J.B. WadeMineral Metabolism

Objectives:

1) Discuss sites of phosphate transport and the importance of the kidney to excretion of excess phosphate.

2) Discuss sites of calcium transport and the cellular mechanisms involved in calcium sensing and transport.

3) Provide an integrated overview of phosphate-calcium homeostasis and the role of the kidney in body mineral homeostasis.

4) Review bone formation and resorption and describe the action of PTH on bone.

Readings: Koeppen and Stanton: Chapt. 9 p.155-167. A. Phosphate Transport

The phosphate transport curve below (Fig. 1) uses a traditional analysis that nephrologists often use to analyze that renal handling of a substance as the plasma concentration of the substance is changed. Plotted are the filtered amount of the substance, the amount reabsorbed and the amount excreted as a function of the plasma concentration. Since phosphate is freely filtered, the amount filtered goes up linearly as a function of the plasma concentration. This filtered phosphate is reabsorbed by the tubules such that when the amount filtered is low all of it is reabsorbed and none excreted. But the ability of renal tubules to reabsorb phosphate is very limited with the phosphate transport maximum (called Tm

in mmol/min) reached when plasma phosphate rises only slightly above the normal level (the region between the arrows at the bottom of Fig 1). Thus a small increase in plasma phosphate can result in enough of an increase in filtered phosphate to saturate phosphate reabsorption ability of the tubules and result in its excretion in the urine. For this reason the kidneys represent a powerful mechanism for regulating excessive plasma phosphate. A high phosphate diet decreases the Tm for phosphate while a low phosphate diet increases it. As discussed in more detail below, PTH (independent of changes in phosphate diet) inhibits phosphate reabsorption by the proximal tubule (via increased cAMP).

Fig. 1

Most (~80%) of phosphate is reabsorbed by proximal tubules via a cotransporter that appears to transport two Na+ ions with either HPO4

-2 or H2PO4-. A small amount of phosphate is reabsorbed

distally and normally about 10% of filtered phosphate is excreted.

Fig. 2

B. Calcium Homeostasis

Total Ca++ in plasma is about 5mEq/L and must be maintained within narrow limits to avoid hyperexcitability of nerve and muscle cells (tetany) due to low plasma Ca++ or cardiac arrhythmia from hypercalcemia. The organs and hormones regulating calcium homeostasis are shown below.

Calcitonin has been shown in animal studies to promote the formation of bone and lower plasma Ca++. The importance of calcitonin in human physiology is debated because removal of the thyroid gland which produces calcitonin (without removal of the parathyroids) does not seem to affect Ca++ homeostasis.

About 50% of plasma Ca++ is ionized and 5% bound to small anions (HCO3

-, HPO4-) and

therefore also filterable. The remaining 45% is bound to plasma proteins and not filtered. Normally 99% of filtered Ca++ is reabsorbed and 1% excreted (~200 mg/d, the amount absorbed by the GI tract). Most Ca++ (70%) is reabsorbed by proximal tubules and the remainder by the loop of Henle (20%), distal tubule (5-10%) and collecting duct (<5%).

A model for cellular Ca++ reabsorption is shown in Fig. 3. Entry into the cell is down a very favorable

electrochemical gradient. Transport across the basolateral membrane is believed to be via a Ca ++ - ATPase and/or the Na+-Ca++ exchanger. Ca++ is also believed to be absorbed passively via the paracellular pathway due to the lumen positive potential present at the end the proximal tubule and the thick ascending limb of the loop of Henle.

Fig. 3

C. The Calcium-Sensing Receptor and the Regulation of TAL Ca++ Reabsorption

Recently a Ca++ sensitive receptor (CaSR) has been cloned that senses extracellular Ca++ concentration. This G-protein-coupled receptor is present in the parathyroid gland where is its sensitivity to Ca++ regulates PTH release from the parathyroid gland. Binding of Ca++ by the CaSR reduces PTH in circulation. Mutations in the CaSR have been identified with diseases of calcium homeostasis. The CaSR is also present in renal nephron segments where it is believed to regulate the transporters that bring about the reabsorption of Ca++. How this might work is illustrated in Figure 4. When Ca++ is high it binds to the CaSR on the basolateral side of the TAL to trigger an inhibition of cAMP, reducing the activation of the NKCC transporter in the apical membrane. Ca++ activation of the CaSR also stimulates arachidonic acid (AA) production, which is believed to lead to inhibition of the cotransporter and the apical K channel. This will cause not only a drop in sodium reabsorption but also a reduced lumen positive potential with the result that less Ca++ will be reabsorbed and more excreted in the urine to correct the hypercalcemia.

Fig. 4

Recent observations indicate that the CaSR also mediates an action of hypercalcemia on ADH action by being present at the apical surface of collecting ducts. It appears that if luminal Ca++ rises such that there is danger of a stone forming, the CaSR signals the down regulation of the AQP2 water channels responsible for water permeability to limit luminal Ca++ concentration. The chain of events leading to increased excretion of Ca++ in a more dilute urine is shown in Figure 5.

Fig. 5

PTH is also an important regulator of renal Ca++ excretion. Although overall it enhances Ca++

reabsorption, PTH actually decreases Ca++ reabsorption proximally (presumably this is related to PTH's inhibition of fluid and phosphate transport at this site) but it dramatically stimulates Ca++ reabsorption by the thick ascending loop of Henle and distal tubule. The net result is a decrease in urinary excretion of Ca++ when plasma Ca++ is low and PTH is high.

D. Overview of Body Ca++ -Phosphate Homeostasis

Maintaining plasma calcium and phosphate within physiological limits requires close coordination between the kidney, bone, gut and parathyroid gland. The response is mediated by PTH and Vitamin D metabolites. Calcium and phosphate balances are tightly linked because the body cannot mobilize bone to maintain plasma calcium without also adding phosphate to the circulation. PTH produced by the parathyroid gland protects the body from this excess phosphate by telling the kidney to stop reabsorbing phosphate and excrete it into the urine. The complex interplay in response to low plasma calcium is shown in Fig. 6. The parathyroid gland (PTG) responds to the low calcium by releasing PTH, which has the effects on the kidney, described above (reduced excretion of calcium, increased excretion of phosphate). In addition, PTH acts on the bone to cause resorption to provide calcium. Phosphate is also moved into circulation but the body is protected from the excess phosphate because the kidney is excreting it into the urine.

Fig. 6

At the same time vitamin D3 (the "sunshine vitamin") is being produced by the skin and converted into 25(OH)D3 by the liver. In hypocalcemia or low phosphate states the kidney converts this to 1,25(OH)2

D3 (also known as calcitriol) which acts on bone to promote resorption and promotes absorption of calcium and phosphate from the gut. This renal conversion of D3 by proximal tubules is stimulated by

PTH but it can also occur in a low phosphate state even when calcium is normal and PTH is low. Conversely, a high plasma phosphate will tend to decrease calcitriol. In this way absorption of calcium and phosphate from the gut is regulated depending on need. They are also mobilized from the bone if needed with the excess ion being excreted in the urine as necessary to maintain calcium-phosphate homeostasis. Patients with renal failure often have disturbed calcium-phosphate homeostasis because their kidneys are unable to excrete phosphate.

Fig. 7

E. The action of PTH on bone

Bones are composed of osteoid which is an extracellular organic matrix onto which calcium-phosphate salts are deposited. Osteoid is synthesized and secreted by the bone-forming cells, osteoblasts, which form a continuous sheet on the surface of newly formed bone (Fig. 7). Osteoblasts that become entrapped in the bone matrix differentiate into osteocytes which interconnect with other trapped osteocytes via thin syncytial processes that run through the canaliculi. The other important bone cell is the osteoclast. These are large multinucleated cells that form from the fusion of monocyte/macrophage cells from the bone marrow. Osteoclasts function in bone resorption by secreting acids that break down the bone components to make Ca++ and phosphate available.

PTH acts indirectly to increase bone dissolution by osteoclasts. Although osteoclasts lack PTH

receptors, neighboring osteoblasts respond to PTH and active vitamin D (calcitriol) to promote bone resorption by digesting osteoid covering bone that impedes the osteoclasts. Most importantly, osteoblasts also secrete cytokines that stimulate the number and activity of osteoclasts with the result that the movement of Ca++ and phosphate into the ECF is increased.

Fig. 8

Action of calcium/phosphate regulating hormones

Bone Kidney GI tract

PTH Increases bone resorption Increases distal calcium No direct action; by indirectly reabsorption; decreases phosphate indirectly stimulates activating osteoclast reabsorption proximally; calcium and phosphateactivity increases renal conversion of absorption by activating

25-(OH)D3 to 1,25-(OH)2D3 renal synthesis of calcitriol

(calcitriol)

Calcitriol Synergizes with PTH Increases calcium Increases calcium

(1,25(0H)2 on bone to reabsorption and phosphate absorption

vitamin D3) stimulate osteoclast across intestinal mucosaactivity