L5 Renal Physiology 5

Overview

This lecture covers acid-base balance: why plasma pH is held in a very narrow range, the two acid loads the body must deal with (CO2 from metabolism and non-volatile acids), and how buffering plus excretion keep pH constant. CO2 is buffered in transit by haemoglobin and removed by alveolar ventilation, so respiratory disturbances act through pCO2. Non-volatile acid is buffered by bicarbonate, protein and phosphate, which consumes HCO3-, so the kidney must both reclaim filtered HCO3- and generate “new” HCO3- while excreting H+ on urinary buffers (titratable acid and ammonia). The Henderson-Hasselbalch equation ties the two control systems together, and the final sections apply the framework to renal tubular acidosis and to the four classes of respiratory and metabolic disturbance.

Body fluid pH and why it matters

• ECF pH is 7.4, normal range 7.36 to 7.45. Intracellular pH is nearer 7.
• pH below 7.35 is acidosis; pH above 7.45 is alkalosis.
• Plasma pH below 7 or above 7.8 is usually fatal.
• Expressed as H+ concentration, pH is regulated within 36 to 44 nmol/L.

pH must be controlled because it influences:

• Enzyme activity (each enzyme has an optimum pH).
• Ion transport mechanisms.
• Hormone function.
• Cellular metabolism, DNA synthesis and growth.
• CNS function. Alkalosis causes headache, drowsiness, tetany and convulsions. Acidosis affects respiratory control and causes drowsiness progressing to coma.

The scale of the problem

• At pH 7 there is only about 0.004 mmol of H+ in body fluid, and it must be kept within about +/- 0.0004 mmol.
• Yet the body produces roughly 13000 mmol of “acid” per day, from CO2 of metabolism and from non-volatile acids of metabolism.
• Control is achieved by two processes: buffering and excretion.

R.F. Pitts, 1953, demonstrated how effective buffering is: injecting a dog with 150 mL of 1 mol/L HCl moved blood pH from 7.44 to only 7.14, whereas adding the same load to a similar volume of water gave pH 7.44 to 1.84. That is buffering of more than 99.99%.

The two day-to-day challenges

1. CO2 produced by cells, acting through the reaction $C{O}_2+H_{2}O\leftrightarrow H_{2}C{O}_3\leftrightarrow H^{+}+HC{O}_3^{-}$.
2. Acid produced from metabolism: phosphoric acid, sulfuric acid, lactic acid and ketone bodies, collectively the “non-volatile acids”.

CO2 as an acid load

• About 13000 mmol CO2 is produced per day, roughly 10 mmol/min, which is potentially 13000 mmol of acid via the hydration reaction.
• CO2 can be removed by the lungs, so it is not normally a problem, but two issues remain: CO2 is not produced in the lungs and so must be buffered in transit, and ventilation must continue.

Buffering CO2 in transit

Mostly buffered in blood by haemoglobin.

• In the tissues: $C{O}_2+H_{2}O\to H_{2}C{O}_3\to H^{+}+HC{O}_3^{-}$ (H+ made), then $H^{+}+O_{2}H{b}^{-}\to HHb+O_2$. HHb buffers the H+, so there is very little pH change from CO2 production.
• In the lungs the reactions reverse: $HHb+O_2\to H^{+}+O_{2}H{b}^{-}$, then $H^{+}+HC{O}_3^{-}\to H_{2}C{O}_3\to C{O}_2+H_{2}O$ (CO2 removed).

Alveolar ventilation

Exhalation of CO2 must continuously match the metabolic production rate.

• Hypoventilation: CO2 is not removed and accumulates, the equation shifts to the right, giving excess H+, that is respiratory acidosis.
• Hyperventilation: excess CO2 is removed, the equation shifts to the left, giving decreased H+, that is respiratory alkalosis.

Case (Annabel): comes to A&E very anxious after what her partner describes as a panic attack, with numbness and tingling in hands and feet and, more recently, cramps and spasms of the hands. Na 137 mmol/L, Cl 112 mmol/L, K 4.2 mmol/L, PCO2 29 mmHg, pH 7.52, HCO3 23 mmol/L.

Non-volatile acids

• Metabolism produces 50 to 80 mmol/day, the net endogenous acid production (NEAP).
• Sources: H2SO4 from sulfur-containing amino acids; H3PO4 from phosphoproteins and phospholipids; organic acids such as lactate and pyruvate from anaerobic exercise; ketoacids from fat metabolism.
• The H+ from these acids must be both buffered and excreted.

Buffering is by multiple mechanisms, described as “whole body buffering”:

• Three main buffers: bicarbonate, protein and phosphate.
• Plus dilution and exchange with bone minerals.
• This prevents pH falling to dangerous levels.

Buffering only hides the H+; it still has to be removed. Using the HCO3-/respiratory mechanism, buffered H+ reacts with HCO3-: $H^{+}+HC{O}_3^{-}\to H_{2}C{O}_3\to C{O}_2+H_{2}O$. The CO2 is exhaled and the H+ is now part of a water molecule, so it is no longer an acid threat. But the process has consumed an HCO3-, which the kidney must replace.

Renal handling of bicarbonate

The kidney handles bicarbonate in two ways:

1. Replacement of filtered HCO3-, referred to as “reabsorption”: “new” HCO3- is synthesised by proximal tubule cells to replace the filtered ions.
2. Replacement of bicarbonate consumed in buffering and excretion of non-volatile acids, or excretion of excess bicarbonate.

1. HCO3- “reabsorption”

Sites and contributions:

• Proximal tubule 80%, via NHE-3.
• Thick ascending limb 10%.
• Distal tubule / collecting duct 10%, via intercalated cells using H+/K+ ATPase and H+ ATPase.
• Some intercalated cells instead secrete HCO3- (in exchange for Cl-).

Mechanism in the proximal tubule cell (lumen to blood):

1. Filtered Na+ and HCO3- arrive in the lumen.
2. The apical Na+/H+ exchanger (NHE-3) secretes H+ into the lumen.
3. Luminal H+ combines with filtered HCO3- to form H2CO3, via carbonic anhydrase.
4. H2CO3 breaks down to H2O and CO2, and the CO2 diffuses into the cell.
5. Inside the cell, CO2 + H2O forms H2CO3 (carbonic anhydrase), which dissociates to HCO3- and H+.
6. The H+ is recycled to the apical exchanger; the HCO3- exits across the basolateral membrane into blood, so bicarbonate is reabsorbed.
7. The basolateral Na+/K+ ATPase pumps 3Na+ out and 2K+ in, driving the apical exchange.

Warning

The reabsorption slide carries an isolated “0” printed at the bottom left, whose meaning is not stated on the slide.

2. Replacement of HCO3- consumed in buffering (renal net acid excretion, RNAE)

• The urinary buffers HPO4²⁻ and NH3 combine with H+ secreted by tubule cells, and the products are excreted in the urine.
• When tubule cells make H+ for secretion they also produce HCO3-, via $C{O}_2+H_{2}O\leftrightarrow H_{2}C{O}_3\leftrightarrow H^{+}+HC{O}_3^{-}$.
• This “new” HCO3- is returned to plasma to replace the HCO3- consumed in buffering non-volatile acid.

Two mechanisms:

1. Titratable acid: urinary buffer HPO4²⁻, in the distal tubule.
2. Ammonia: glutamine to NH3 to NH4+, starting in the proximal tubule, passing through the medulla, and acting in the collecting duct (glycoproteins).

Titratable acid, cell level: filtered HPO4²⁻ in the lumen combines with H+ secreted by an ATP-driven H+ transporter to form H2PO4-, which is excreted. Inside the cell, CO2 + H2O gives H2CO3 (carbonic anhydrase), which yields HCO3- (exiting to blood) and H+ (secreted into the lumen).

Ammonia, cell level (late distal / collecting duct acid-secreting cell): H+ secreted by an ATP-driven transporter combines with NH3 in the lumen to form NH4+, which is excreted. Inside the cell, CO2 + H2O gives H2CO3 (carbonic anhydrase), yielding HCO3- to blood and H+ to the lumen. The NH3 comes from glutamine, converted by glutaminase to glutamate plus NH3.

Ammonia handling along the nephron:

• In the proximal-tubule-type cell, glutamine is metabolised to produce A⁼ and 2NH4+ (with H+/NH3 exchange and Na+-coupled transport). The A⁼ is converted to 2HCO3-, which exits to blood, and the NH4+ exits into the tubular fluid via a Na+/H+-type exchanger.
• NH4+ is then carried down the descending limb and recycled/concentrated through the loop of Henle into the medullary interstitium, feeding the collecting duct region.
• In cortical/outer-medullary collecting duct cells, NH3 diffuses into the tubular fluid and combines with secreted H+ (generated by CO2 + H2O via carbonic anhydrase, with HCO3- exiting to blood) to form NH4+, which is trapped in the lumen.
• A further collecting-duct-type cell shows NH4+ and H+ moving directly between tubular fluid and blood across both membranes.

Warning

The nephron ammonia-handling slide has no title, heading or accompanying text; the description above is a reading of the labelled pathway elements only, and the slide does not state its overall message.

Putting it together: the fate of a non-volatile acid

For new non-volatile acid HA:

• HA dissociates to A- + H+, and the H+ is buffered. This is the acid made.
• $H^{+}+HC{O}_3^{-}\to C{O}_2+H_{2}O$: bicarbonate is consumed.
• A- is filtered by the kidney.
• The kidney supplies H+ and generates new HCO3-.
• Filtered A- and the secreted H+ recombine to form HA, which is excreted.

The Henderson-Hasselbalch framework

$pH=6.1+\log \frac{[HC{O}_3^{-}]}{0.03\times pC{O}_2}$

• The numerator, HCO3-, is under renal control.
• The denominator, pCO2, is under respiratory control.
• The underlying reaction is $C{O}_2+H_{2}O\leftrightarrow H_{2}C{O}_3\leftrightarrow H^{+}+HC{O}_3^{-}$.

Maintaining balance therefore needs:

• Appropriate respiratory function, to maintain CO2.
• Appropriate renal function: the kidney must be able to function, must handle HCO3- appropriately, and must secrete acid appropriately.

Renal tubular acidosis

Two failure modes map onto the two renal requirements.

Proximal RTA (inappropriate HCO3- handling)

• Defect in the mechanism for “reabsorbing” HCO3-.
• HCO3- is lost, producing acidosis.
• Causes: congenital, paraproteins (myeloma), PTH, drugs.

Distal (“classic”) RTA (inappropriate acid secretion)

• Defect in the non-volatile acid secretion mechanisms.
• Acid is not excreted, producing acidosis.
• Causes: congenital, inflammation (for example SLE), toxins (amphotericin), hyperkalaemia, Conn’s.

Case (Beryl, proximal): several problems including vertebral collapse and recurrent UTIs; urine is alkaline and contains abnormal amounts of glucose, phosphate and amino acids. ABG on air: PaCO2 33 mmHg, pH 7.25, HCO3 14 mmol/L. Interpretation: glucose, PO4 and amino acids in the urine point to the proximal tubule; proximal tubule failure to reabsorb HCO3- means HCO3- is lost, causing acidosis (Fanconi syndrome).

Case (Camilla, distal): treated for SLE for 5 years but discontinued medication a few months ago because of side effects, and has become increasingly unwell with problems including diffuse lupus nephritis. ABG on air: PaCO2 22 mmHg, pH 7.37, HCO3 13 mmol/L, urine pH 7. Interpretation: urine pH 7 means she is not secreting non-volatile acid, causing acidosis; SLE gives tubulointerstitial inflammation, and the problem is distal tubule secretion of non-volatile acid (titratable acid and NH4+).

The four primary disturbances

Respiratory disturbances (pCO2)

• Too much CO2: hypoventilation, for example chronic bronchitis, or high inspired CO2, gives respiratory acidosis.
• Too little CO2: hyperventilation, for example anxiety, altitude or hypoxia, gives respiratory alkalosis.

Metabolic disturbances (HCO3-)

• Too much HCO3-, an acid deficit: excess bicarbonate from ingestion or renal overproduction, or loss of acid secretions (gastric contents), gives metabolic alkalosis.
• Too little HCO3-, an acid excess: loss of bicarbonate (gut), inadequate renal bicarbonate production, or addition/excessive production of organic acids, gives metabolic acidosis.

Self-test

1. State the normal ECF pH and range, the pH thresholds defining acidosis and alkalosis, and the values that are usually fatal.
2. List four consequences of disordered pH given in the lecture, including the CNS effects of alkalosis and of acidosis.
3. Explain what the Pitts 1953 dog experiment demonstrates, using the pH values obtained.
4. Distinguish the two day-to-day acid challenges the body faces, giving examples of each.
5. Describe how H+ generated from CO2 is buffered in transit in the blood, and what happens to that buffering in the lungs.
6. Predict the acid-base consequence of hypoventilation and of hyperventilation, explaining the direction the CO2 hydration reaction shifts in each.
7. List the sources of non-volatile acid and state the daily quantity produced.
8. Name the three main buffers of whole body buffering, and state what else contributes.
9. Explain why buffering alone is insufficient and why the kidney must be involved.
10. State the proportions of bicarbonate “reabsorption” occurring at each nephron segment, with the transporter involved at each.
11. Describe the steps by which a proximal tubule cell reabsorbs filtered bicarbonate.
12. What happens to bicarbonate handling if carbonic anhydrase is unavailable in the proximal tubule reabsorption pathway?
13. Distinguish the titratable acid mechanism from the ammonia mechanism for forming new bicarbonate, including the buffer and the nephron site of each.
14. Describe the handling of ammonium along the nephron from glutamine metabolism to luminal trapping.
15. Write out the Henderson-Hasselbalch equation and state which control system governs each of its two variables.
16. Distinguish proximal from distal renal tubular acidosis by defect, consequence and causes.
17. A patient has alkaline urine containing glucose, phosphate and amino acids, with pH 7.25, HCO3 14 mmol/L and PaCO2 33 mmHg. What is the diagnosis and why?
18. A patient with untreated SLE and diffuse lupus nephritis has HCO3 13 mmol/L and urine pH 7. Explain the mechanism of her acidosis.
19. List the causes of metabolic alkalosis and of metabolic acidosis given in the lecture.
20. Integrative: trace a single molecule of non-volatile acid HA from production to excretion, naming what is made, what is consumed, what is filtered and what the kidney contributes.

Answers

Reveal answers

1. ECF pH 7.4, range 7.36 to 7.45 (cells nearer pH 7). pH below 7.35 is acidosis, above 7.45 is alkalosis. Plasma pH below 7 or above 7.8 is usually fatal. Regulated as H+ within 36 to 44 nmol/L.
2. Enzyme activity (optimum pH), ion transport mechanisms, hormone function, and cellular metabolism/DNA synthesis/growth. CNS: alkalosis gives headache, drowsiness, tetany and convulsions; acidosis affects respiratory control and causes drowsiness progressing to coma.
3. Injecting a dog with 150 mL of 1 mol/L HCl dropped blood pH only from 7.44 to 7.14, whereas the same load in a similar volume of water gave 7.44 to 1.84. This shows buffering removes more than 99.99% of the acid load.
4. CO2 produced by cells, acting through $C{O}_2+H_{2}O\leftrightarrow H_{2}C{O}_3\leftrightarrow H^{+}+HC{O}_3^{-}$; and non-volatile acids from metabolism, such as phosphoric, sulfuric and lactic acid and ketone bodies.
5. In cells, CO2 + H2O forms H2CO3 then H+ + HCO3-; the H+ reacts with O2Hb- to give HHb + O2, so haemoglobin buffers the H+ and there is very little pH change. In the lungs, HHb + O2 gives H+ + O2Hb-, and the H+ recombines with HCO3- to form H2CO3 then CO2 + H2O, and the CO2 is removed.
6. Hypoventilation: CO2 is not removed and accumulates, the reaction shifts right, excess H+, respiratory acidosis. Hyperventilation: excess CO2 is removed, the reaction shifts left, decreased H+, respiratory alkalosis.
7. 50 to 80 mmol/day (NEAP): H2SO4 from sulfur-containing amino acids, H3PO4 from phosphoproteins and phospholipids, organic acids such as lactate and pyruvate from anaerobic exercise, and ketoacids from fat metabolism.
8. Bicarbonate, protein and phosphate, plus dilution and exchange with bone minerals.
9. Buffering only hides the H+; it still has to be got rid of. Buffered H+ reacts with HCO3- to give H2CO3 then CO2 + H2O, the CO2 is exhaled and the H+ ends up in water, but an HCO3- has been consumed and must be replaced by the kidney.
10. Proximal tubule 80% via NHE-3; thick ascending limb 10%; distal tubule/collecting duct 10% via intercalated cells using H+/K+ ATPase and H+ ATPase. Some intercalated cells instead secrete HCO3- in exchange for Cl-.
11. Apical NHE-3 secretes H+ into the lumen; luminal H+ plus filtered HCO3- forms H2CO3 via carbonic anhydrase; H2CO3 breaks down to H2O and CO2; CO2 diffuses into the cell; intracellular CO2 + H2O forms H2CO3 (carbonic anhydrase) then HCO3- + H+; H+ is recycled to the apical exchanger and HCO3- exits basolaterally to blood. The basolateral Na+/K+ ATPase (3Na+ out, 2K+ in) drives the exchange.
12. Carbonic anhydrase is required both to convert luminal H2CO3 to H2O and CO2 and to form intracellular H2CO3 from CO2 and H2O, so both the luminal disposal of filtered HCO3- and the intracellular generation of HCO3- for basolateral exit would fail, and filtered bicarbonate would not be reclaimed. [slide does not elaborate beyond showing carbonic anhydrase at both steps]
13. Titratable acid uses the urinary buffer HPO4²⁻ in the distal tubule: secreted H+ (ATP-driven transporter) plus filtered HPO4²⁻ forms H2PO4-, which is excreted. The ammonia mechanism uses glutamine converted to NH3 then NH4+, starting in the proximal tubule, passing through the medulla and acting in the collecting duct (glycoproteins). In both, intracellular CO2 + H2O via carbonic anhydrase gives the HCO3- returned to blood.
14. In the proximal-tubule cell, glutamine is metabolised to A⁼ and 2NH4+; A⁼ becomes 2HCO3- exiting to blood, and NH4+ enters the tubular fluid via a Na+/H+-type exchanger (in the collecting duct cell the source is glutamine to glutamate + NH3 via glutaminase). NH4+ is carried down the descending limb and recycled/concentrated through the loop of Henle into the medullary interstitium, feeding the collecting duct. There NH3 diffuses into tubular fluid and combines with secreted H+ to form NH4+, which is trapped in the lumen and excreted, while the associated HCO3- exits to blood.
15. $pH=6.1+\log ([HC{O}_3^{-}]/(0.03\times pC{O}_2))$. HCO3- is under renal control; pCO2 is under respiratory control.
16. Proximal RTA: defect in the mechanism for reabsorbing HCO3-, so HCO3- is lost and acidosis follows; causes are congenital, paraproteins (myeloma), PTH and drugs. Distal (“classic”) RTA: defect in non-volatile acid secretion mechanisms, so acid is not excreted and acidosis follows; causes are congenital, inflammation such as SLE, toxins such as amphotericin, hyperkalaemia and Conn’s.
17. Proximal RTA in Fanconi syndrome. Glucose, phosphate and amino acids in the urine localise the defect to the proximal tubule, and proximal tubule failure to reabsorb HCO3- causes bicarbonate loss and acidosis.
18. Urine pH 7 shows she is not secreting non-volatile acid, so acid is not excreted and acidosis develops. SLE causes tubulointerstitial inflammation, and the problem is distal tubule secretion of non-volatile acid (titratable acid and NH4+).
19. Metabolic alkalosis (too much HCO3-, acid deficit): excess bicarbonate from ingestion or renal overproduction, and loss of acid secretions (gastric contents). Metabolic acidosis (too little HCO3-, acid excess): loss of bicarbonate from the gut, inadequate renal bicarbonate production, and addition or excessive production of organic acids.
20. HA dissociates to A- + H+ and the H+ is buffered (acid made). The H+ reacts with HCO3- to give CO2 + H2O, so bicarbonate is consumed and the CO2 is exhaled. A- is filtered at the glomerulus. The kidney secretes H+ and generates new HCO3- to replace what was consumed. In the tubular fluid the filtered A- and secreted H+ recombine as HA, which is excreted.