Impaired NCC Activity and Regulation on Dietary High Dietary Salt Intake in Aged Sprague Dawley Rats Is Associated with Increased Salt Sensitivity of Blood Pressure
Authors: Kim Kiyoung , Puleo Franco , Frame Alissa A. , Wainford Richard D. 
Published: April 2020

BackgroundHypertension (HTN) is one of the leading causes of death worldwide and its prevalence increases with age. Studies revealed that enhanced global sympathetic tone is related to increases in blood pressure (BP) in aged human subjects. Dietary sodium intake is associated with the regulation of blood pressure, as increased sodium retention is a well‐established precursor of hypertension risk. An excessive dietary sodium intake leads to increases in BP in individuals that demonstrate the salt sensitivity of BP and evoke the development of salt sensitive HTN (SSH). Recent studies have demonstrated that excessive release of norepinephrine by increased sympathetic nerve activity upregulates renal sodium chloride cotransporter (NCC) activity to promote sodium reabsorption. However, the underlying mechanisms of the regulation of sodium homeostasis and blood pressure in response to increase in sympathetic tone with age remain unclear.HypothesisNCC‐mediated sodium retention driven by increased sympathetic tone with age contributes to age‐related HTN. Furthermore, the activity and regulation of NCC are impaired on a high salt (HS) diet and this impaired NCC activity and regulation drives the development of age‐dependent SSH.MethodsThree different age groups (3, 8, 16 month old (MO)) of male Sprague‐Dawley (SD) rats were fed a normal salt (NS; 0.6% NaCl) or HS (4% NaCl) diet for 21 days respectively. On day 21, basal MAP, NCC activity (peak natriuresis to IV hydrochlorothiazide (2mg/kg) infusion), and vascular sympathetic tone (peak MAP change in response to ganglionic blockade (IV hexamethonium; 30mg/kg)) were measured in vivo. Plasma and renal NE concentrations were also assessed via ELISA. The expression of total NCC, phosphorylated NCC, with‐no‐lysine [K] kinases (WNK) 1, WNK4, STE20/SPS1‐related proline‐alanine‐rich protein kinase (SPAK), oxidative stress responsive kinase 1 (OxSR1), and phosphorylated SPAK/OxSR1 were assessed via immunoblotting (N=6/gp).ResultsFirst, 3MO SD rats are normotensive in both diet groups and show dietary sodium‐evoked suppression of NCC activity, expression, and phosphorylation and upstream regulatory kinases including WNK1, SPAK, and OxSR1 on a HS diet relative to a NS diet. Second, SD rats show age‐dependent HTN, increases in renal sympathetic tone, NCC activity and expression, and WNK1 expression. Last, SD rats show aged‐dependent SSH, increased sympathetic tone, and impaired suppression of NCC activity, phosphorylation, and the expression of kinases SPAK and OxSR1.ConclusionSD rats show age‐dependent HTN that is coupled with increases in sympathetic tone, NCC activity and expression, which strongly suggests that the NCC contributes to the development of age‐dependent HTN. Moreover, the NCC may play a pivotal role in the development of age‐dependent SSH as aged SD rats show impaired dietary‐sodium evoked suppression of NCC activity and regulation. Collectively, these findings suggest that increased sympathetic tone may drive NCC activity that evokes age‐dependent HTN and SSH.
MO, month old; NS, 0.6% NaCl; HS, 4% NaCl.

Variables

3 MO

8 MO

16 MO

NS

HS

NS

HS

NS

HS

MAP (mmHg)

124 ± 2

126 ± 3

135 ± 4
#

143 ± 5
#

149 ± 3
#
†

169 ± 1
*
#
†

ΔUNaV to HCTZ (μeq/min)

9 ± 1

7 ± 1
*

18 ± 2
#

16 ± 1
#

15 ± 5
#

16 ± 6
#

Plasma NE (nmol/L)

44 ± 4

28 ± 4
*

55 ± 3
#

42 ± 4
*
#

54 ± 4
#

41 ± 4
*
#

Renal NE (pg/mg)

612 ± 36

368 ± 32
*

835 ± 48
#

722 ± 44
*
#

974 ± 13
#
†

1019 ± 134
#
†

ΔMAP to hexamethonium (mmHg)

−33 ± 4

−24 ± 3
*

−64 ± 5
#

−65 ± 6
#

−60 ± 3
#

−67 ± 7
#

Total NCC expression (fold change)

1.00 ± 0.22

0.32 ± 0.09
*

1.91 ± 0.25
#

1.13 ± 0.15
*
#

1.82 ± 0.23
#

0.75 ± 0.11
*

Total pNCCT53 expression (fold change)

1.00 ± 0.20

0.19 ± 0.05
*

1.24 ± 0.46

0.87 ± 0.22

0.49 ± 0.18

1.73 ± 0.22
*
#

WNK1 expression (fold change)

1.00 ± 0.20

0.49 ± 0.06
*

1.07 ± 0.34

1.36 ± 0.15

2.15 ± 0.38
#
†

1.60 ± 0.29
#

WNK4 expression (fold change)

1.00 ± 0.11

0.86 ± 0.07

1.95 ± 0.14
#

1.28 ± 0.10
#

1.05 ± 0.07
†

1.17 ± 0.09

SPAK expression (fold change)

1.00 ± 0.12

0.48 ± 0.09
*

1.25 ± 0.09

1.30 ± 0.23
#

0.42 ± 0.04
#
†

0.41 ± 0.13
†

OxSR1 expression (fold change)

1.00 ± 0.12

0.62 ± 0.05
*

1.09 ± 0.12

1.10 ± 0.06
#

1.19 ± 0.20

1.17 ± 0.08
#

pSPAK/OxSR1 expression (fold change)

1.00 ± 0.24

0.84 ± 0.30

0.55 ± 0.23

1.42 ±0.24
*

1.12 ± 0.17

2.86 ± 0.43
*
#
†

*p<0.05 vs. respective NS group;
#p<0.05 vs. respective 3 MO group;
† p<0.05 vs. respective 8 MO group.