Fischer 344 rats

For volume of distribution experiments, a biological replicate of three was performed to ensure reproducibility and accuracy of our data, as previously reported in literature by our group. For flow cytometry, a biological replicate of five was performed to repeat the experiment on three different days, accounting for any processing variables and to ensure day to day variation. Moreover, preliminary experiments demonstrated that N=5 was sufficient to ensure the detection of significant differences. Animals that died during procedures were not processed further for analysis. For the therapeutic efficacy study, an initial number of 10 animals per group was previously shown in our group to ensure statistical differences. Animals dying without waking up from surgery (after tumour implantation or CED) were excluded. Animals were randomly assigned to the different groups to prevent any confounding variables from syringes or stereotactic frames. Investigators were not formally blinded to experimental groups.
All procedures were performed in accordance with the guidelines and policies of the Yale Animal Resource Center (YARC) and approved by the Institutional Animal Care and Use Committee (IACUC). Male Fischer 344 rats (Charles River Laboratories, 200–220 g) were used. Surgical procedures were performed using standard sterile surgical techniques.

The Animal Ethics Committee of the University Rovira i Virgili (Tarragona, Spain) approved all the procedures. Twenty-four 8-weeks-old male Fischer 344 rats (Charles River Laboratories, Barcelona, Spain) were housed in pairs in cages at 22 °C under two different light schedules in order to emulate season’s day length: long day photoperiod (n = 12, LD, 18:6h light/dark cycle) or short-day photoperiod (n = 12, SD, 6:18 h light/dark cycle). After 1 month under these conditions, animals in each photoperiod were orally supplemented with lyophilized red grapes [29 (link)] (100 mg per kg of body weight/day) (grape, n = 12) as an autumn fruit for 10 weeks or a control vehicle (control, n = 12). Rats were fed an ad libitum standard diet (2.90 kcal/g; A04, Panlab, Barcelona, Spain) and, after 14 weeks, they were deprived of food for 1 h and sacrificed by decapitation. Each brain was rapidly removed after death, flushed with cold isotonic saline buffer, weighed, frozen in liquid nitrogen and stored at −80 °C until further analysis.

Three-month-old male and female Fischer 344 rats were purchased from Charles River Laboratories. For the ddPCR validation study, 8 per sex/group (total n = 32) were used. For assessment of pSyn inclusions in the SN and FISH, 8 males per group and 10 females per group were used (total n = 36). Rats were housed 1-3 per cage in a room on 12 h light/dark cycle, and food and water were provided ad libitum. In all cases where an animal was single housed, enrichment was provided. All animal work was performed in the Michigan State University Grand Rapids Michigan State Research Center vivarium. All procedures were approved and conducted in accordance with the Michigan State University Institute for Animal Care and Use Committee (IACUC) at Michigan State University.