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In order to produce HCO3- from glutamine and to subsequently reabsorb it, H+ ions need to be secreted. The kidney does this by via ammonium ions in the PCT. But why are the ammonium ions then reabsorbed in the TAL before being put back into the collecting duct? Wouldn't it be more efficient if they were just left in the tubule fluid where they would reach the collecting duct eventually? The first method seems to be a longer way of achieving the same result.
Isn't it all to do with the reabsorption of water, i.e. everything gets flushed out in the Bowman's capsule and then some stuff is reabsorbed to lower the water potential of blood, so water is reabsorbed as well.
I guess I figured it was easier to have a leakier sieve at the Bowman's capsule than try to only release the waste products via facilitated diffusion.
Why reabsorb $NH_4^+$ in the thick ascending limb if you're just going secrete it in the collecting duct
This is an excellent question. When you find an odd, seemingly inefficient aspect of physiology, it's an opportunity to learn something new.
This alternating secretion, reabsorption, and secretion starts to make sense when you look at the anatomy of the nephron.
There is no appreciable ammonia in the filtrate at Bowman's capsule (free ammonia is efficiently converted to urea in the liver by the urea cycle, so it's in very low concentrations in the blood). As you say, it is produced in the proximal convoluted tubule. The amount that is produced is regulated by the acid load (more acid, more ammonia production). The proximal tubule is in the cortex (top of the photo).
$NH_4^+$ is in equilibrium with $NH_3$ in the tubule, and renal tubular cells and junctions are relatively permeable to $NH_3$ here, so the more $NH_4^+$ is produced and actively pumped into the lumen, the more $NH_3$ backflow leaves the tubule and enters the ECF. Because of this, the primary work of the PCT in acid base homeostasis by the kidney is not secretion (though $NH_4^+$ is actively secreted here), but rather in the production of new $NH_4^+$ and new $HCO_3^-$.
The bulk of the work of secretion happens in the collecting duct. There are a number of passive and active transport mechanisms for the secretion of $NH_4^+$ here, especially in the medullary collecting duct. These mechanisms are aided by a high concentration of $NH_4^+$ and $NH_3$ in the medulla (at the bottom of the picture).
So where does this high concentration of ammonia come from? It's from the reabsorption of $NH_4^+$ in the thick ascending limb. This is reabsorption (the tubular cells take $NH_4^+$ from the lumen and send it to the ECF), but you can think of it more as taking it from the cortex and putting it in the medulla. The filtrate flows along the lumen of the tubule, toward the collecting duct. Much of it is in equilibrium between the lumen and the ECF before it gets to the thick ascending limb. At the thick ascending limb, the permeability of the tight junctions and the tubular cell membrane changes. Solutes (the one we're interested in right now is $NH_4^+$) are taken out of the lumen and left behind in the ECF. Because the filtrate flows but not the ECF, and because the tubular cells are permeable before the thick ascending limb, and impermeable after, this substantially concentrates these solutes in the medulla, and dilutes the solutes in the cortex.
This is a process known as the countercurrent multiplier. The same principle is used to create a hyperosmotic ECF (and consequently, allow for a hyperosmotic urine if you've been wandering in the desert for hours).
Here's a diagram of the nephron from a decent review article that illustrates each of these processes for ammonia handling:
The organs involved in regulation of external acid-base balance are the lungs are the kidneys.
The lungs are important for excretion of carbon dioxide (the respiratory acid) and there is a huge amount of this to be excreted: at least 12,000 to 13,000 mmols/day.
In contrast the kidneys are responsible for excretion of the fixed acids and this is also a critical role even though the amounts involved (70-100 mmols/day) are much smaller. The main reason for this renal importance is because there is no other way to excrete these acids and it should be appreciated that the amounts involved are still very large when compared to the plasma [H + ] of only 40 nanomoles/litre.
There is a second extremely important role that the kidneys play in acid-base balance, namely the reabsorption of the filtered bicarbonate. Bicarbonate is the predominant extracellular buffer against the fixed acids and it important that its plasma concentration should be defended against renal loss.
In acid-base balance, the kidney is responsible for 2 major activities:
- Reabsorption of filtered bicarbonate: 4,000 to 5,000 mmol/day
- Excretion of the fixed acids (acid anion and associated H + ): about 1 mmol/kg/day.
Both these processes involve secretion of H + into the lumen by the renal tubule cells but only the second leads to excretion of H + from the body.
The renal mechanisms involved in acid-base balance can be difficult to understand so as a simplification we will consider the processes occurring in the kidney as involving 2 aspects:
Thick Ascending Limb
The primary site of sodium reabsorption in the Loop of Henle is the thick ascending limb (TAL). The TAL is impermeable to water. Sodium (Na + ) reabsorption is active- the driver is the Na + /K + ATPase on the basolateral membrane which actively pumps three Na + ions out the cell into the interstitium and two potassium(K + ) ions into the cell. By creating a low intracellular concentration of sodium, the inside of the cell becomes negatively charged, creating an electrochemical gradient.
Sodium then moves into the cell (from the tubular lumen) down the electrical and chemical gradient, through the NKCC2 transporter on the apical membrane This transporter moves one Na + ion, one K + ion and two Cl – ions across the apical membrane. .
Potassium ions are transported back into the tubule by ROMK channels on the apical membrane to prevent toxic build up within the cell. Chloride ions are transported into the tissue fluid via CIC-KB channels.
The overall effects of this process are:
- Removal of Na + whilst retaining water in the tubules – this leads to a hypotonic solution arriving at the DCT.
- Pumping Na + into the interstitial space contributes to a hyperosmotic environment in the kidney medulla (see below)
There is also significant paracellular reabsorption of magnesium, calcium, sodium and potassium.
Thin Ascending Limb
Sodium reabsorption in the thin ascending limb is passive. It occurs paracellularly due to the difference in osmolarity between the tubule and the interstitium.
As the thick ascending limb is impermeable to water, the interstitium becomes concentrated with ions, increasing the osmolarity. This drives water reabsorption from the descending limb as water moves from areas of low osmolarity to areas of high osmolarity. This system is known as counter-current multiplication.
For further explanation of counter-current multiplication, please see this helpful video: https://www.youtube.com/watch?v=Vqce2dtg45U
Thin Descending Limb
The descending limb is highly permeable to water, with reabsorption occurring passively via AQP1 channels. Very low amounts of urea, Na + and other ions are also reabsorbed. . As mentioned above, water reabsorption is driven by the counter-current multiplier system set up by the active reabsorption of sodium in the TAL.
Fig 1 – Diagram showing ion and water reabsorption within the Loop of Henle.
Mechanism of type 4 renal tubular acidosis
There are several mechanisms of hyperkalemia and metabolic acidosis in this heterogenous group of disorders. The major roles in the pathogenesis are played by a decrease in renal ammonia excretion and by the increase in paracellular chloride reabsorption which results from this.
The role of hyperkalemia in the impairment of renal ammonia clearance
In the classical literature, much is made of the degree to which renal ammoniagenesis is impaired by hyperkalemia, and how this decreases H+ excretion. Of course, the relevance of H+ and NH3 excretion is minimal – after all, fresh water is a near-infinite source of H+ ions. The whole point of excreting NH4+ is to have a weak cation to excrete together with chloride, so that one does not waste one’s sodium and potassium.
Now, the impairment of renal ammoniagenesis can be viewed in terms of its influence on chloride excretion. The consequence of low urinary ammonia is chloride retention, and a decreasing strong ion difference. This decrease in the rate of ammonia generation has been attributed to hyperkalemia. This can be demonstrated, at least in rats. The major defect seems to be due to the impaired medullary ability to concentrate ammonium in its interstitial fluids. Remember that ammonium excreted in the proximal tubule is reabsorbed in the thick ascending limb as a part of a countercurrent multiplication mechanism which concentrates ammonia in the renal medulla. The highly concentrated ammonia is then excreted into the medullary collecting ducts. It has been shown that hyperkalemia interferes with the mechanism of ammonia concentration by interfering with the reabsorption of ammonium at the thick ascending limb. The job of reabsorbing ammonium belongs to the famously frusemide-related Na+/K+/2Cl- cotransporter, for the services of which potassium and ammonia compete.
So the decrease in ammonia reabsorption leads to decreased ammonia concentration and thus to diminished ammonia levels in the lumen of the distal convoluted tubule. This is the last series of gap junctions which are permeable to chloride (as it is known that cortical collecting duct gap junctions are pretty tightly shut to everything, chloride included). Diminished ammonia levels here echo diminished chloride levels. There is no chloride excretion without ammonium excretion.
Further downstream, in the cortical collecting duct, paracellular transport of chloride is now impossible. Under normal conditions, the actions of the ENaC channel would result in reabsorption of sodium here. Likewise, the ROMK potassium channel would excrete potassium into the lumen.
Now, let us consider what may happen if aldosterone receptors are not being activated. Sodium extraction from the tubular lumen would be greatly decreased thus potassium secretion would be greatly decreased because the driving electric potential difference is gone. Potassium stays in, and sodium stays out, which is essentially a single-phrase description of the electrolyte abnormalities in hypoaldosteronism.
Lastly, whatever chloride is present in the lumen of the cortical collecting duct becomes exposed to the activity of the chloride-bicarbonate kAE1 exchanger, which can increase the chloride retention even further (in a similar fashion to its role in the pathogenesis of type 1 (distal) renal tubular acidosis).
In this fashion, one can summarise by saying that type 4 tubular acidosis is a condition where multiple mechanisms conspire to decrease the renal capacity for chloride excretion, by interfering with the excretion of ammonium.
Ammonia production and transport in response to acidosis
Metabolic acidosis stimulates ammonia production and transport by renal epithelial cells. Acidosis stimulates glutamine uptake into the proximal tubule and upregulates the expression of ammonia-producing enzymes, glutaminase, GDH, and PEPCK6, 7, 9, 10). Metabolic acidosis also increases the apical NHE3 activity and protein abundance in the proximal tubule18).
As mentioned earlier, ammonia reabsorption in the thick ascending limb leads to medullary interstitial ammonia accumulation, thereby driving its secretion into the collecting duct. Metabolic acidosis stimulated NKCC2 mRNA and protein expression in the rat and increased NHE4 mRNA expression in mouse thick ascending limb cells11, 12).
Rh B Glycoprotein (Rhbg) and Rh C Glycoprotein (Rhcg) are recently recognized ammonia transporter family members. Chronic HCl ingestion increased Rhcg protein expression and altered its subcellular distribution in the collecting duct19, 20). Both global and collecting duct-specific Rhcg knockout mice excreted less urinary ammonia under basal conditions and developed more severe metabolic acidosis after acid loading21, 22).
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As the thin descending limb makes its way into the medulla the surrounding tissue becomes ever increasingly hypertonic and therefore the osmotic gradient increases the deeper the limb goes. As the limb is permeable to water it leaves the tubule via osmosis. The tube then bends back on itself and heads back up the cortex. The thin ascending limb is entered. Here salt enters the tubule passively due to the hypertonicity of the medulla creating a gradient. This results in a very high salt concentration at the bottom of the loop. The fluid moves on and enters the thick ascending limb. This has salt transporters and so salt is pumped into the medulla via active transport causing more water to leave the thin descending limb. The vasa recta has a similar countercurrent uptake system and only removes what is absorbed maintaining the medulla in a hypertonic state.
This is a challenging concept which is hard to explain. It is explained below in a different way:
- "Descending limb is permeable to water but not solutes"
- "The thick ascending loop is not permeable to water but solutes are pumped out"
- "Therefore, osmolarity of peritubular space is elevated, which draws water out of the descending limb"
- "Therefore, solute concentration of the fluid in the ascending limb is higher. causing more pumping"
- "Therefore, osmolarity of peritubular space is elevated, which draws more water out of descending limb"
Courtesy of Dr Ali Mobasheri (University of Nottingham School of Veterinary Medicine and Science)
Due to the transport of sodium and chloride from the thick ascending limb, the concentration of the urine as it passes up this limb decreases back down to around the level it was when it entered the loop. However the volume is greatly reduced. The collecting duct is where most concentration happens however it is only possible thanks to the incredibly high concentrations of NaCl at the bottom of the loop. This returning of the concentration back to the same level as when it entered the loop is important for retaining salt and also allows the concentration to be finely controlled by the collecting ducts without the loss of salt.
When there is excess water in the body, the excess fluid passes through the loop of henle because the fluid entering the loop is less concentrated already. The solutes only have so much osmotic potential and therefore are unable to draw the excess water from the lumen. This contributes to allowing the kidneys to produce dilute urine.
RENAL GLUTAMINE METABOLISM
Significant renal catabolism of glutamine occurs only during metabolic acidosis [ 23 ]. The measured rat renal arterial-venous difference is normally less than 3% of the arterial plasma concentration of glutamine. However, metabolic tracer studies indicate that in postabsorptive humans, a slightly greater proportion of the plasma glutamine is extracted by the kidneys [ 14 ]. Approximately 20% of the plasma glutamine is filtered by the glomeruli and enters the lumen of the nephron. The filtered glutamine is reabsorbed primarily by the epithelial cells of the proximal convoluted tubule [ 66 ]. It is initially transported across the apical brush border membrane, and subsequently most of the recovered glutamine is returned to the blood via transport across the basolateral membrane.
Utilization of the small fraction of extracted plasma glutamine requires its transport into the mitochondrial matrix where glutamine is deamidated by glutaminase and then oxidatively deaminated by glutamate dehydrogenase. Mitochondrial uptake of glutamine occurs via an electroneutral uniporter [ 67 ]. Kinetic measurements indicated that the rate of glutamine transport in isolated rat renal mitochondria is not rate-limiting for glutamine catabolism [ 68 ]. Therefore, either the activity of the mitochondrial glutamine transporter or the glutaminase must be largely inactivated in vivo during normal acid-base balance to account for the effective reabsorption of glutamine.
Increased renal ammoniagenesis and gluconeogenesis from plasma glutamine constitute an adaptive response that partially restores acid-base balance during metabolic acidosis (Fig. 5) [ 69 ]. Thus, the renal catabolism of glutamine is rapidly activated following the acute onset of metabolic acidosis. Within 1–3 h, rat renal extraction of plasma glutamine exceeds the percent filtered by the glomeruli [ 21 ]. Thus, the direction of the basolateral glutamine transport must be reversed in order for the proximal convoluted tubule cells to extract glutamine from both the glomerular filtrate and the venous blood. In addition, the transport of glutamine into the mitochondria is acutely activated [ 70 ]. Further responses include a prompt acidification of the urine that results from an acute activation of the apical Na + /H + exchanger [ 71 ]. This process facilitates the rapid removal of cellular ammonium ions and ensures that the bulk of the ammonium ions generated from the amide and amine nitrogens of glutamine are excreted in the urine. Finally, a pH-induced activation of α-ketoglutarate dehydrogenase reduces the intracellular concentrations of α-ketoglutarate and glutamate [ 72 ]. Thus, increased renal catabolism of glutamine initially results from a rapid activation of key transport processes, an increased availability of glutamine, and a decreased concentration of the products of the glutaminase and glutamate dehydrogenase reactions.
During chronic metabolic acidosis, the kidney continues to extract more than one-third of the total plasma glutamine even though many of the acute adaptations are partially compensated (Fig. 5). Renal catabolism of glutamine is now sustained by increased expression of the genes that encode the mitochondrial glutaminase [ 18 ], the mitochondrial glutamate dehydrogenase [ 73 ], and the cytoplasmic phosphoenolpyruvate carboxykinase (PEPCK) [ 74 ]. All three adaptations occur solely within the proximal convoluted tubule, the primary site of renal ammoniagenesis [ 75 ]. The decreases in plasma pH and HCO3 − concentration during metabolic acidosis produce a comparable and sustained decrease in the intracellular pH (pHi) within the proximal convoluted tubule [ 76 ]. Thus, the adaptive increases in glutaminase and PEPCK activities may be initiated by a decrease in pHi. Both adaptations result from increased rates of synthesis of the proteins [ 77 , 78 ] that correlate with comparable increases in the levels of their respective mRNAs [ 79 , 80 ]. However, the increases in glutaminase and glutamate dehydrogenase result from the selective stabilization of their mRNAs, whereas the increase in PEPCK activity results from enhanced transcription of the PEPCK gene [ 5 ]. The levels of the mitochondrial glutamine transporter [ 70 ], the SN1 glutamine transporter [ 33 ], the apical Na + /H + exchanger, NHE3 [ 81 ], the basolateral Na + -3HCO3 − co-transporter [ 81 ], the apical Na + /dicarboxylate co-transporter, NaDC-1, [ 82 ] and the medullary Na + K + -2Cl – co-transporter [ 83 ] are also increased during chronic acidosis.
The excretion of ammonium ions is facilitated by a countercurrent mechanism [ 84 ]. Increased renal ammoniagenesis occurs primarily within the proximal convoluted tubule. The increased expression of the apical Na + /H + exchanger facilitates an increased acidification of the fluid within the lumen of the proximal tubule that ensures the initial trapping of the ammonium ions. Ammonium ions are subsequently reabsorbed within the thick ascending limb of the loop of Henle. This leads to the formation of a steep cortical to papillary concentration gradient that provides the driving force to secrete ammonium ions into the luminal fluid of the collecting ducts. The gradient is formed by the appropriate localization of various isoforms of the Rh-glycoproteins that function as selective ammonium ion transporters [ 85 ]. As a result, during chronic acidosis, ammonium ion excretion continues to provide an expendable cation that facilitates the excretion of titratable acids while conserving sodium and potassium ions. The increased Na + /H + exchanger activity also promotes the tubular reabsorption of HCO3 − ions. In addition, the α-ketoglutarate generated from glutamine is converted to glucose or oxidized to CO2. Either process requires the cataplerotic activity of PEPCK to convert intermediates of the tricarboxylic acid cycle to phosphoenolpyruvate. This product is either utilized in gluconeogenesis or converted to pyruvate that re-enters the mitochondria and is oxidized to CO2. However, both pathways generate two HCO3 − ions per mole of α-ketoglutarate. The increase in basolateral Na + -3HCO3 − co-transporter activity facilitates the vectoral translocation of reabsorbed and of de novo-synthesized HCO3 − ions into the renal venous blood. Thus, the combined adaptations also create a net renal release of HCO3 − ions that partially compensate the systemic acidosis.
VII. DISTAL CONVOLUTED TUBULE
The specific role of the distal convoluted tubule (DCT) in ammonia transport is incompletely understood. Because of technical issues, isolated perfused tubule studies examining DCT ammonia transport have not been performed. A limited number of micropuncture studies have examined ammonia transport between the rly” and “late” micropuncturable distal tubule, a region which includes both the distal convoluted tubule and portions of the connecting segment. There appear to be low rates of ammonia secretion, and the rate of ammonia secretion is not increased by in vivo chronic metabolic acidosis (266). Mathematical modeling studies also suggest the DCT secretes ammonia (315).
Why are ammonium ions reabsorbed in the thick ascending limb during acid secretion? - Biology
REGULATION OF BODY POTASSIUM
K + is the major intracellular ion. Only 2% is in the ECF at a concentration of only 4 mEq/L. K is taken up by all cells via the Na-K ATPase pump. It is one of the most permeable ion across cell membranes and exits the cells mostly via K channels (and in some cells via K-H exchange or via K-Cl cotransport).
K is the major ion determining the resting membrane electrical potential, which in turn, limits and opposes K efflux. Thus changes in K concentrations (particularly in the ECF) have marked effects on cell excitability (heart, brain, nerve,muscle).
K is the mayor intracellular osmotically active cation and participates in cell (intracellular) volume regulation (exits with Cl when cells swell).
A constant cell K concentration is critical for enzyme activities and for cell division and growth.
Intracellular K participates in acid base regulation through exchange for extracellular H and by influencing the rate of renal ammonium production.
Regulation of extracellular K is by tissue buffering (uptake of K excess) and by slower renal excretion.
2. Cellular K buffering
When K is added to the ECF, most of the added K is taken up by the cells, reducing the ECF K + increase. Similarly, if K is lost from the ECF, some K + leaves the cells, reducing the ECF K decline.
Buffering of ECF K through cell K uptake is impaired in the absence of aldosterone or of insulin or of catecholamines.
Cell K exit to the ECF increases when osmolarity increases (as in diabetes mellitus) and in metabolic acidosis, when it is exchanged for ECF protons (H + ). When cells die, they release their very high K content to the ECF.
3. Renal regulation of Potassium
In normal function, renal K excretion balances most of the K intake (about 1.5 mEq/Kg per day). The kidneys excrete about 15 % of the filtered K load of 10 mEq/Kg per day.
Along the proximal tubule the K concentration remains nearly equal to that in plasma. Since the PCT reabsorbs about 2/3 of the filtrate water, it also reabsorbs about 2/3 (66%) of the filtered K. This reabsorption is mostly passive and is driven by the positive tubule electrical potential present along the S2 and S3 segments and by paracellular solvent drag.
Along the descending limb of the loop of Henle, K is secreted into the tubule lumen from the interstitium. Along the thick ascending limb, K is reabsorbed via Na-K-2 Cl cotransport. In the loop, there is net K reabsorption of 25% of the filtered K.
Along the distal tubule and collecting ducts, there is net secretion of K which is stimulated by aldosterone and when there is dietary K excess. Secretion decreases and becomes net reabsorption in K deficiency. Regulation of renal K excretion is in the CD and is mostly by changes in the rate of K secretion.
In the CD, K secretion is by the principal cells (via luminal K channels and basolateral Na-K ATPase) and K reabsorption is by the alpha intercalated cells via a luminal H-K ATPase.
K secretion from principal cells into the CD lumen is enhanced by luminal and cellular determinants:
Luminal determinants that stimulate K secretion are increases in tubule urine flow (which reduces the intratubular K concentration), the delivery of sodium to the CD, and the delivery of poorly reabsorbed anions (other than Cl) to the CD. Na delivery followed by its reabsorption increases K secretion by increasing the lumen negative electrical potential and by stimulating the activity of the Na-K ATPase which results in enhanced accumulation of K in the cells. The presence in the CD of poorly reabsorbed anions (SO4 2- , excess of HCO3, beta hydroxybutyrate, or HPO4 2- )enhances the negativity of the CD lumen, favoring K secretion.
Cellular determinants of K secretion are the activity and abundance of K channels at the luminal cell membrane and of Na-K ATPase at the basolateral membrane. Both of these are enhanced primarily by aldosterone, and also by ADH (by decreasing urine flow, ADH reduces K secretion, but by increasing luminal permeability, ADH promotes it) and by dietary K excess. K deficiency is associated with increased activity and expression of luminal H-K ATPase in the alpha intercalated cells of the CD, which act to promote reabsorption of K from the lumen.
Clinical Relevance - Diabetes Insipidus (DI)
This form of diabetes also involves the classic presentation of polyuria (increased frequency of urination), and subsequent polydipsia (excessive thirst). It can be due to either insufficient ADH release from the posterior pituitary gland (central diabetes insipidus), or the collecting ducts not responding to ADH (nephrogenic diabetes insipidus). If the actions of ADH are ineffective, less water will be reabsorbed from the filtrate. This means there will be a greater volume of filtrate, hence producing a greater volume of urine, causing the polyuria and polydipsia.
A water deprivation test can be used as a confirmatory test for diabetes ins, after ruling out other common causes of polydipsia, such as hypercalcaemia. Management depends on the cause in central DI, desmopressin (synthetic ADH) can be used whilst thiazide diuretics are used in the treatment of nephrogenic DI.