Effects of Acute Sepsis on Renal Structure and Sympathetic Innervation in Mice
Authors: Alkhateeb Tuqa , Ozment Tammy R. , Youngberg George A. , Howell Mary E. , Stuart Charles A. , Price Jennifer B. , Jones Thomas C. , Williams David L. , Blair Tesha E. , Hoover Donald B. 
Published: November 2023

Sympathetic nerves have major roles in renal physiology and the pathophysiology of sepsis. Recent work has shown that rapid sprouting of sympathetic nerves occurs in the spleen of septic mice. This study was done to investigate effects of acute sepsis on renal morphology in mice and to test the hypothesis that sprouting of sympathetic nerves also occurs in septic kidney. Sepsis was induced in male C57BL/6 or ChAT‐eGFP mice at 4–6 months of age by cecal ligation and puncture (CLP). Sham control mice underwent similar surgery without CLP. Both sham and CLP mice received a subcutaneous injection of resuscitation fluid (1 ml lactated Ringers) immediately after surgery. Mice were euthanized with isoflurane at 16 h post‐surgery and the kidneys were removed for histological or biochemical evaluation. Serum was collected to measure levels of cystatin C, an indicator of renal function. Paraffin sections were stained with PAS or H&E. Digital images of stained renal corpuscles were collected and used to measure area of the glomerulus and Bowman's capsule. ChAT‐eGFP mice were used to evaluate sympathetic nerve density and screen for eGFP+ non‐neuronal cholinergic cells in kidney. Frozen, 100 μm longitudinal sections of kidneys were immunolabeled for the sympathetic nerve marker, tyrosine hydroxylase (TH), and for eGFP. Images of stained sections were collected by confocal microscopy and marker densities quantified using ImageJ. Bowman's space was significantly smaller in septic mice (Sham: 633 ± 92 and CLP: 304 ± 33 μm2; n=3 per group, P<0.05), and many cortical tubular cells had a bubbly appearance. Both of these results suggest acute renal pathology, however, cystatin C levels were not elevated (Sham: 376 ± 22 and CLP: 325 ± 11 ng/ml, n=6 per group, P>0.05). Acute sepsis did not affect sympathetic nerve density determined by confocal analysis or the abundance of TH determined by Western blotting. However, preliminary evidence indicates that renal levels of the sympathetic neurotransmitter, norepinephrine, are reduced in the kidney at 16 h post‐CLP (Sham: 353 ± 25 and CLP: 129 ng/g ± 20; n=6 per group, p<0.0001). While the kidney lacks cholinergic innervation, we were surprised to find that many tubular cells stained for eGFP, suggesting they might have a cholinergic phenotype. Acute sepsis did not affect either the density of GFP+ cells in confocal analysis or the abundance of GFP in Western blots. We conclude that septic mice, show only minor renal pathology in the early stages of disease. Furthermore, we found no evidence for noradrenergic nerve sprouting in the kidneys, whereas in a previous study, noradrenergic nerve density in the spleen more than doubled using the same model. This difference suggests that sprouting of sympathetic nerves is not a general phenomenon in sepsis and may be limited to lymphoid tissue. Further studies are needed to understand the mechanism and consequences of reduced norepinephrine in the kidney and to determine the role of cholinergic tubular cells in renal physiology and pathophysiology.

Support or Funding Information

AHA 11SDG53330002 Career Scientist Development Award (TRO); NIH GM107949 (DBH); NIH GM53522 and NIH GM117179 (DLW); NIH GM083016 (CL and DLW); NIH HL071837 (CL) and C06RR0306551.