what to do and does so each time anew, but it throws “its own” dice, one that it has shaped itself before and one that changes at least slightly every time actions were based on what the dice returned.

Interestingly, in the work of Pamir et al 2014 Front Behav Neurosci on bees, the animals are suggested to learn in an all-or-none manner, so to speak switching from one rigidity (not-showing PE towards an odour) to another rigidity (showing PE).

I wonder whether canonical learning mutants would be less well “selectable” than the ones in Figure 2…?

Line 439: I cannot quite certainly follow what was done here; this means I did not get the difference/ relationship between Figures 2C versus D. This would seem essential to judge a key conclusion, though.

If I understood correctly, it might help to label the X-axis with “Number of times choosing light” or so (I hope I got the sign right here…). Using the abstract expressions “Tube #1, …” would be intuitive only with the suitable graphic/ pictogram.

Maybe arrange Figure 3 such that the significantly different data are grouped together?

Maybe the data from figure 3 would allow some sort of machine-learning approach to see how many “kinds of flies” there are?

Line 569: It should be “than” and not “then” here.

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Reviewer #1: No

Reviewer #2: No

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Collective action or individual choice: Spontaneity and individuality contribute to decision-making in Drosophila

PONE-D-21-09940R1

Dear Dr. Gorostiza,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

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Kind regards,

Efthimios M. C. Skoulakis, PhD

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

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2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

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3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

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5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: I am satisfied with the authors edits and additions. Congratulations on your interesting results.

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7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No