The manuscript JMB4-08-20-0095 by Bohns F. et al. describes a deep investigation of bone mineralization following alendronate treatment in an osteoporotic induced model of zebrafish. Tail regrown was choose as a model for mineralization investigation, glucocorticoid treatment was performed to obtain the osteoporotic model and the bisphosphonate Alendronate as example of anti-resorptive drug..
Although the effect of both used drugs is not a novelty the type of analyses are interesting and can be of relevance for the bone field especially in drug screening investigations.
The data support the conclusion, and the manuscript is clearly written.
Comments
1. A relevant limit of the study is the small number of animals per group, especially considering the variability of zebrafish. In Raman study, only one dish was used! This limit should be clearly stated and the standard length of the animals used in the various experiments should be indicted.
2. In the study, the authors evaluate the tail regrowth; it will be interesting also to have the data from not cut fish tail or a reference for that should be provided if available in literature. As pointed out ALN is impairing osteoclasts, but there is no mention of the location and role of osteoclasts in tail regrowth. The discussion is extremely long and should be shortened; nevertheless, this point should be addressed.
3. It will be nice to have data also for a CTRL group treated with ALN; this is missing from the study.
4. Please clarify better the Alizarin Red S staining time points, it is not clear if it has been done only once at the beginning of the study or multiple times.
5. It is not clear why cross section slides were used for surface profile and mechanical testing. It should be explained in the test.
6. It will be nice to have a schematic representation of the cut performed for the analysis of mineral morphology and content.

All the points raised on the first submission were properly addressed.