Metabolic rate in common shrews is unaffected by temperature, leading to lower energetic costs through seasonal size reduction

Small endothermic mammals have high metabolisms, particularly at cold temperatures. In the light of this, some species have evolved a seemingly illogical strategy: they reduce the size of the brain and several organs to become even smaller in winter. To test how this morphological strategy affects energy consumption across seasonally shifting ambient temperatures, we measured oxygen consumption and behaviour in the three seasonal phenotypes of the common shrew (Sorex araneus), which differ in size by about 20%. Body mass was the main driver of oxygen consumption, not the reduction of metabolically expensive brain mass. Against our expectations, we found no change in relative oxygen consumption with low ambient temperature. Thus, smaller body size in winter resulted in significant absolute energy savings. This finding could only partly be explained by an increase of lower cost behaviours in the activity budgets. Our findings highlight that these shrews manage to avoid one of the most fundamental and intuitive rules of ecology, allowing them to subsist with lower resource availability and successfully survive the harsh conditions of winter.


INTRODUCTION
I have a little confusion with the expression of the aim of the study. If my meaning of Authors' intention is correct, scientific reasons for which the work was done are presented on page 5, lines 91-92 and lines 108-109. But now, they are hidden in the main text. Both arguments should be strongly emphasized together with a general description of methods used for such comparisons. It ones read then oxygen consumption was measured outside at ambient environmental temperatures (lines 111-112), but no word about a way in which a class of behavior was recording. Please, re-write lines 110-114 to make that paragraph (very essential for Introduction) more informative and scientific important.
Pages 5-6, lines 114-120: Re-think and make hypotheses more clear. In Discussion there is the statement 'we expected both body mass and environmental temperature to influence organismal metabolic rate' (page 12, lines 286-287). I can't see any detailed speculation about an effect of body mass reduction in shrews on their metabolic rate in that part of Introduction.
Page 4, line 84: Shouldn't be 'surface to volume ratio'? METHODS Page 6, line 140: Put information how long shrews have been waiting for the beginning of the experiments. The time course between capturing and experiments was the same for individuals from each season or not? If not, way?
Page 6, lines 141-144: I have mixed feelings with that information. It describes the method for incorporation digested food into metabolism and assessment of turnover rate of body fat, which was one of the aim in previous Authors' study (I suppose it is Keicher L., O'Mara M.T., Voigt C.C., Dechmann D.K. 2017. Stable carbon isotopes in breath reveal fast incorporation rates and seasonally variable but rapid fat turnover in the common shrew (Sorex araneus). J. Exp. Biol. 220, 2834-2841. Moreover, it looks like the same group of shrews was used in the previous paper and this work. And of course, a way like this is acceptable in results sharing and publishing, but I can see no scientific need for putting information like that here. Results on incorporation rates and fat turnover were published and they are not presented in actually reviewed paper. I suggest removing that information.
Page 6, line 144: Phrase 'two and seven days after capture' is confusing. It is not clear to which step of experiment it relates to. And again, does it describe the period of time before the start of oxygen consumption analyses? Please, specify it.
Page 7, line 147: Remove the phrase 'for 12 hours'. It is mentioned below, in line 164 (and refers to all shrews used in measurements of oxygen consumption).
Page 7, line 157: Please decide which form 'braincase' or 'brain case' is correct and apply it throughout the paper.  Figure 3E' instead of ' Figure 3D'. OTHER COMMENTS: Figure 1 and 2. The sample size for winter subadults and spring adults on both figures is the same (n=8 and n=6, respectively). Explain the difference in the number of summer juveniles presented on Figure 1 and 2 of (in Methods, consistently).
Decision letter (RSOS-191989.R0) 10-Mar-2020 Dear Dr Schaeffer On behalf of the Editors, I am pleased to inform you that your Manuscript RSOS-191989 entitled "Metabolic rate in common shrews is unaffected by seasonal temperature, leading to reduction of energetic costs through size reduction." has been accepted for publication in Royal Society Open Science subject to minor revision in accordance with the referee suggestions. Please find the referees' comments at the end of this email.
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Once again, thank you for submitting your manuscript to Royal Society Open Science and I look forward to receiving your revision. If you have any questions at all, please do not hesitate to get in touch. Two expert reviews and I reviewed the manuscript "Metabolic rate in common shrews is unaffected by seasonal temperature, leading to reduction of energetic costs through size reduction." A common ecological rule is that animals are larger in colder environments. This study investigated the energetic and behavior consequences of the paradoxical seasonal reductions in body size in the common shrew. The study shows that the combination of seasonal changes in behavior and morphology results in energy savings. Both reviewers felt that the study was scientifically sound and contributed to our understanding of thermal strategies and adaptations. The reviewers liked the integration of physiology and behavior and felt that it was represented a novel contribution to the literature on thermal strategies. I agree with the reviewers that parts of the introduction could be rearranged and edited to enhance the paper. I recommend making the hypothesis/aims of the study more salient and clarifying the description of the metabolic theory of ecology so that it more accurately reflects the literature on that topic. One reviewer's comments focused on how conductance and body temperature differences might relate to the presented results; please integrate these concepts to enhance your discussion. Address the comments about the methods to increase the ability for replication of the study.
Reviewer comments to Author: Reviewer: 1 Comments to the Author(s) This paper investigates changes in metabolic rate in response to seasonal changes in temperature and phenotypic changes in body size and relative brain size to further elucidate the mechanisms behind Dehnel's Phenomenon. The methods and data are sufficiently executed. The interpretation of the results are adequate.
Abstract: Generally good. I would suggest including a brief explanation for how the authors ruled out the following in the abstract "Body mass was the main driver of oxygen consumption, not the reduction of metabolically expensive brains". For example, through the use of generalized mixed effects models. This is also unclear in the objectives paragraph 110-120.

8
Introduction: The authors state that MTE provides an important framework to tease apart the effect of body size and temperature as the main energetic costs for endotherms. This isn't exactly true. The temperature term in MTE (Brown et al. 2004) is included to accommodate ectotherms that vary in body temperature in response to ambient temperatures. The major extension of MTE and Scholander-Irving to accommodate impacts of temperature in endotherms to body size and metabolic rate comes from Fristoe et al. (2015 PNAS). That and Scholander showed how changes in environmental temperature influences changes in body temperature via metabolic rate and conductance. It seems the alternative hypothesis the authors are testing is that the mass-specific metabolic rate would be greater for body size in winter to offset heat lost to cold, although the authors do not find support for this. This may inform predictions in changes in body temperature, which the authors do not report here (and that's OK). But may wish to include in future studies. But I do think this section of the introduction should be revised to clearly explain the link between MTE and predictions.
Line 45: "It states that temperature" do the authors mean environmental or body temperature or both? See above explicit distinction in theory. This should be clear throughout when temperature is used, for example, in lines 58-59 is it body or environmental temp?
Line 85: Are the author's referring to metabolic scope?
Line 89: If S. araneus at 10 g has the same BMR as 25 g vole and two that of a similar sized whitetoothed shrew, the does it have a higher body temperature, higher conductance or operate and colder environmental temperatures (e.g., Fristoe et al. 2015)? The authors do not report body temperature or conductance, but these links need to be clear in the context of the theory.
Line 113: again, you must be referring to environmental temperature unless you measured body temperature too?
Line 232: Please reference previously published studies you refer to.
Line 239: Please cite the previously published data source.
Paragraph 312-333: is very long. Consider breaking in to two.

Reviewer: 2
Comments to the Author(s) General: I found the study as an interesting and thought-provoking. Authors continuously produce papers discussing morphological and physiological adaptations to seasonal changes of ambient temperatures in small, non-hibernating mammals (e.g. Dechmann et al. 2017, LaPoint et al. 2017, Lázaro et al. 2017and 2018. Here, along with changes in body mass and skull size in common shrews across three seasons (summer, winter and spring), Authors estimated individual's energy consumption (absolute and mass-specific) together with five classes of behavior activity (rest, eat, drink, run and walk). And these two last measurements, metabolic rate and behavior, add new facts to the number of papers testing animals adaptations to challenging environmental factors. The demonstration of seasonal phenotypic plasticity in morphological, behavioral and metabolic traits is fundamental not only for better understanding one of winter strategy in small mammals called Dehnel's phenomenon, but also for the whole evolutionary physiology. From this perspective the study is important. However, I have several comments, which can help Authors make some corrections and improve the paper.

INTRODUCTION
I have a little confusion with the expression of the aim of the study. If my meaning of Authors' intention is correct, scientific reasons for which the work was done are presented on page 5, lines 91-92 and lines 108-109. But now, they are hidden in the main text. Both arguments should be strongly emphasized together with a general description of methods used for such comparisons. It ones read then oxygen consumption was measured outside at ambient environmental temperatures (lines 111-112), but no word about a way in which a class of behavior was recording. Please, re-write lines 110-114 to make that paragraph (very essential for Introduction) more informative and scientific important.
Pages 5-6, lines 114-120: Re-think and make hypotheses more clear. In Discussion there is the statement 'we expected both body mass and environmental temperature to influence organismal metabolic rate' (page 12, lines 286-287). I can't see any detailed speculation about an effect of body mass reduction in shrews on their metabolic rate in that part of Introduction.
Page 4, line 84: Shouldn't be 'surface to volume ratio'? METHODS Page 6, line 140: Put information how long shrews have been waiting for the beginning of the experiments. The time course between capturing and experiments was the same for individuals from each season or not? If not, way?
Page 6, lines 141-144: I have mixed feelings with that information. It describes the method for incorporation digested food into metabolism and assessment of turnover rate of body fat, which was one of the aim in previous Authors' study (I suppose it is Keicher L., O'Mara M.T., Voigt C.C., Dechmann D.K. 2017. Stable carbon isotopes in breath reveal fast incorporation rates and seasonally variable but rapid fat turnover in the common shrew (Sorex araneus). J. Exp. Biol. 220, 2834-2841). Moreover, it looks like the same group of shrews was used in the previous paper and this work. And of course, a way like this is acceptable in results sharing and publishing, but I can see no scientific need for putting information like that here. Results on incorporation rates and fat turnover were published and they are not presented in actually reviewed paper. I suggest removing that information.
Page 6, line 144: Phrase 'two and seven days after capture' is confusing. It is not clear to which step of experiment it relates to. And again, does it describe the period of time before the start of oxygen consumption analyses? Please, specify it.
Page 7, line 147: Remove the phrase 'for 12 hours'. It is mentioned below, in line 164 (and refers to all shrews used in measurements of oxygen consumption).
Page 13, line 324: Put ' Figure 3E' instead of ' Figure 3D'. OTHER COMMENTS: Figure 1 and 2. The sample size for winter subadults and spring adults on both figures is the same (n=8 and n=6, respectively). Explain the difference in the number of summer juveniles presented on Figure 1 and 2 of (in Methods, consistently).

31-Mar-2020
Dear Dr Schaeffer, It is a pleasure to accept your manuscript entitled "Metabolic rate in common shrews is unaffected by temperature, leading to lower energetic costs through seasonal size reduction." in its current form for publication in Royal Society Open Science. The comments of the reviewer(s) who reviewed your manuscript are included at the foot of this letter.
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Dear Dr. Downs and Dr. Dunn We are, of course, excited about your conditional acceptance of our paper in Royal Society Open Science. We have gone through all the instructions in your email carefully, and added the required information, particularly also the end sections (new line numbers 344ff). We also thank you and the reviewers for the very welcome comments. Please find our responses below in bold as well as in the revised manuscript (as suggested we submit a version with and without tracked changes, the indicated line numbers refer to the tracked version). Please note that we have slightly changed the title as the previous one seemed a bit bumbling to us now. We hope we have been able to clarify and address everything to your satisfaction and look forward to hearing from you.

Paul Schaeffer and co-authors
Associate Editor Comments to Author (Dr Cynthia Downs): Two expert reviews and I reviewed the manuscript "Metabolic rate in common shrews is unaffected by seasonal temperature, leading to reduction of energetic costs through size reduction." A common ecological rule is that animals are larger in colder environments. This study investigated the energetic and behavior consequences of the paradoxical seasonal reductions in body size in the common shrew. The study shows that the combination of seasonal changes in behavior and morphology results in energy savings. Both reviewers felt that the study was scientifically sound and contributed to our understanding of thermal strategies and adaptations. The reviewers liked the integration of physiology and behavior and felt that it was represented a novel contribution to the literature on thermal strategies. >> Thank you for your positive review and the comments which we respond to in detail below I agree with the reviewers that parts of the introduction could be rearranged and edited to enhance the paper. I recommend making the hypothesis/aims of the study more salient, and clarifying the description of the metabolic theory of ecology so that it more accurately reflects the literature on that topic. >> We have tried to address these points as described in detail below. We have rewritten sections of the introduction to reflect reviewer comments, particularly as related to conductance and the specific hypotheses.
One reviewer's comments focused on how conductance and body temperature differences might relate to the presented results; please integrate these concepts to enhance your discussion.

>> We have added a short comment on this point (new line numbers 292ff).
Address the comments about the methods to increase the ability for replication of the study. >> We have addressed the comments in the methods to better explain how the animals were handled and give our rationale for why this should be included. We hope this addresses this concern (various line numbers, see responses to reviewers below).
Reviewer comments to Author: Reviewer: 1 Comments to the Author(s) This paper investigates changes in metabolic rate in response to seasonal changes in temperature and phenotypic changes in body size and relative brain size to further elucidate the mechanisms behind Dehnel's Phenomenon. The methods and data are sufficiently executed. The interpretation of the results is adequate.
Abstract: Generally good. I would suggest including a brief explanation for how the authors ruled out the following in the abstract "Body mass was the main driver of oxygen consumption, not the reduction of metabolically expensive brains". For example, through the use of generalized mixed effects models. This is also unclear in the objectives paragraph 110-120. >>There may be different schools with regards to this, but as the methods used are not what is the important message of the paper we have excluded the explanation of the statistical methods used to support the conclusion in both of these sections. These are clarified in the methods and results sections.
Introduction: The authors state that MTE provides an important framework to tease apart the effect of body size and temperature as the main energetic costs for endotherms. This isn't exactly true. The temperature term in MTE (Brown et al. 2004) is included to accommodate ectotherms that vary in body temperature in response to ambient temperatures. The major extension of MTE and Scholander-Irving to accommodate impacts of temperature in endotherms to body size and metabolic rate comes from Fristoe et al. (2015 PNAS). That and Scholander showed how changes in environmental temperature influences changes in body temperature via metabolic rate and conductance. It seems the alternative hypothesis the authors are testing is that the mass-specific metabolic rate would be greater for body size in winter to offset heat lost to cold, although the authors do not find support for this. This may inform predictions in changes in body temperature, which the authors do not report here (and that's OK). But may wish to include in future INTRODUCTION I have a little confusion with the expression of the aim of the study. If my meaning of Authors' intention is correct, scientific reasons for which the work was done are presented on page 5, lines 91-92 and lines 108-109. But now, they are hidden in the main text. >> Yes, these set the stage for the questions asked in the final paragraph. We have clarified those questions to better follow from these points (new lines 113ff).
Both arguments should be strongly emphasized together with a general description of methods used for such comparisons. It ones read then oxygen consumption was measured outside at ambient environmental temperatures