An open-source platform for head-fixed operant and consummatory behavior

  1. Adam Gordon-Fennell
  2. Joumana M Barbakh
  3. MacKenzie T Utley
  4. Shreya Singh
  5. Paula Bazzino
  6. Raajaram Gowrishankar
  7. Michael R Bruchas
  8. Mitchell F Roitman
  9. Garret D Stuber

  • Curated by eLife

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    Gordon-Fennell et al. present a low-cost, open-source platform for measuring action elicitation and consummatory behavior in head-fixed animals. The findings are important because they allow animals to perform a truly voluntary action whilst their head is held still, and the evidence supporting them is both comprehensive and compelling (in some cases even exceptional). The results have the potential to have a broad impact in the field as many labs start to move towards measuring head-fixed behavior effectively, although this is said with the caveat that such behavior will never be an ideal replication of naturalistic behavior.

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Abstract

Head-fixed behavioral experiments in rodents permit unparalleled experimental control, precise measurement of behavior, and concurrent modulation and measurement of neural activity. Here, we present OHRBETS (Open-Source Head-fixed Rodent Behavioral Experimental Training System; pronounced ‘Orbitz’), a low-cost, open-source platform of hardware and software to flexibly pursue the neural basis of a variety of motivated behaviors. Head-fixed mice tested with OHRBETS displayed operant conditioning for caloric reward that replicates core behavioral phenotypes observed during freely moving conditions. OHRBETS also permits optogenetic intracranial self-stimulation under positive or negative operant conditioning procedures and real-time place preference behavior, like that observed in freely moving assays. In a multi-spout brief-access consumption task, mice displayed licking as a function of concentration of sucrose, quinine, and sodium chloride, with licking modulated by homeostatic or circadian influences. Finally, to highlight the functionality of OHRBETS, we measured mesolimbic dopamine signals during the multi-spout brief-access task that display strong correlations with relative solution value and magnitude of consumption. All designs, programs, and instructions are provided freely online. This customizable platform enables replicable operant and consummatory behaviors and can be incorporated with methods to perturb and record neural dynamics in vivo.

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  1. Version published to 10.7554/elife.86183 on eLife
  2. eLife

    Author Response

    Reviewer #1 (Public Review):

    Weaknesses

    1. I was curious as to how novel this setup is. Although I do not do head-fixed research myself, I thought there were already some open-source, relatively cheap systems available. I'm not sure how the current setup differs from those already available. Personally, even if this system involves only the wheel turning, as this is a truly operant response, that is novel enough for my liking.

    The novelty of the system stems from the synergistic combination of functionality, the low-cost open source nature of the design, and the breadth of behavioral procedures the system is able to support. The use of a wheel as an operant response was adapted from the International Brain Laboratory rig which has been used extensively for visual discrimination tasks. We adapted this wheel design to make the response closer to lever pressing through the use of the wheel brake, which ensures that subjects have to rotate the wheel in discrete rotational bouts rather than continuously spinning the wheel and potentially disengaging and allowing the wheel to rotate independently. There are no examples of systems capable of delivering 5+ solutions within a behavioral session or conducting valence testing with a modification of real-time place preference without the cost and complexity associated with virtual reality. We believe that the combination of factors, the flexibility and scalability of the system makes OHRBETS a novel and useful system for diverse motivation and consumption behaviors in head-fixed mice.

    1. It would be useful to have a bit more detail in the manuscript (not just on the GitHub link - in supplemental material perhaps?) on how to build such a system, just to get a sense of how difficult building such a system might be and how many components it has.

    With this submission we have included detailed assembly instructions as a supplement to the main manuscript and added reference to the file within the methods section. We have also added details, including time estimates, to the methods section.

    1. I wasn't sure how to feel about the comparisons across experimental set-ups in Figures 2 and 3. Usually, these sorts of comparisons are not considered statistically valid due to the many variables that differ between set-ups. However, I do see that the intent here is a bit different - i.e. is to show that despite all these alterations in variables the behavioural outputs are still highly correlated. However, without commenting on this intent, I did find these comparisons a little jarring to read.

    Thank you for highlighting this. We have added in a justification for why we measured the consistency in behavior measured with each head-fixed system.

    1. The only dataset I was not wholly convinced by was that in Figure 3 (real-time place preference and aversion). I think the authors have done the best job that they can of replicating such a procedure in a head-fixed mouse, but the head-fixed version is going to necessarily differ from the freely moving version in a fundamental way when the contextual cues and spatial navigation form part of the RTPT task. Giving a discrete cue, such as a tone, just is not a sufficient substitute for contextual cues, and I think the two types of task would engage fundamentally different brain cells and circuits (e.g. only the free-moving version is likely to engage place cells in the hippocampus).

    To avoid confusion regarding the place component of the real-time place preference assay name, we have renamed the head-fixed assay for assessing valence to Wheel-Time Preference (WTP). We have also added a full paragraph to the discussion where we outline the differences in the task requirements and relevant neuronal circuits between the freely-moving RTPP and head-fixed WTP. We understand that the head-fixed task is not a perfect analog of the RTPP task, however based on the similarity in the resulting time spent in the stimulation chamber/zone we believe that the WTP is able to replicate the valence assessment that many in the field uses RTPP to measure. We believe that the WTP with OHRBETS opens up new possibilities for assessing preference in head-fixed mice and this justifies keeping the figure within the main manuscript.

    To thoroughly address the potential confound of spatial information during the multi-spout experiment, we have added an additional supplemental figure (Figure 4- figure supplement 5) that depicts the proportion of trials with licking and added a paragraph to the discussion centered on the potential confound associated with learning the solution identity.

    1. Personally, I found having the statistics in a separate file confusing.

    Thank you for raising this concern. With our initial submission, we were concerned that including all of the statistics within the main text would make the paper difficult to read due to the extensive amount of statistics. With this submission, in addition to the statistics table, we have included statistics within the figure legends and main text where applicable.

    1. Line 589-594. Suggesting the medial/lateral shell recording results mean that the medial shell 'tracks value, and the range of values during the multi-spout consumption of gradients of NaCl is greater than the range of values during multi-spout consumption of gradients of sucrose" seems to engage in circular logic to me. That is, the authors should use behavioural data to infer what the animal is experiencing and whether it is a change in value, and/or a greater change in value during NaCl vs. sucrose consumption, and only then should they make an inference about what the larger medial shell response means.

    Thank you for identifying this potential site of confusion. To address this concern we have modified the language to better communicate our interpretation of the data.

    “If we assume that the range of values is greater during multi-spout consumption of gradients of NaCl compared to gradients of sucrose, as indicated by a greater range in licking behavior (Figure 8- Figure Supplement 4), then the greater range of dopamine release in the NacShM could imply that dopamine release in this structure tracks value.”

  3. eLife

    eLife assessment

    Gordon-Fennell et al. present a low-cost, open-source platform for measuring action elicitation and consummatory behavior in head-fixed animals. The findings are important because they allow animals to perform a truly voluntary action whilst their head is held still, and the evidence supporting them is both comprehensive and compelling (in some cases even exceptional). The results have the potential to have a broad impact in the field as many labs start to move towards measuring head-fixed behavior effectively, although this is said with the caveat that such behavior will never be an ideal replication of naturalistic behavior.

  4. eLife

    Reviewer #1 (Public Review):

    Gordon-Fennell et al., here present a relatively low-cost, open-source platform for head-fixed operant and consummatory behaviour, called OHRBETS (prounounced Orbitz). This setup provides a great advantage over other systems in that it enables the animal to perform a truly operant response (i.e.one that fulfills the criterion of bidirectionality) whilst head-fixed. The authors provide thorough evidence of the utility of this setup, showing that a number of behavioural paradigms can be performed whilst the animal is head-fixed, as well as consummatory behaviours, optogenetic manipulations, and photometry recordings. These findings will be of broad interest to neuroscientists across multiple fields.

    Strengths:
    1. The work presented here is extremely thorough and explores multiple different types of paradigms. There is a huge amount of data that will be immensely useful to individuals who hope to use this setup and build on these findings. The setup is generally well-explained.
    2. The statistics reported are generally quite strong and the sample sizes are sufficient (although strictly speaking ANOVA and Tukey should not be used together - Tukey's 'overall' test is a test of the maximal comparison, if the maximal comparison is not significant then no other pairwise comparison will be).
    3. The open-source nature of the system is a great advantage as the fact that it is relatively low cost (as long as a lab has access to a 3D printer). This and similar endeavours will promote equality throughout the field.
    4. The response here is truly operant as it is bidirectional. In other words, the animal shows that its response is governed by the relation between that response and the outcome, not stimulus-outcome associations like so many other so-called 'operant' responses (e.g. licking, food approach behaviours). Here, the stimuli are kept constant but the animal will either turn the wheel to the left or to the right to receive the food, depending on which direction is reinforced. This means that the animal cannot be governed solely by a stimulus-outcome response as in Pavlovian conditioning, because their response would not flexibly reverse the way that it is shown to here, particularly in Figure 1Q.
    5. The accumbens shell recordings are interesting data in their own right (i.e. not simply to demonstrate the viability of the system), particularly the heterogeneity of the responses in the medial and lateral shells. This could be interesting for future studies to follow up on.
    6. The correlational data between the head-fixed and free-moving versions of paradigms is, for the most part, quite convincing.

    Weaknesses
    1. I was curious as to how novel this setup is. Although I do not do head-fixed research myself, I thought there were already some open-source, relatively cheap systems available. I'm not sure how the current setup differs from those already available. Personally, even if this system involves only the wheel turning, as this is a truly operant response, that is novel enough for my liking.
    2. It would be useful to have a bit more detail in the manuscript (not just on the GitHub link - in supplemental material perhaps?) on how to build such a system, just to get a sense of how difficult building such a system might be and how many components it has.
    3. I wasn't sure how to feel about the comparisons across experimental set-ups in Figures 2 and 3. Usually, these sorts of comparisons are not considered statistically valid due to the many variables that differ between set-ups. However, I do see that the intent here is a bit different - i.e. is to show that despite all these alterations in variables the behavioural outputs are still highly correlated. However, without commenting on this intent, I did find these comparisons a little jarring to read.
    4. The only dataset I was not wholly convinced by was that in Figure 3 (real-time place preference and aversion). I think the authors have done the best job that they can of replicating such a procedure in a head-fixed mouse, but the head-fixed version is going to necessarily differ from the freely moving version in a fundamental way when the contextual cues and spatial navigation form part of the RTPT task. Giving a discrete cue, such as a tone, just is not a sufficient substitute for contextual cues, and I think the two types of task would engage fundamentally different brain cells and circuits (e.g. only the free-moving version is likely to engage place cells in the hippocampus).
    5. Personally, I found having the statistics in a separate file confusing.
    6. Line 589-594. Suggesting the medial/lateral shell recording results mean that the medial shell 'tracks value, and the range of values during the multi-spout consumption of gradients of NaCl is greater than the range of values during multi-spout consumption of gradients of sucrose" seems to engage in circular logic to me. That is, the authors should use behavioural data to infer what the animal is experiencing and whether it is a change in value, and/or a greater change in value during NaCl vs. sucrose consumption, and only then should they make an inference about what the larger medial shell response means.

    Overall this is a very solid paper in which the authors achieve their aims of demonstrating an open-source system for head-fixed operant and consummatory behavioural assessment, that is successfully employed across a number of different behavioural assays as well as in conjunction with optogenetic manipulations and fibre photometry recordings.

  5. eLife

    Reviewer #2 (Public Review):

    The manuscript by Gordon-Fennell et al. presents an open-source platform for the analysis of behavior in a head-fixed apparatus (termed OHRBETS). In addition to providing instruction on how to assemble and implement the apparatus itself, the authors validate its use across a set of procedures broadly relevant to the field of behavioral neuroscience - including operant conditioning and fluid consumption protocols run in conjunction with optical manipulation and/or recording of neural activity.

    The manuscript is comprehensive and clearly very strong. It also has the potential to have a broad impact in the field as many labs start to move towards effective head-fixed behavior. I also appreciate the fact that this manuscript includes a range of very strong behavioral tests - including experiments where several reinforcer options are available. This could be used for studies assessing taste, preference, reinforcer value, etc. Overall, the manuscript is impactful and my enthusiasm for it is high.

  6. eLife

    Reviewer #3 (Public Review):

    Head-fixed preparations should always be conceived more as a necessity (for example, to avoid damaging expensive lab equipment) than as a final path towards which the entire field of neuroscience must go. The ideal will always be to move towards a more naturalistic and ecological approach to understanding behavior. Said that. The Davis Rig seems to be a thing of the past, welcome the Open-Source Head-fixed Rodent Behavioral Experimental Training System (OHRBETS). OHRBETS represents a significant advantage over the Davis Rig equipment to measure oromotor palatability responses in a brief access test, to perform positive and negative reinforcement, and even real-time place preference in a head-fixed preparation.

    This is a well-written manuscript; the work and results are impressive. The manuscript is quite relevant to the Neuroscience field and will be of general interest. The experiments were carefully done. It is expected that OHRBETS will be widely used in multiple Neuroscience labs.

  7. Version published to 10.1101/2023.01.13.523828v1 on bioRxiv