Na + Transport 1 and 2 Linda Costanzo, Ph.D. OBJECTIVES: After studying this lecture, the student should understand: 1. The terminology applied to single nephron function, including the meaning of TF/P and the double ratio. 2. How to calculate fractional water reabsorption using TF/P inulin 3. The pattern of sodium reabsorption along the nephron. 4. Features of and transporters involved in sodium reabsorption in the early proximal tubule, late proximal tubule, thick ascending limb, early distal tubule, and late distal tubule and collecting ducts. I. TERMS TO DESCRIBE SINGLE NEPHRON FUNCTION Up to this point, we have discussed primarily whole kidney function (e.g., GFR, urine, clearance, excretion). Now we will turn our attention to the nephron, which is the functional unit of the kidney. There is a specific vocabulary of the nephron, with terms analogous to that of the whole kidney (e.g., tubular fluid is analogous to urine). [TF] x is the concentration of substance X in tubular fluid. (Tubular fluid is the fluid inside the nephron...also called luminal fluid.) [P] x is the concentration of substance X in plasma and is considered to be constant. SNGFR is the single nephron glomerular filtration rate.
[TF/P] x is the concentration of substance X in tubular fluid relative to the concentration in plasma. There are three possibilities for the value of this ratio, which are explained as follows: [TF/P] X = 1.0. X has not been reabsorbed or secreted (all freely filtered substances in Bowman's space), or X is reabsorbed in proportion to water (e.g., Na in proximal tubule). For example, [TF/P] X =1.0 for all freely filtered substances in Bowman s space (no reabsorption or secretion has taken place yet). For another example, [TF/P] Na = 1.0 throughout the proximal tubule because Na + is reabsorbed in exact proportion to water. [TF/P] X < 1.0. X is reabsorbed more than water, causing the concentration of X in tubular fluid to fall below that in plasma. For example, [TF/P] glucose starts at 1.0 in Bowman s space, but then falls below 1.0 along the proximal tubule as glucose is reabsorbed more than water. [TF/P] X > 1.0. X is reabsorbed less than water or X is secreted. If X is reabsorbed less than water (or if X is secreted into tubular fluid), the concentration of X in tubular fluid rises above that in plasma. For example, [TF/P] urea is > 1.0 in cortical collecting ducts in the presence of ADH because water is reabsorbed and urea is not. For another example, [TF/P] K is > 1.0 in cortical collecting ducts because this part of the nephron secretes K +.
[TF/P] inulin is the concentration of inulin in tubular fluid relative to the concentration of inulin in plasma. This specific [TF/P] X ratio is used to measure water reabsorption since inulin, once filtered, is "inert" (i.e., is neither reabsorbed or secreted). Thus, the amount of inulin in tubular fluid is constant along the nephron (because inulin is not reabsorbed or secreted) but the concentration of inulin in tubular fluid is determined by how much water remains; as water is reabsorbed, the tubular fluid concentration of inulin increases. For example, if 50% of the filtered water has been reabsorbed, then the tubular fluid inulin concentration doubles and the [TF/P] inulin = 2.0. (Don't forget that the "P" in TF/P is always assumed to be constant.) Calculate water reabsorption with this ratio as follows: Fraction of filtered water reabsorbed= 1-1 [TF/P] inulin For example, if tubular fluid is sampled at the end of the proximal tubule, and the [TF/P] in ratio is measured as 3.0, what fraction of the filtered water has been reabsorbed up to that point? What fraction of the filtered water remains in the lumen of the nephron? Fraction of filtered water reabsorbed = 1-1 [TF/P] in = 1-1/3 = 2/3, or 66.7% reabsorbed If 2/3 of the filtered water has been reabsorbed, then 1/3 of the filtered water remains in the lumen of the neprhon. [TF/P] x [TF/P] inulin, or the "double ratio" is the fraction of the filtered load of a substance remaining in the nephron at any point. If the "double ratio" is 0.3, then 30% of the filtered load of the substance remains in tubular fluid, and 70% of the filtered load must have been reabsorbed. If you re wondering why the double ratio corresponds to fraction of filtered load of a substance remaining in the nephron, here s the derivation (in italics, just FYI...):
% of filtered load remaining = excretion rate of x at any point in nephron divided by filtered load of x in nephron Excretion rate at any point in nephron = [TF] x x V SNGFR = single nephron GFR= [TF] inulin [P] inulin x V Filtered load of X = SNGFR x [P] x = [TF] inulin x V/[P] inulin x [P] x Substituting, and cancelling V: % of filtered load remaining = [TF/P] x /[TF/P] inulin We will use [TF/P] X and the double ratio together to describe how substances are handled in the nephron. For example, if, at the end of the proximal tubule, the double ratio for Na + is 0.33 and [TF/P] Na is 1.0, we would say that 33% of the filtered Na + remains in the nephron at the end of the proximal tubule, that 67% of the filtered Na + was reabsorbed by the proximal tubule, and that this Na + reabsorption must have been in exact proportion to water reabsorption (since [TF/P] Na was 1.0). II. OVERALL NA + BALANCE Na + is the major ECF cation and, with accompanying anions Cl - and HCO 3 -, constitutes the major ECF solute. As we have already discussed, the amount of Na + in ECF determines ECF volume and therefore also determines blood volume and blood pressure. Thus, regulation of Na + balance is the most important function of the kidneys. On an average daily diet of 150 meq of Na + ingested, the kidneys must excrete 150 meq of Na + to keep us in Na + balance (neglecting small non-renal losses such as via sweat). If the kidneys excrete less Na + than is ingested, then we are in positive Na + balance; if the kidneys excrete more Na + than is ingested, we are in negative Na + balance.
Figure 1. Na + handling in the nephron. Arrows show locations of Na + reabsorption; numbers are percentages of the filtered load reabsorbed or excreted. Na + is reabsorbed along the nephron as follows: 67% of the filtered load in the proximal tubule, 25% in the thick ascending limb of Henle, 5% in the early distal tubule, and 3% in late distal tubule and collecting duct. Cumulatively, this is more than 99% of the filtered load reabsorbed, leaving less than 1% of the filtered load to be excreted. (For a person in Na + balance, 1% of the filtered load excreted corresponds to the daily Na + excretion that would equal daily Na + ingestion.) III. PROXIMAL CONVOLUTED TUBULE: EARLY AND LATE PROXIMAL The entire proximal convoluted tubule reabsorbs 67% or 2/3 of the filtered Na +. A major feature of proximal Na + reabsorption (and total solute reabsorption as well) is that it is linked directly to water reabsorption. Thus, Na + (and solute) reabsorption is proportional to water reabsorption and we call the process isosmotic. The basis for isosmotic reabsorption will be explained later in the lecture. Proximal tubule is divided between an "early" part (first half, nearest the glomerulus) and "late" part (second half). The cellular mechanisms for Na + reabsorption are different in the two parts, so they will be discussed separately.
A. Early proximal tubule Figure 2. Cellular mechanisms of Na + reabsorption in the early proximal tubule. The transepithelial potential difference is the difference between the potential in the lumen and the potential in blood, -4 mv. ATP, Adenosine triphosphate. Early proximal tubule has the following features: Na + -glucose, Na + -amino acid, and Na + -phosphate cotransporters in the luminal membrane Na + -H + exchange in the luminal membrane (linked to filtered HCO 3 - reabsorption, which will be covered in the acid-base portion of the course) Preferential reabsorption of HCO 3 - over Cl - (as the anion accompanying Na + reabsorption) Na + - phosphate cotransport is inhibited by parathyroid hormone (PTH) and responsible for the phosphaturic effect of PTH. Lumen-negative transepithelial potential difference due to Na + - glucose cotransport
Always isosmotic reabsorption [TF/P] Na and [TF/P] osm = 1.0 B. Late proximal tubule Figure 3. Cellular mechanisms of Na + reabsorption in the late proximal tubule. The transepithelial potential difference is +4 mv. ATP, Adenosine triphosphate. Late proximal tubule has the following features: High luminal Cl - concentration (created by preferential reabsorption of HCO 3 - in early proximal) Cl - reabsorption by Cl - -formate exchange in luminal membrane and by Cl - diffusion between cells (down Cl - concentration gradient) Lumen positive transepithelial potential difference created by Cl - diffusion Always isosmotic reabsorption [TF/P] Na and [TF/P] osm = 1.0 IV. PROXIMAL TUBULE: ISOSMOTIC REABSORPTION, GLOMERULOTUBULAR BALANCE Solute (mainly Na +, HCO 3 -, Cl -, glucose and amino acids) and water reabsorption are always proportional to each other in proximal tubule. They are linked mechanistically, so the reabsorption process is isosmotic. A consequence of this
proportional, isosmotic process is that, along the entire proximal tubule, [TF/P] Na = 1.0 and [TF/P] osm = 1.0. In other words, the concentration of Na + and total osmolarity do not change in proximal tubule fluid, even though lots of sodium and total solute are reabsorbed. The mechanism of isosmotic reabsorption is explained in the figure below. Figure 4. Mechanism of isosmotic reabsorption in the proximal tubule. Dashed arrows show the pathways for reabsorption; circled numbers correspond to the text. π c, Peritubular capillary colloid osmotic pressure. Solute crosses the luminal membrane (as described for the early and late proximal tubule). The luminal membrane is permeable to water, so water follows solute into the cell. Na + is pumped out across the lateral membranes into the lateral intercellular space by the Na + -K + pump; again, water follows. In the lateral space, an isosmotic fluid accumulates. From that point on, reabsorption of this fluid is driven by Starling forces across the peritubular capillary. The major Starling force driving proximal tubule reabsorption is the high oncotic pressure, π c, of peritubular capillary blood (recall how this high π c is created!). Glomerulotubular balance is a regulatory feature of the proximal tubule. It says that glomerular filtration of Na + (and solute and water) are balanced by reabsorption (i.e. by the tubule), resulting in constant fractional reabsorption of 67%. This balance or constancy ensures that ECF Na + content, and therefore ECF volume, will be maintained. The physiologic parameter that maintains glomerulotubular balance is the oncotic pressure of peritubular capillary blood (π c ). Thus, if for some reason GFR were to spontaneously increase, there would be increased filtration of fluid out of glomerular capillaries, increased π c in
peritubular capillary blood, and increased driving force for proximal reabsorption, i.e., constant fractional reabsorption. Normally, glomerulotubular balance prevails and reabsorption is 67% of the filtered load. However, changes in ECF volume (volume contraction and volume expansion) can "override" and change the percentage reabsorbed. In ECF volume expansion, fractional reabsorption is decreased (excretion is increased) and in ECF volume contraction, fractional reabsorption is increased (excretion is decreased). Figure 5. Effects of ECF volume expansion (A) and ECF volume contraction (B) on isosmotic fluid reabsorption in the proximal tubule. Changes in Starling forces in the peritubular capillary blood are responsible for the effects. π c, Peritubular capillary colloid osmotic pressure; Pc, peritubular capillary hydrostatic pressure. V. THICK ASCENDING LIMB OF HENLE'S LOOP Thick ascending limb of Henle's loop reabsorbs 25% of the filtered Na +. In contrast to the proximal tubule, which always reabsorbs isosmotically, thick ascending limb reabsorbs solute without water. Because it is impermeable to water, it also is called the diluting segment (if solute is reabsorbed and water is left behind, the tubular fluid becomes dilute). Therefore, both [TF/P] Na and [TF/P] osm are <1.0 in the thick ascending limb.
Figure 6. Cellular mechanism of Na + reabsorption in the thick ascending limb of the loop of Henle. The transepithelial potential difference is +7 mv. ATP, Adenosine triphosphate. Thick ascending limb has the following features: Na + -K + -2Cl - cotransporter in luminal membrane Cells are impermeable to water and tubular fluid is diluted as Na + and solute are reabsorbed Also called diluting segment Lumen positive transepithelial potential difference due to action of Na + -K + -2Cl - cotransporter. This feature is puzzling at first, because it seems that the cotransporter would be electroneutral. However, note on the figure that some K + diffuses from cell back into lumen, and consequently, net negative charge enters cell, leaving the lumen with a positive charge. Furosemide and other loop diuretics inhibit the Na + -K + -2Cl - cotransporter and thereby inhibit Na + reabsorption, the basis for their diuretic action [TF/P] Na and [TF/P] osm < 1.0 Important for countercurrent multiplication (another lecture) Ca 2+ and Mg 2+ are reabsorbed between the cells, driven by the lumenpositive potential. Loop diuretics abolish the lumen-positive and inhibit Ca 2+ reabsorption VI. DISTAL TUBULE AND COLLECTING DUCTS Together, the distal tubule and collecting ducts reabsorb 8% of the filtered Na +. In terms of cellular mechanisms, these segments are divided into early distal tubule and late distal tubule/collecting duct.
A. Early distal tubule Early distal tubule reabsorbs about 5% of the filtered Na +. Like thick ascending limb, the cells are impermeable to water and it is called the cortical diluting segment. As a result both [TF/P] Na and [TF/P] osm are < 1.0. Figure 7. Cellular mechanism of Na + reabsorption in the early distal tubule. The transepithelial potential difference is -10 mv. ATP, Adenosine triphosphate. Early distal tubule has the following features: Na + -Cl - cotransporter in luminal membrane Cells are impermeable to water and tubular fluid is diluted as solute is reabsorbed Called cortical diluting segment Thiazide diuretics (chlorothiazide; hydrochlorothiazide) inhibit the Na + -Cl - cotransporter, which is the basis for their diuretic action Ca 2+ reabsorption is active and stimulated by parathyroid hormone (PTH) and thiazide diuretics B. Late distal tubule/collecting duct Together, the late distal tubule and collecting duct reabsorb 3% of the filtered Na +. There are two cell types interspersed along these segments,
the principal cells and the intercalated cells. For completeness, both cell types will be described here, although it is the principal cells that reabsorb Na +. These segments are notable in that they are the site of action of important hormones: aldosterone (regulates Na + reabsorption, K + secretion, and H + secretion) and ADH (regulates water reabsorption) 1. Principal cells Figure 8. Cellular mechanism of Na + reabsorption in the principal cells of the late distal tubule and collecting duct. The transepithelial potential difference is -50 mv. ATP, Adenosine triphosphate. Principal cells have the following features: Na + channels in luminal membrane (called EnaC, for epithelial Na + channel) Aldosterone induces these Na + channels and thereby increases Na + reabsorption Na + reabsorption is fine-tuned in the principal cells by the action of aldosterone Water reabsorption is variable, via aquaporin2 (AQP2) in the luminal membrane that are regulated by ADH K + secretion via K + channels (discussed in next lecture) K + -sparing diuretics such as spironolactone and amiloride inhibit the action of aldosterone, block the Na + channels, decrease which is the basis for their diuretic action 2. Intercalated cells have the following features
Reabsorption of K + via the H + - K + ATPase (under conditions of low K + diet) Secretion of H + via the H + ATPase, which is increased by aldosterone Figure 9 VII. PRACTICE QUESTIONS 1. If [TF/P] Na+ /[TF/P] inulin is 0.33 at the end of the proximal convoluted tubule, what fraction of the filtered load of Na + has been reabsorbed?
2. If [TF/P] inulin = 3.0, what percentage of the filtered H 2 O has been reabsorbed? 3. How will an increase in filtration fraction alter isosmotic reabsorption in the proximal tubule and why? 4. How do changes in ECF volume alter isosmotic reabsorption in the proximal tubule? 5. How does the Na + -K + -2Cl - cotransporter produce a lumen-positive transepithelial potential difference in the thick ascending limb? 6. How does reabsorption of solute (NaCl) in the thick ascending limb cause dilution of tubular fluid? What is [TF/P] osm of fluid leaving the thick ascending limb? 7. What are major differences between the transport functions of the early distal tubule and the late distal tubule/collecting duct? 8. Which of the following exhibits increased Na + reabsorption in the late distal tubule and collecting ducts? Person with syndrome of inappropriate ADH Person with aldosterone-secreting tumor Person taking a thiazide diuretic Person taking spironolactone ANSWERS 1. 67% 2. 67% 3. Increases it (increased filtration from glomerular capillaries increases π c ) 4. Increased ECF volume decreases isosmotic reabsorption; decreased ECF volume increases isosmotic reabsorption 5. See notes 6. In thick ascending limb, more solute is reabsorbed than water, diluting the tubular fluid. [TF/P] osm <1.0 7. Early distal, Na + -Cl - cotransport, no H 2 O; late distal/collecting duct, Na + channels, H 2 O depends on ADH, aldosterone stimulates Na + reabsorption and K + secretion 8. Person with syndrome of inappropriate ADH no Person with aldosterone-secreting tumor yes Person taking a thiazide diuretic no Person taking spironolactone no