Proximal tubule-specific glutamine synthetase deletion alters basal and acidosis-stimulated ammonia metabolism

Abstract

Lee H, Osis G, Handlogten ME, Lamers WH, Chaudhry FA, Verlander JW, Weiner ID. Proximal tubule-specific glutamine synthetase deletion alters basal and acidosis-stimulated ammonia metabolism. Am J Physiol Renal Physiol 310: F1229 -F1242, 2016. First published March 23, 2016 doi:10.1152/ajprenal.00547.2015.-Glutamine synthetase (GS) catalyzes the recycling of NH 4 ϩ with glutamate to form glutamine. GS is highly expressed in the renal proximal tubule (PT), suggesting ammonia recycling via GS could decrease net ammoniagenesis and thereby limit ammonia available for net acid excretion. The purpose of the present study was to determine the role of PT GS in ammonia metabolism under basal conditions and during metabolic acidosis. We generated mice with PT-specific GS deletion (PT-GS-KO) using Cre-loxP techniques. Under basal conditions, PT-GS-KO increased urinary ammonia excretion significantly. Increased ammonia excretion occurred despite decreased expression of key proteins involved in renal ammonia generation. After the induction of metabolic acidosis, the ability to increase ammonia excretion was impaired significantly by PT-GS-KO. The blunted increase in ammonia excretion occurred despite greater expression of multiple components of ammonia generation, including SN1 (Slc38a3), phosphate-dependent glutaminase, phosphoenolpyruvate carboxykinase, and Na ϩ -coupled electrogenic bicarbonate cotransporter. We conclude that 1) GS-mediated ammonia recycling in the PT contributes to both basal and acidosis-stimulated ammonia metabolism and 2) adaptive changes in other proteins involved in ammonia metabolism occur in response to PT-GS-KO and cause an underestimation of the role of PT GS expression.acid-base; ammonia; glutamine synthetase; proximal tubule A MAJOR FUNCTION of the kidney is to maintain acid-base homeostasis under basal conditions and in response to a variety of physiologic disturbances (29). In the kidney, ammonia 1 metabolism is a central component of basal net acid excretion, and changes in ammonia metabolism and urinary excretion are the primary components of the renal response to metabolic acidosis (62,64,66). Accordingly, understanding the molecular mechanisms of renal ammonia metabolism is central to understanding mammalian physiology and pathophysiology.Renal ammonia metabolism involves integrated functions of intrarenal ammoniagenesis and epithelial cell-specific NH 3 and NH 4 ϩ transport. Ammoniagenesis involves cellular glutamine uptake and metabolism through an enzymatic process involving phosphate-dependent glutaminase (PDG), glutamate dehydrogenase, and phosphoenolpyruvate carboxykinase (PEPCK) (7,59,60,63,65,66). Ammonia produced in the kidney undergoes regulated transport of both NH 3 and NH 4ϩ by a variety of specific proteins, including Naϩ -ATPase, and Rh glycoproteins B and C (Rhbg and Rhcg, respectively) (10,25,28,63,66,67). This integrated interaction of intrarenal ammonia production and transport facilitates highly regulated renal ammonia metabolism and excretion.However, not all ammonia...

Recent advances in understanding renal ammonia metabolism and transport

“…Because previous studies have shown that glutamine synthetase is expressed in cells involved both in ammonia production, i.e., proximal tubule cells, and cells involved in transepithelial ammonia transport, i.e., distal convoluted tubule and intercalated cells, and that glutamine synthetase expression can be differentially regulated in these different cell populations [20], the authors of this recent study used cell-specific glutamine synthetase deletion to determine the specific role of proximal tubule glutamine synthetase in renal ammonia metabolism. Proximal tubule-specific deletion increased basal ammonia excretion (Figure 3), consistent with decreased ammonia recycling leading to increased net ammonia generation and excretion [25]. There also was an impaired ability to respond to an acid load [25], indicating the ability to alter glutamine synthetase-mediated ammonia recycling is critical to increasing net ammoniagenesis.…”

“…Proximal tubule-specific deletion increased basal ammonia excretion (Figure 3), consistent with decreased ammonia recycling leading to increased net ammonia generation and excretion [25]. There also was an impaired ability to respond to an acid load [25], indicating the ability to alter glutamine synthetase-mediated ammonia recycling is critical to increasing net ammoniagenesis. These findings demonstrate the presence of a “yin-yang” mechanism regulating “net” ammoniagenesis, one involving ammonia generation and another involving ammonia recycling.…”

“…Right panel shows that increased ammonia excretion occurred despite no change in urinary pH. Figure from [25] and reprinted with permission.…”

NBCe1-A Regulates Proximal Tubule Ammonia Metabolism under Basal Conditions and in Response to Metabolic Acidosis

“…The proximal tubule can recycle ammonia via the cytosolic enzyme, GS, which decreases net ammonia generation. [32][33][34][35] Metabolic acidosis normally decreases GS expression. 32,34 GS expression in NBCe1-A KO mice was not suppressed despite the basal metabolic acidosis ( Figure 10).…”

“…[32][33][34][35] Metabolic acidosis normally decreases GS expression. 32,34 GS expression in NBCe1-A KO mice was not suppressed despite the basal metabolic acidosis ( Figure 10). In the cortex, acid loading decreased GS expression in both WT and KO mice, but expression was significantly greater in KO mice despite more severe acidosis.…”