Proximal Tubule Tm Mechanisms PDF

WSUSOM Medical Physiology Rossi-Renal Physiology Page 1 of 15 Proximal Tubule Tm Mechanisms Proximal Tubule Tm Mechanisms Learning Objectives: 1. Proximal tubule A. Compare relative quantitative transport that occurs in the proximal tubule...

WSUSOM Medical Physiology Rossi-Renal Physiology Page 1 of 15 Proximal Tubule Tm Mechanisms Proximal Tubule Tm Mechanisms Learning Objectives: 1. Proximal tubule A. Compare relative quantitative transport that occurs in the proximal tubule with the rest of the nephron B. Recognize that reabsorption in the proximal tubule is iso-osmotic and the underlying mechanisms responsible C. Distinguish between Tm and non-Tm reabsorptive processes in the proximal tubule and the substances handled by each of these mechanisms D. Identify the concept and mechanisms involved in glomerular tubular balance E. Distinguish between different types of transport a. primary and secondary active transport, facilitated diffusion, passive transport and endocytosis b. co-transport, counter-transport (or exchanger) or channel c. water transport via osmotic forces F. Tm reabsorption a. Articulate the concepts of threshold and Tm and be able to calculate them b. Recognize the influence glomerular filtration rate exerts on threshold c. Understand the concept of splay d. Distinguish the major categories of solute that are reabsorbed via Tm mechanisms and the unique characteristics of the major categories, including the transporters involved G. Tm secretion 1) Recognize that the concept of threshold does not apply to secretory processes 2) Identify the major cations and anions that are secreted by the proximal tubule and the general transport mechanisms involved WSUSOM Medical Physiology Rossi-Renal Physiology Page 2 of 15 Proximal Tubule Tm Mechanisms This lecture focuses on the Rx and Sx components of the excretion equation, that is, the reabsorptive and secretory functions of the tubules, respectively. Excretion Ux * V = (GFR * Ax) - Rx + Sx Clearance Cx = Ux* V = GFR _ Rx + Sx Ax Ax Ax Normal GFR is 125 ml/min. In one day 180 liters of fluid are filtered into Bowman’s space each day: 125 ml/min * 1,440 min/day = 180,000 ml/day = 180 L/day Since typically, humans excrete 1 - 2 liters of urine per day, the tubules must - reabsorb 178 - 179 liters of fluid each day - reclaim essential solutes that are filtered at the glomerulus - eliminate solutes (including drugs, toxins, etc.) that are not filtered in sufficient quantity Theoretical Problem: If a 70 kg “ideal” male with a GFR of 125 ml/min does not reabsorb any fluid, how long would it take for him to excrete his total body water (as if this were possible)? Total body water (TBW) = 0.6 * 70 kg = 42 L 42 L ÷ 0.125 L/min = 336 min = 5.6 hr More scary, if [Na] in ECV = 150 mM, then Na content of ECV = 2,100 mmol. At a GFR of 125 ml/min, 18.75 mmol Na are filtered/min. It would take only 112 minutes (< 2 hrs) to lose all ECV Na. Note that 80% of the ATP used by the kidney (2 kg/d) is spent to reabsorb Na. Recall: ECV = 1/3 * TBW; ICV = 2/3 * TBW. PV = 1/4 * ECV; ISV = 3/4 * ECV (Note that I use 150 mM for extracellular Na concentration. This is because strictly speaking I will be discussing Na dissolved in plasma water and it is the Na dissolved in plasma water that the cells respond to. Those of you who have seen lab results or worked in hospitals know that “normal” Na is 140 mM. This is the Na concentration in plasma or serum, which is normally only 93% water. When the lab measures the [Na] it calculates it as if all the serum were water. This is actually not true. This is why the fluids given to patients “normal saline” is 0.9% NaCl or ~ 154 mM instead of 140 mM!) WSUSOM Medical Physiology Rossi-Renal Physiology Page 3 of 15 Proximal Tubule Tm Mechanisms Just a reminder of where we are going…following the filtered fluid and its contents into the proximal tubule… Proximal Tubule Proximal tubule ~60-80% of filtered solutes and water reabsorbed isotonically as solutes reabsorbed, water reabsorbed passively by osmosis solutes secreted, but negligible QUANTITATIVELY overall Proximal tubular mechanisms responsible for reclaiming and eliminating solute/fluid: 1. Reabsorption: Tm and non-Tm mechanisms 2. Secretion - Tm mechanisms 3. H2O reabsorption “Tm” means “tubular maximum” or “transport maximum” Although conceptually the Tm may be considered analogous to the Bmax of an enzyme, the Tm is not for a single transporter but for the proximal tubule as whole. Several transporters may contribute to the Tm. For example, for glucose which we will discuss below there are two transporters, SGLT1 and SGLT2 but ONE and only ONE Tm for the tubule. WSUSOM Medical Physiology Rossi-Renal Physiology Page 4 of 15 Proximal Tubule Tm Mechanisms Glomerulo-tubular Balance Spontaneous changes in GFR lead to parallel changes in proximal tubule reabsorption ­ or ¯ in GFR leads to proportional ­ or ¯ in proximal tubule reabsorption, respectively Mechanisms - Parallel change in pc in peritubular capillaries → parallel change in solute and H2O reabsorption - Change in filtered load of glucose ® parallel change in Na reabsorption Glomerulo-tubular balance: When body Na+ balance is normal, an increase in GFR results in a higher filtered load of Na (GFR x ANa). If this is not accompanied by a proportional increase in proximal tubular reabsorption of Na+ and H2O, too much Na+ and water will be delivered downstream (more than the distal segments can handle) and result in more Na+ and water being excreted. Thus, a constant fraction (»80%) of the filtered load of and H2O is reabsorbed from the proximal tubule despite variations in GFR. The net result of glomerulo-tubular balance is to mitigate the influence that even small changes in GFR have on the amount of Na+ and H2O delivered to the more distal nephron segments and ultimately the amount of Na+ and H2O excreted in the urine. Two mechanisms are responsible for G-T balance: 1. Starling forces: At constant RPF, a rise in GFR raises protein concentration in the plasma leaving by the efferent arteriole. This plasma enters the peritubular capillaries; the higher pc in the peritubular capillaries increases reabsorption of solutes and water. 2. A rise in GFR increases filtered load of glucose and amino acids. The rate of Na+ reabsorption depends on the filtered load of glucose and amino acids (see below as they are “co-transported”). So, as GFR increases, Na+ reabsorption increases as well. G-T balance occurs ONLY due to spontaneous changes in GFR with NORMAL Na+ balance (ie., normal volume, normal blood pressure, etc.). There are pathologic situations when glomerulo-tubular balance does not occur, such as severe volume loss, when proximal reabsorption may increase while GFR decreases. Under these circumstances, extrinsic influences signal the proximal tubule so that up to 90% of the filtered fluid and solute may be reabsorbed. WSUSOM Medical Physiology Rossi-Renal Physiology Page 5 of 15 Proximal Tubule Tm Mechanisms Tm : Tubular Transport Maximum Tm reabsorption: from TF to P (lumen to capillary) carbohydrates amino acids metabolites HPO42- and H2PO4- Tm secretion: from P to TF organic acids (PAH, penicillin, indomethacin) organic bases (creatinine, cimetidine, uric acid) Tm Reabsorption UgluV = (GFR * Aglu) – Rglu when Rglu = Tmglu, then glucose appears in urine Colors would be good here...see slide show on video if it helps you...or bring colored pencils to class. (see full explanation below) non-Tm Reabsorption A maximal transport rate is not apparent for some substances reabsorbed by the PCT Ionic: Na, K, Cl, HCO3 Nonionic: urea This does not imply that transport proteins are not involved in non-Tm reabsorptive mechanisms!!! It just means that there does not seem to be a maximum level or ceiling above which the substance cannot be transported. WSUSOM Medical Physiology Rossi-Renal Physiology Page 6 of 15 Proximal Tubule Tm Mechanisms H2O Reabsorption by Proximal Tubule Proximal tubule has high water permeability 60-80% of filtered solutes are reabsorbed by the proximal tubule cells Water follows by osmosis in isotonic fashion via aquaporin 1 (see explanation below re: “micro environments”) Proximal tubule fluid is always isotonic to renal cortical ISF Water in the interstitium then moves into the peritubular capillary because of high oncotic pressure (remember the proteins stayed behind after filtration in the glomerulus) In the proximal tubule, water is reabsorbed passively by osmosis and isotonically via water channels known as aquaporin 1. (see animation during lecture) The driving force for H2O is provided by the active reabsorption of solute, primarily Na+. As Na+ and other solutes are reabsorbed across the apical (luminal) membrane into the cell and then are extruded by the basolateral membrane into the ISF, small osmotic gradients develop from the TF to the intracellular compartment and then from the intracellular compartment to the ISF. This sets up a “micro-environment” just below the cell membrane surface, with higher osmolality, to cause water to move into the cell (likewise, pulling water out of the cell on the basolateral side). These tiny changes in osmolality ~6 mosm/kgH2O are the driving force for water to move. Overall, however, the movement of water when looked at a “macro” level appears to be, and is CALLED, isotonic movement. This provides the osmotic force that permits H2O to flow from the TF into the cell then to the ISF and finally into the peritubular capillary. Some H2O passes through the tight junction between the cells because of the osmotic gradient, but more goes through the cells in the proximal tubule via aquaporin 1. Recall also that the peritubular capillary just came off the efferent arteriole so that the plasma has a very high protein concentration and, therefore, a high oncotic pressure. Thus, fluid and salts are drawn into the capillary and reabsorption is achieved. WSUSOM Medical Physiology Rossi-Renal Physiology Page 7 of 15 Proximal Tubule Tm Mechanisms Tm for Glucose Tm = the maximum rate of transport by the tubule for that substance; it was considered a constant (until very recently, but still a good approximation under most circumstances) Threshold = plasma concentration at which reabsorptive mechanism is saturated. In other words, the threshold is the highest plasma concentration at which UgluV = 0 If 0 = (GFR * A glu) - Rglu and when Aglu = threshold Then when Tm = maximum Rglu 0 = (GFR * threshold) – Tm Thus Tm = GFR * threshold Note: If GFR doubles, then threshold is halved. IN untreated diabetes mellitus: Aglucose may be > threshold The reabsorption of glucose is a classic example of a proximal tubule Tm mechanism. The diagram shows filtered, reabsorbed, and excreted glucose as a function of arterial glucose concentration, Aglucose. In this example,GFR = 125 ml/min. Definition of concepts: 1. threshold 2. Tm 3. splay Be sure to keep the concepts of threshold and Tm separate. Everyone tends to confuse them! Threshold - the plasma concentration below which the substance (in this case, glucose) is not found in the urine. On the diagram below, the threshold is ~3 mg/ml, or 300 mg/dl, which is 3x the normal plasma concentration of 100 mg/dl. E.g., , if the threshold for glucose is 3 mg/ml and GFR is 125 ml/min then Tm for glucose is 375 mg/min (125 ml/min * 3 mg/ml = 375 mg/min) Tm for reabsorption - the maximum reabsorptive rate of a substance and is constant (exception is Tm for phosphate). Tm is constant, but threshold may vary and depends on both GFR and Tm. Example: If GFR doubles (from 100 to 200 ml/min) and the Tm is constant, then the threshold would be halved (from 300 to 150 mg/dl). This actually happens in pregnancy where GFR can increase by 50-100% and the threshold for glucose decreases accordingly. So pregnant women with normal serum glucose can have glucose in the urine. Splay – Not all nephrons are identical. Thus, the threshold is not a definitive point for the kidney as a whole, there is WSUSOM Medical Physiology Rossi-Renal Physiology Page 8 of 15 Proximal Tubule Tm Mechanisms variation among individual nephrons, so that some non-linearity, or splay, in the reabsorption and excretion curves occurs near threshold. Tm Reabsorption – Glucose, an example Colors are helpful here...see slide show on video if it helps you...or bring colored pencils to class. As glucose gets filtered, the amount in the tubular fluid rises (blue dashed line). Note that the rate of rise is identical to the GFR which is the slope of the red line. When the amount of glucose filtered is lower than the Tm (dot-dash line), all of the glucose gets reabsorbed and none appears in the urine. Once the amount of glucose filtered exceeds the Tm, that is, the maximum reabsorptive rate is reached, the glucose in the filtrate exceeds the amount that can be reabsorbed and glucose begins to appear in the urine (solid black line) and is excreted. The maximum PLASMA concentration at which the amount filtered can be fully reabsorbed is the THRESHOLD. In other words, the PLASMA concentration above which glucose begins to first appear in the urine is the THRESHOLD. There are many ways of phrasing this. You may also wish to follow the animated sections of the lecture presentation…so as not to make the notes even longer. Uglucose * V = (GFR * Aglucose) - Rglucose Tm = maximum Rglucose Effect of Change in GFR on Threshold Note the solid line for filtered load (= GFR x Aglu). Also, recall that reabsorption reaches max when FR the line denoting “filtration rate” crosses the line FR wG Lo for Tm. Dropping a vector to the glucose hG concentration when this line crosses the line for Hig Tm gives the threshold. When the GFR increases the amount of glucose filtered per unit time also increases. The “filtration rate” line becomes steeper (dash-dot). Notice how this line crosses the Tm line at a lower concentration of blood glucose, that is, the threshold for glucose decreases. Pregnancy is an example of this. WSUSOM Medical Physiology Rossi-Renal Physiology Page 9 of 15 Proximal Tubule Tm Mechanisms Likewise, when GFR decreases, the slope of the filtration rate line is less steep, the line (dashed) crosses the Tm line later and the threshold increases. Chronic kidney disease is an example of this. Tm: characteristics of 2º Active Transport Saturability analogous to the Vmax in enzyme reactions transporter (carrier); substrate (glucose) [glucose] ® ­ Rglu until carrier is “saturated” carrier always saturated with respect to Na+ (150 mM) vs glucose (5.6 mM), so Na+ is never rate limiting Specificity e.g., transport of the “D” not the “L” optical isomer Competitiveness affinities: also transports fructose, galactose, xylose Note that inulin (5-carbon sugar) or mannose (6-carbon sugar) do NOT compete All Tm mechanisms exhibit three characteristics of secondary (or tertiary) active transport, listed above. 1. Saturability - (analogous to enzymatic reactions) At a constant transporter number in the membrane, and increase in substrate concentration will increase the transport rate until all the carriers (transporters) for that substance are saturated. When the carriers are saturated and transporting at the maximum rate, the Tm is achieved. In the case of glucose (and amino acids, phosphate, metabolites) reabsorbed by the proximal tubule, Na+ is also a substrate and is co-transported with glucose. Since the Na+ concentration is so high (150 mM) relative to glucose (100 mg/dl = 0.56 mM), the Na+- glucose co-transporter is always saturated with Na+, so that the Na+ concentration is never rate limiting. 2. Specificity - carriers are specific for the molecules they transport. e.g., “D” but not “L” glucose is transported glucose and galactose but not mannose are transported. 3. Competitiveness - The carrier has affinities for glucose, fructose, galactose and xylose. These four carbohydrates compete for the same carrier. E.g., If the plasma concentration of fructose increases, it will decrease the apparent maximum reabsorptive rate (Tm) for glucose. WSUSOM Medical Physiology Rossi-Renal Physiology Page 10 of 15 Proximal Tubule Tm Mechanisms Cellular Transport: Glucose Cell characteristics Na,K-ATPase on basolateral mmb luminal and basolateral mmb permeability to K >> Na PD of ICF is negative vs TF NaICF [K]ISF. The membranes are more permeable to K+ than Na+, so the PD (potential difference) across both apical and basolateral membranes approximates the Nernst potential for K+ (-80mV inside neg). Thus, there are large electrical and chemical gradients for Na+ to move from TF into ICF (intracellular fluid). The Na-glucose co-transporter, therefore, moves Na+ downhill coupled to glucose movement uphill into the cell. Glucose then moves, via GLUT2, from ICF to ISF (interstitial fluid) and then into the plasma. SGLT1 and GLUT1 are present in the later part of the proximal tubule. Glucose is reabsorbed by secondary active transport. Energy is derived from coupled movement of a solute (Na+) down its electro-chemical gradient and permits glucose to be reabsorbed against its electrochemical gradient. Secondary active transport can transport solute against enormous gradients! Under normal conditions, filtered glucose is completely reabsorbed, so that TF glucose concentration decreases to zero, whereas ISF glucose concentration is approximately that of plasma. Indeed, nearly an infinite gradient!!! Primary active transport (eg., Na,K-ATPase) depends directly on ATP hydrolysis to provide the energy for transport a solute against its electrochemical gradient. WSUSOM Medical Physiology Rossi-Renal Physiology Page 11 of 15 Proximal Tubule Tm Mechanisms The Na,K-ATPase in the basolateral membrane is KEY in maintaining [Na+] in the ICF low. If the Na,K-ATPase is poisoned, ICF [Na+] will increase and the driving force for secondary active transport (the downward movement of Na+ from TF into the cell) will be dissipated and Na coupled transport will stop. Diabetes mellitus - Normally the plasma concentration, Aglucose is much lower than threshold for glucose; therefore, all the filtered glucose is reabsorbed. In untreated diabetes mellitus, Aglucose may be as high as 500 to 1000 mg/dl, >> threshold. Under these conditions, more glucose is filtered than can be reabsorbed. The maximum reabsorptive capacity for glucose (Tm) is exceeded. Consequently, glucose is excreted in the urine. SGLT2 is the target of the NEW diabetes medications that you are hearing about in all the TV commercials: the gliflozins. (These are exciting new drugs that have been found to decrease cardiovascular risk and deterioration of diabetic kidney disease!) SGLT2 is a high capacity/low affinity transporter that transports Na+ and glucose at ratio 1:1 and can establish a cell to TF glucose ratio of 70. SGLT1 is a low capacity/high affinity transporter that transports Na+ and glucose at ratio 2:1 and can establish a cell to TF glucose ratio of 4,900. NEW INFORMATION Textbooks and my notes SGLT2 inhibitor used to say that the Tm for glucose does not change and that it is largely the same in all humans. No one bothered to check until the new SGLT2 SGLT2 inhibitor inhibitors came out. Note in the figure that the Tm for type 2 diabetics is higher than that for healthy controls. Inhibition of the SGLT2 transporter decreases the Tm for glucose in both healthy and type 2 diabetic subjects. Diabetes Care 36:3169–3176, 2013 Question: What would you expect urinary excretion of glucose to do if a person is given a dose of a gliflozin drug? (increase/decrease) What would you expect Na excretion to do after the person is given the SGLT2 inhibitor? (increase/decrease) WSUSOM Medical Physiology Rossi-Renal Physiology Page 12 of 15 Proximal Tubule Tm Mechanisms FACTS about Tm Mechanisms ALL are at least secondary active transport (some may be tertiary) ALL exhibit saturation specificity competition ALL derive energy from downhill movement of Na into the cell For many years, it has been generally accepted that ALL Tm mechanisms are in the proximal tubule. New findings in the distal nephron may alter this concept, but remain controversial at this time. Other Substances with Tm Reabsorption Amino acids: 3 types (at least) apical transporters: basic, acidic, other specificity for “L” isomer saturable but concentration