This research manuscript discusses the impact of gut inflammation on 6-OHDA-induced DA neurodegeneration in the development of Parkinson's Disease. The timing of this research is crucial as most epidemiological studies suggest gut inflammation's involvement in the development of PD pathology. Though the study is important and timely the authors are suggesting it as a novel rat model of gut inflammation and PD pathology that does not go with the results the authors have presented. The direction of the research manuscript as a development of novel model need to be changed due to lack of sufficient data to prove the novelty of the model. There are some major concerns which need to be addressed before acceptance of this manuscript.
Comments
1. In the abstract the authors say the aim is to identify the exacerbation of DA neuron degeneration due to gut inflammation, however, they did not observe the exacerbation of PD pathology. So this has to be changed.
2. Authors should cover the already published studies with DSS and neurotoxins of PD in the introduction.
3. The methods and statistical part is very clearly defined. However it will be better to provide the catalogue numbers of chemicals and antibodies used in the present study.
4. The authors have provided the explanation for similar DA neurodegeneration in both 6-OHDA and 6-OHDA + DSS group while greater motor impairment in 6-OHDA + DSS group with the levels of pro-inflammatory cytokines. However, the significance of pro-inflammatory cytokines in the 6-OHDA +DSS group is minimal. The authors can suggest the 6-OHDA insult is already so high inducing almost 60% DA neuronal loss that it may circumvent the effect on DA neurodegeneration in the presence of DSS or should provide a better rationale.
5. The reason this can not be considered a novel rat model is because 1. The authors have not sacrificed rats time-dependently and observed the development of phenotype. 2. For the animal model the authors should cover other factors too which are important in the PD perspective like alpha-synuclein pathology, and GFAP activation which is unfortunately not covered in this study.
6. Why the authors have not covered the striatum, is there are any staining the authors have performed in striatum?
7. Regarding microglial infiltration in 6-OHDA +DSS group which is less than 6-OHDA group, do the authors have better explanation. Is it possible the different stages of inflammation?
8. DA neuron degeneration figure. Can the authors provide high resolution image at high magnification?
9. There are some limitations of this study like the authors have not included the DSS alone group which would have provided a better comparison.

The manuscript can be accepted for publication