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This review reflects comments and contributions from Marina Schernthanner, Femi Arogundade and Pablo Ranea-Robles. Review synthesized by Jonny Coates.

The study leverages the phenotype presented by the renal Glut2 KO mice (glycosuria with normal glycemia) to investigate how the body senses this glucose loss and the mechanisms behind metabolic homeostasis processes that lead to enhanced glucose production so glycemia remains stable. The use of a genetically modified mouse model with renal Glut2 knockout provides a controlled system for studying the specific role of renal glucose transporters in glucose homeostasis. The study involves various methods, including measurements of glucose production, metabolomics, gene expression related to the hypothalamic-pituitary-adrenal axis, afferent renal nerve ablation, and analysis of secreted proteins. The authors point to a kidney/hypothalamus axis and suggest the involvement of different acute phase proteins in this homeostatic response. The limitations of the study are acknowledged, and further research is suggested to delve deeper into the role of secretory proteins and the specific source of endogenous glucose production after afferent renal denervation. The manuscript is well written and the results are potentially of interest. The study's findings have potential implications for the field of diabetes treatment, as they suggest a mechanism that may explain why SGLT2 inhibitors don't achieve their full potential in lowering blood glucose levels. However, we think that some of the conclusions are merely based on descriptive assessments of changes occurring in the renal Glut2 KO mice. There are a lack of details in the reporting of some of the results and, in particular, in the discussion section, that would also require a bit more explanation from the authors. Right now, it could be hard for the reader to place this research in context. We have summarized our comments below

Major comments:

• The study mentions the use of male and female mice, but it's important to know the sample sizes for each experimental group and how gender might influence the results. Additionally, the authors should provide more details about the control groups and their matching criteria to ensure the validity of comparisons. Moreover, the exact genetic information for the knockout mice i.e. what is the CreER driver that makes it kidney-specific? Is missing. It is currently inconsistent in terms of sex and age of mice used for different experiments. 

Minor comments:

• While the metabolomics analysis is described, more information is needed about the biological significance of the changes observed in the metabolites. How do these changes relate to the compensatory glucose production, and are they causally linked?

• The paper would benefit from improved organization and clarity, particularly in the results and discussion sections. 

• Crh+ cells in control image of fig 2 are not clear. The authors could consider highlighting the are where these cells are present, or add an inset showing a zoomed image of some positive cells

• How specific is the use of capsaicin to selectively suppress afferent renal nerve activity? Does this impact other neurons? Either citations or experimental data should be included here. 

• The conditions of mice in Sup Fig. 1 are not clear and should be stated clearly in this part of the text and in the figure legend.

• The study is transparent about its limitations and raises important questions for future research. This acknowledgment of limitations contributes to the scientific rigor of the work.

• While control groups are mentioned, it's not clear how these controls were chosen or matched to the experimental group. Further information is needed on how these controls were used to make valid comparisons.

• While the study describes the experimental procedures in detail, it's essential to provide information on how many times these experiments were replicated to assess the reproducibility of the results. This is especially crucial given the complex methods used.

• Blocking the HPA axis and assessing responses in KO and WT mice would strengthen the data in Fig 2

• Investigating or showing the levels of glucagon and adrenaline to delineate mechanisms of tissue-specific glucose production would further strengthen the data presented. 

• Is it possible to measure glucose production under denervation conditions? That would support the conclusion if the increased glucose production is blunted 

• Not everyone might be familiar with the abbreviation 2D-DIGE. Explaining this before first use would be beneficial. 

• Supp fig 2 could be fused with Fig 4 to make the argument more convincing.

• The authors state that “It is possible that afferent renal denervation in the present study attenuated only hepatic glucose production through the hypothalamus without affecting the compensatory increase in renal (local) glucose production”. Addressing this would significantly strengthen the manuscript, particularly given that the title includes “hypothalamus-kidney axis”. 

Comments on reporting:

• The paper mentions the use of statistical tests but lacks information on the specific statistical tests performed for each analysis. It's crucial to provide details on the tests used, assumptions made, and how p-values were adjusted for multiple comparisons, if applicable.

Suggestions for future studies:

• Extend the research to human subjects, particularly individuals with diabetes treated with SGLT2 inhibitors. Investigate whether similar mechanisms and pathways are at play in humans, and whether these findings have clinical relevance.

• Investigate the specific roles of secreted proteins, such as acute phase proteins and major urinary proteins, in glucose regulation and potential interactions with the kidney-hypothalamus axis.

• Explore how the kidney-hypothalamus axis integrates with other nervous system and endocrine signals involved in glucose regulation, such as insulin and glucagon.

• Conduct in-depth studies on the impact of afferent renal nerve activity on glucose homeostasis and the signaling pathways involved. Investigate the role of sensory nerves in detecting glycosuria and triggering compensatory responses.

Competing interests

The author declares that they have no competing interests.