DOCA Pellet Implantation

DOCA-salt and DOCA-salt+OX1R-shRNA groups were subjected to subcutaneous implantation of a DOCA pellet (75 mg, 21-day release, Innovative Research of America, FL, United States) prior to beginning the high salt drink treatment. Rats were given 5% isoflurane for anesthesia induction followed by 2–3% isoflurane to maintain adequate anesthesia during the procedure. An incision was made in the retro-scapular region, and the DOCA pellet was placed subcutaneously. The wound was sutured, and rats were given the same post-operative care as above. However, directly following the procedure, all rats’ drinking water was switched to a saline solution (1% NaCl and 0.2% KCl) for the remainder of the 21 days. As previously mentioned, DOCA treatment is usually paired with uninephrectomy to exacerbate the development of hypertension. However, we decided to exclude the kidney removal following a previously established model (Kandlikar and Fink, 2011) to obtain a more gradual progression of hypertension development that more closely mimics that seen in a humans, as well as to eliminate many of the adverse effects that traditional DOCA-salt and uninephrectomy incurs (Gavras et al., 1975; Wada et al., 1995; Loch et al., 2006).