Pressure natriuresis induced alterations in renal sodium transporters in diabetic and non-diabetic rats

Hypertension increases cardiovascular risk and renal disease in type 1 diabetes mellitus (T1DM)(1). The normal natriuretic response (pressure natriuresis, PN) that offsets increased blood pressure (BP) is markedly suppressed in T1DM rats(2). PN represents a rapid downregulation of renal tubular sodium transport(3), so we hypothesised that this downregulation is suppressed when BP is increased in T1DM. To investigate this and provide mechanistic insight, we measured the effect of T1DM on mRNA and protein expression of the sodium–hydrogen antiporter 3 (NHE3) and the thiazide-sensitive sodium transporter (NCC), with and without acute ramps in BP.

Adult male Sprague-Dawley rats were IP injected with streptozotocin (STZ, 35-50mg/kg) or sodium citrate vehicle. After 2 weeks, blood glucose confirmed T1DM status in STZ-injected rats. Animals were anaesthetised (thiopental 50 mg/kg IP). IV saline with 2% bovine serum albumin and 0.125% FITC inulin was infused for 2 hours at 0.2mL/10g/hr and BP was measured directly from the carotid artery. PN was induced by acutely ramping BP with sequential ligation of coeliac and cranial mesenteric arteries, then the distal aorta. Arteries were not ligated in sham rats. Finally, renal cortex samples were perfusion-fixed for immunofluorescence, immediately homogenised for Western blotting, or frozen for qPCR. There were 4 experimental groups: non-T1DM and T1DM rats that underwent induction of PN or sham (n=6 per group). Data were analysed by one-way ANOVA.

Ligation of arteries increased BP in both groups by ~40mmHg, with no difference between T1DM and non-T1DM rats (P=0.264). Sham blood pressure remained unchanged (P=0.363). Abundance of NHE3 mRNA was increased by T1DM (P=0.006), but following acute ramps in BP, NHE3 mRNA increased only in non-T1DM rats (P<0.001). Protein expression of NHE3 was unchanged by T1DM and decreased following acute BP ramps in both groups (non-diabetic P=0.024; T1DM P=0.003), but the effect was greater in T1DM rats (P=0.010). Abundance of NCC mRNA was unaffected by T1DM but was reduced following acute BP ramps in T1DM rats compared to non-T1DM rats (P=0.042). T1DM did not affect phosphorylated (active) NCC protein expression in the sham group (P=0.999) or impact the reduction in NCC protein expression (P=0.075) induced by acute BP ramps (T1DM P=0.002; non-T1DM P=0.024). Total NCC protein expression was unaffected by T1DM or BP ramps (P>0.05 for all). Immunofluorescence identified reduced NCC at the apical membrane of tubules when BP was increased.

T1DM does not suppress PN in rats by preventing downregulation of expression of NCC or NHE3 mRNA or protein. However, we have also identified a disconnect between increased BP and the expression of 2 sodium transporters in different parts of the nephron, regardless of diabetic status. Functional studies localising the nephron segment with inappropriate sodium transport are required to aid targeting and restoration of the PN response in T1DM rats.
Acknowledgement
Supported by Kidney Research UK (RP02/2019)
References
(1) Kim, Y. T. et al. (2020) ‘Risk of Cardiovascular Disease and Chronic Kidney Disease According to 2017 Blood Pressure Categories in Diabetes Mellitus’, Hypertension. doi: 10.1161/HYPERTENSIONAHA.120.15320.
(2) Culshaw, G. J. et al. (2019) ‘Impaired pressure natriuresis and non-dipping blood pressure in rats with early type 1 diabetes mellitus’, Journal of Physiology, 597(3), pp. 767–80. doi: 10.1113/JP277332.
(3) McDonough, A. A. (2010) ‘Mechanisms of proximal tubule sodium transport regulation that link extracellular fluid volume and blood pressure’, American Journal of Physiology - Regulatory Integrative and Comparative Physiology. doi: 10.1152/ajpregu.00002.2010.

Keywords: Pressure natriuresis, sodium balance, diabetes mellitus