Effects of Atrial Natriuretic Peptide knockout on renal mitochondrial bioenergetics in salt‐sensitive hypertension
Authors: Sultanova Regina , Nikiforova Anna , Domondon Mark , Vasileva Valeriia Y. , Stadler Krisztian , Ilatovskaya Daria 
Published: April 2020

Background

Atrial Natriuretic Peptide (ANP) encoded by Nppa, is a hormone known to promote salt excretion and BP reduction; there are clinical data implicating inherently low levels of ANP in the development of salt‐sensitive (SS) hypertension. In the heart and fat tissue, ANP is known to affect lipid peroxidation, mitochondrial bioenergetics and biogenesis. However, there is a gap in knowledge regarding the effects of ANP on mitochondria in the kidney. We hypothesized here that in SS hypertension ANP deficiency causes renal mitochondrial dysfunction.

Methods

SS hypertension was induced in SSNPPA−/− (ANP knockout in Dahl SS background) and SSWT (wild type Dahl SS) rats by placing them on a high salt diet (HS, 4% NaCl) or a normal salt diet (control, NS, 0.4% NaCl) for 21 days. A combination of in vivo techniques with studies performed on isolated renal mitochondria (seahorse respiration and spectrofluorimetry assays) were used to test the role of ANP knockout in the mitochondria bioenergetics.

Results

Our previous studies demonstrated that during a HS challenge SSNPPA−/− rats exhibit exacerbated salt‐sensitivity of blood pressure, reduced sodium excretion, decreased urine flow, and aggravated kidney injury. In addition, there is a significant reduction in renal cortical cGMP production in the HS fed SSNPPA−/− rats vs SSWT. cGMP is known to exert protective effects in mitochondria; in order to test mitochondrial function in our model, we measured membrane potential and levels of superoxide and hydrogen peroxide (H2O2) in mitochondria isolated from kidney cortex. Specifically, TMRM, Amplex Red and MCLA were used to detect mitochondrial membrane potential, H2O2 and superoxide, respectively. We report a decrease in mitochondrial membrane potential in the SSNPPA−/− rats vs SSWT (both on NS and HS diets), and an increase in mitochondrial H2O2 and superoxide levels in the same groups. Swelling assay revealed a trend for an increased mitochondria swelling rate in the SSNPPA−/− rats. Pilot calcium uptake studies implicated calcium influx via MCU (mitochondrial calcium uniporter) and mitochondrial permeability transition pore (mPTP) opening in this process. Interestingly, seahorse respirometry performed on cortical mitochondria revealed elevation of the oxygen consumption rate (OCR) in the knockout rats on HS, while in SSWT rats HS diet feeding reduced OCR (both vs NS). In addition, we observed activation of the antioxidant system in the SSNPPA−/− rats as opposed to SSWT (on HS diet, assessed in a Trolox‐based assay).

Conclusions

Taken together, our data indicate that lack of ANP, via disruption of protective cGMP‐mediated pathways, results in significant alterations in mitochondrial bioenergetics in the renal tissue. By establishing the mechanistic link between ANP and mitochondrial function, further studies will advance the understanding of the mitochondria‐mediated mechanisms affecting renal disease progression.

Support or Funding Information

The study was supported by NIH R00 DK105160 and Dialysis Clinic Inc Reserve Funds.