Obligatory reabsorption and secretion in the proximal convoluted tubule

Question 1

Define blood urea nitrogen. What is the effect of GFR increase on BUN ? Identify a treatment option for increased BUN.

Answer 1

Blood urea nitrogen (BUN) = Measures nitrogen in the blood as urea (waste product of amino acid catabolism). If GFR decreases (as in renal disease) BUN increases. Increased BUN can be treated with a reduced protein diet (decreased amino acids). Normal range 7-18 mg/dL or 1.2 – 3 mM.

Question 2

Define oncotic P.

Answer 2

Osmotic pressure for large molecules (e.g. proteins). These attract H2O, but number of particles per kg is low.

Question 3

Define osmolarity.

Answer 3

Number of solute particles per litre (mOsm/L)

Question 4

Define osmolality.

Answer 4

Number of solute particles per kg solute (mOsm/kg H2O). NOT temperature dependent and therefore correct term for biology.

Question 5

Define osmosis.

Answer 5

Movement of H2O across cell membranes. Driving force is osmotic
pressure difference.

Question 6

Define osmotic P.

Answer 6

Determined by number of solute particles in solution. Not affected by size of particle, charge or mass. Generally expressed in terms of osmolarity or osmolality.

Question 7

What is the formula for specific gravity ? Based on this, what is the specific gravity of distilled water ?

Answer 7

Weight of a set volume of solution / Weight of same volume of pure water

Distilled water is therefore 1.000

Question 8

What is the specific gravity of normal urine ?

Answer 8

1.007 due to presence of solutes.

Question 9

What is the osmolality of 1 mmol/L solution of CaCl2 ? of glucose ? of NaCl ?

Answer 9

mmol/L solution of CaCl2 has an osmolality of 3 mOsm/kg H2O, since CaCl2 is comprised of Ca2+ and 2Cl-.

1 mmol/L solution of glucose has osmolality of 1 mOsm/kg H2O.

1 mmol/L solution of NaCl has an osmolality of 2 mOsm/kg H2O, since NaCl is comprised of Na+ and Cl-.

Question 10

What is the main site for obligatory reabsorption ?

Answer 10

Proximal Tubule

Question 11

Is obligatory reabsorption regulated ? What is the one factor that does regulate the amount of obligatory reabsorption that takes place ?

Answer 11

No

Only thing that
regulates reabsorption of solutes is the amount of solutes filtered in first place from glomerulus (i.e. no intrinsic factor other than availability of ions/metabolites)

Question 12

Identify the main solutes/metabolites which are reabsorbed in the PT.

Answer 12

Glucose
AAs
Magnesium
Calcium
Sodium
Potassium
H20
Phosphate
Urea

Question 13

What are the main functions of the PT ?

Answer 13

1) Most of the recovery of water, glucose, Na+ etc.
2) Actively secretes a number of compounds for excretion with urine (e.g. antibiotics, waste products of metabolism), and metabolises some of the amino acids

Question 14

Are proteins filtered by the glomerulus ? Why or why not ?

Answer 14

Almost all proteins not filtered by glomerulus because of negatively charged BM (so stay in blood) but small amount of proteins do come through.

There are proteases on PT cell surface that break those proteins down into AAs, which then get reabsorbed.
There are also some proteins that come through that are smaller, and are taken up by
endocytosis (into cell).

Question 15

Identify the possible pathways taken by molecules and ions across the tubule epithelium in reabsorption.

Answer 15

1. Transcellular route (through the apical surface into the cell body, into the interstitial fluid, then into bloodstream)
2. Paracellular route (through the leaky tight junctions between cell bodies, into the bloodstream)

Question 16

Identify the main forces involved in the obligatory reabsorption from the PT.

Answer 16

1. Ion gradients across the basolateral membrane– active transport 3 Na out 2K in
2. This sets up an electrochemical gradient (pumping more positive charge out than in) of about -3mV (tubule lumen negative compared to interstitial fluid; causes paracellular efflux of cations)
3. Osmotic gradient set up by pumping Na out of the cell in to the interstitial space
4. Water moving along the paracellular path due to osmotic pressure drags solutes along with it (especially negative electrolytes, because transluminal negative potential in PT means cations tend to fight solvent drag). Known as solvent drag
5. Chemical concentration of solutes left behind when water leaves the tubule facilitates a chemical gradient (can then use transcellular route to diffuse down their concentration gradient)

Question 17

Is the lumen negative relative to the interstitial fluid along the entirety of the tubules ?

Answer 17

No, lumen negative in PT and distal tubule and collecting ducts, BUT lumen positive relative to interstitial fluid in loop of Henle and TAL.

Question 18

Where does a cotransporter with sodium move substances ?

Answer 18

Co-transport with sodium moves substances into the cell

Question 19

Where does an exchanger with sodium move substances ?

Answer 19

Counter transport (exchange) with sodium moves them out

Question 20

Give an example of secondary active transport.

Answer 20

Sodium pump (actuve transport) sets up Na+ concentration gradient. This gradient is then used by cell to transport other substances.

Question 21

Identify the main reabsorption pathways in the PT (i.e. which molecules in/out, using which transport systems etc.).

Answer 21

APICAL SURFACE (out means into tubular fluid, in means into cell)

1) Na+ H+ exchanger (1 Na+ in, 1 H+ out), using Na+ gradient. In the tubular fluid, H+ combine with filtered bicarbonate to form carbonic acid. Extracellular carbonic anhydrase then catalyses dissociation of carbonic acid into water and CO2, still in tubular fluid. CO2 diffuses back into cell (freely permeable to lipid membrane) whereas H2O enters cell through aquaporins. Intracellular carbonic anhydrase converts H2O and CO2 back into carbonic acid, which dissociates into H+ and bicarbonate. H+ is used in the Na+ H+ exchanger (into tubule lumen), whilst the the bicarbonate is excreted/secreted from basolateral membrane.
2) Ion channels for divalent cations (e.g. Ca++ channel)
3) Na+ glucose/AA cotransport (uses Na+ gradient to transport metabolites into cell)

BASOLATERAL SURFACE (out means into isf then blood, in means into the cell)
1) Na+ K+ ATPase (3 Na+ out, 2K+ in)

2) HCO3- Cl- exchanger (HCO3- out, Cl- in), or HCO3- Na+ cotransport (HCO3- out, Na+ out) to secrete HCO3- obtained from apical surface.
3) K+ Cl- cotransport (both out, uses K+ gradient to reasborb Cl-)
Overall, bicarbonate, chloride and potassium all leave down their concentration gradients.

4) Na+ Ca++ exchanger (Na+ in, Ca++ out, uses Na+ gradient to reabsorb Ca++ against its concentration gradient)
5) Glucose/AA transport (out of cell)

ALSO
Increased concentrations of Cl- and K+ in isf due to secondary active transport (K+ diffuses down concentration gradient into isf, Cl- follows). Because of increase in osmolarity, water will follow through paracellular route, into isf. Solutes dissolved in this water (e.g. Cl-) will follow (not cations, because because transluminal negative potential in PT means cations tend to fight solvent drag, so net flow of anions into isf whereas in other parts of tubule, where lumen is positive, net intake of cations into isf) through solvent drag, through paracellular route

ALSO
Entry of other solutes: coupled to Na+ entry on apical membrane; facilitated diffusion on the basolateral membrane. Other solutes have their own channels through which they diffuse, this is facilitated by the process of concentration (water loss) and Na + entry.

ALSO
Organic ions transporters on both apical and basolateral membranes to reabsorb wanted ones (e.g. nucleotides) from tubular fluid, or to secrete ones (e.g. antibiotics, absorbed pesticides) from blood

Question 22

Why is it possible to segregate between proteins (i.e. transporters) of basolateral membrane and those of the apical membrane ?

Answer 22

Because of tight junctions (proteins cannot diffuse across these)

Question 23

Define Tmax, in reabsorption. What are the units for it ? Are there any circumstances in which Tmax can be exceeded ?

Answer 23

Tmax is “the point at which increases in concentration of a substance do not result in an increase in movement of a substance across a cell membrane” – transport maximum.
Measured in mg/min or mmol/min (rate).

If blood conc. is high, Tmax can be exceeded (e.g. high blood concentration of glucose in diabetes mellitus: normally, almost 100% of
glucose can be reabsorbed by PT. In diabetes, blood glucose higher, exceeds
Tmax of PT for glucose, so glucose appears in urine)

Question 24

Graph amount of glucose filtered versus plasma glucose concentration.

Answer 24

Refer to slide 3 on page 5 of lecture.

Normally, plasma glucose concentration should be 100-120 mg/100 mL. Up to 120 mg/100 mL, that amount can be reabsorbed. Once above 180 (range for diabetes), Tmax for glucose is exceeded, so more filter but can only reabsorb this maximum right here. Hence, concentration of plasma glucose increases even more. Amount reabsorbed is saturated, so amount excreted increases.

Question 25

State the formula to obtain the amount filtered.

Answer 25

Amount filtered (mg/min)= plasma conc. (mg/ml) x GFR (ml/min)

Question 26

Define treshold in the context of reabsorption. What happens when this threshold is exceeded ?

Answer 26

• Threshold is the point at which the amount filtered is equivalent to Tmax
• Above threshold, substance appears in urine (when normally 100% reabsorbed), so above threshold, conc. of glucose in urine is proportional to plasma concentration (because saturated re-uptake system, so if increase amount in blood, comes out in urine)

Question 27

What transporter is mainly responsible for glucose reabsorption ?

Answer 27

Na-glucose transporter can reclaim ~375mg/ min glucose from blood

Question 28

Which factors may cause an increase in Tmax ?

Answer 28

If plasma conc. or GFR increase (i.e. there is more than normal limits in the tubule), Tmax may be exceeded as reabsorption process cannot cope

Question 29

Define PAH in the context of reabsorption.

Answer 29

Para-aminohippuric acid, substance which is actively secreted. When in blood, filtered out into filtrate and PT secretes it. Helps to work out blood flow to kidney (e.g. if looking at stenosis of arterioles going to kidney)

Question 30

Graph plasma concentration of PAH versus amount of PAH filtered.

Answer 30

Refer to slide 2 on page 6 of lecture.

Get linear relationship between amount in plasma and amount filtered.

At very low concentrations, amount filtered is tiny compared to amount secreted. Out of your 600 mL of plasma filtering through kidneys, 125 mL is filtered. Remaining blood goes to
vasa recta, then next to PT. PT takes PAH out of blood and actively secretes it. Can normally clear about 95% of PAH by this mechanism.

Secreted PAH is therefore higher than filtered PAH because you get active secretion. But you do get Tmax once saturate secretory mechanisms, so start seeing PAH is urine (so amount excreted start increasing even more, because coming through filtered part rather than secreted part).

Question 31

What is the relationship between amount filtered, amount present in plasma, amount appearing in urine, and amount secreted ?

Answer 31

♦ Amount filtered is proportional to the amount present in the plasma
♦ Amount appearing in urine (excreted) = amount filtered + amount secreted (from blood to tubule)
♦ Once max secretion level is reached, secretion line levels off so amount excreted is based on concentration in the plasma

Question 32

Identify the main molecules involved in obligatory reabsorption in the PT, and state the proportion of each that is reabsorbed.

Answer 32

• 100%: glucose, amino acids, lactate
• 90%: bicarbonate
• 65%: water and sodium
• 55%: potassium
• 50%: chloride

Question 33

Where does control of reabsorption/secretion (i.e. fine control) occur ?

Answer 33

In the distal part of the tubule.

Question 34

Describe the movement of the following in reabsorption:

• Urea
• Lipid-soluble substances
• Phosphate
• Protein

Answer 34

• Urea: mainly through urea transporters (but also some simple diffusion) reabsorbs 50-60% (rest lost)
• Lipid-soluble substances: simple diffusion through epithelial cells
• Phosphate: sodium-linked transport
• Protein: small amount digested to amino acids by proteases, within the tubule cells then reabsorbed (also some proteins taken up by receptor mediated endocytosis)

Question 35

Describe the effect of parathyroid hormone on reabsorption. Where does this act ?

Answer 35

Activity of carriers changed by parathyroid hormone:

• Stimulates re-uptake mechanism for Calcium (retain it)
• Inhibiting re-uptake of Phosphate

Mainly acts in DT rather than PT (fine control, between reabsorption or keeping it in urine)

Question 36

Define clearance of a substance, in the context of reabsorption.

Answer 36

The clearance of any substance excreted by the kidney is the volume of plasma which is cleared of the substance per unit time (ml/min). Theoretical V, since in reality no substance is completely cleared during its passage through the kidney (even PAH, which is actively secreted, since 5% of it is still reabsorbed)

Question 37

Identify the renal processes which determine and modify the composition of urine. How can we calculate amount excreted based on the values of the aforementioned processes ?

Answer 37

A) Glomerular filtration (normally stays the same, 120-125 mL/min)
B) Tubular reabsorption
C) Tubular secretion

Amount excreted in urine = A - B + C

Question 38

What are the possible ranges of clearance ?

Answer 38

Range is from zero (substance is filtered and then reabsorbed – glucose; or is never filtered – protein) to the equivalent of renal plasma flow (where all of the substance filtered by the kidney ends up in urine, such as PAH)

Question 39

How much blood flow does the kidney get ?

Answer 39

1100 mL/min

Question 40

Define renal plasma flow.

Answer 40

Amount of fluid entering the kidney that is potentially filterable (600ml/min)

Question 41

Define and state the formula for Filtration Fraction.

Answer 41

Filtration Fraction (F.F.) = fraction of the plasma entering the kidney that filters into the lumen of the renal tubules

FF = GFR/RPF = 125/600 = 20%

i.e. of 600 mL/min coming into kidney, only 125 mL filtered (20%).

Question 42

State the Fick principle.

Answer 42

Input Renal Artery = Output Renal Vein + Ureter

Plasma concentration x in renal artery (Pax) x Renal plasma flow in renal artery (RPFa) = (Plasma concentration x in renal vein (Pvx) x renal plasma flow in renal vein (RPFv)) + (Urine concentration of x (Ux) x Urine flow rate (V))

Question 43

How do we calculate renal clearance ?

Answer 43

To equate the urinary excretion rate of x to its renal arterial plasma concentration (Pa) it is necessary to determine the rate at which x is removed from the plasma – the clearance C x.

Pax x Cx = Ux x V
Cx= (Ux x V)/Pax

Where V = urine flow rate (ml/min) and U = urine concentration of x (mg/ml)

Question 44

What are the requirements for a substance to be a good measure of GFR ? Identify a substance used to measure GFR, and state any issues associated with its use.

Answer 44

To measure GFR the substance must be:
•  Freely filtered at the glomerulus 
•  Neither secreted or reabsorbed 
•  Not metabolised
•  Not toxic

Inulin – ideal substance but is a plant sugar and needs to be infused to establish constant plasma concentrations

Question 45

How can we calculate GFR using inulin ?

Answer 45

Amount filtered = Amount excreted (since inulin not secreted or reabsorbed)

Pin x GFR = Uin x V

GFR = (Uin x V)/ Pin

Question 46

Identify any formulas which may help us determine if a substance is secreted, or reabsorbed.

Answer 46

May use clearance rations to determine the renal transport mechanism (if net reabsorption, net secretion, neither etc.)

[(Uphos x V) ∕ P phos] / [(Uin x V) ∕ P in]

(U phos/P phos) / (U in/P in)

where phos= phosphate and in= inulin

♦ Clearance ratio - equal to 1, nether secreted nor reabsorbed
♦ Clearance ratio - greater than 1, substance secreted
♦ Clearance ratio - less than 1, substance reabsorbed

Question 47

Which of the following are possible renal routes ?

1) Filtered and secreted
2) Filtered and partially reabsorbed
3) Filtered and all reabsorbed

Answer 47

All three are possible:

1) Filtered and secreted
2) Filtered and partially reabsorbed
3) Filtered and all reabsorbed