PTH that is secreted by the parathyroid gland in response to variations of serum Ca²⁺ results in changes in intestinal absorption of calcium via enhanced 1,25(OH)₂ D (D3) production and changes in DCT renal calcium absorption. The regulation of PTH is intimately controlled by calcium concentration and occurs at the level of transcription and changes in intracellular degradation of PTH (25). PTH release is triggered by hypocalcemia and modulated by prostaglandin E2, dopamine, and adrenergic agonists (25). In the parathyroid gland Ca²⁺ is sensed by the CaSR which controls PTH secretion (26). CaSR induces through cyclic AMP transcription of parathyroid hormone (PTH) (26). FGF23 receptor (Klotho) expressed on the parathyroid gland also regulates PTH secretion (27). Klotho is stimulated by hyperphosphatemia to positively regulate the secretion of PTH secretion. If the serum calcium concentration drops this results in increased PTH secretion, which induces 1-alpha-hydroxyalse transcription in the kidneys and promotes 25-alpha-hydroxylase in the liver (28) ( Figures 2 , 6 ).

In the DCT, PTH and vitamin D are involved specifically also in regulation of calcium at the transport level through regulation of calbindin (intracellular protein) and TRPV5 and TRPV6 as well as CD28/calbindin. Depletion of vitamin D results in decreased expression whereas both high levels of vitamin D and PTH result in higher expression of TRPV5 and 6 and CD28 (1). Once PTH is secreted, the effect of increasing vitamin D3 production also stimulates calcium and phosphate absorption through the GI tract. Vitamin D3 then enters the enterocyte resulting in binding to the intracellular vitamin D receptor, and expression of various proteins. Calcium transport can occur via channels, active transport, and paracellular transport across enterocytes—and these processes are regulated by vitamin D3. The specific proteins include calbindin, PMCA3 (mediator complex subunit Med27), and the exchanger NCX1 expressed in the luminal surface of enterocytes. This is in addition to channels like TRPV6, and paracellular transport via claudins 2, 12, and 15 across the enterocyte (29) (see Figure 2 ). There are other proteins active in calcium transport in the enterocyte: calcium channel, voltage-dependent, L type, and alpha 1D subunit (Cav 1.3) are involved as well in calcium entry into enterocytes; active transport is mediated by the plasma membrane Ca²⁺-ATPase (PMCA1b), and finally Calbindin9k allows increased enterocyte calcium absorption in response to vitamin D3 ( Supplemental Figure A ).

Changes in urinary calcium excretion can occur secondary to an alteration on the blood calcium concentration with concomitant changes in the filtered load rate and rate of tubular calcium absorption. Hypercalcemia results with increased urine calcium excretion, due to a higher filtered load and lower rate of calcium reabsorption by the nephron (30). Hypercalcemia can cause renal vasoconstriction which tends to lower the filtered load (30). Renal calcium excretion drops in hypocalcemia mainly through the mechanism of a lower filtered load and a compensatory increase in tubular calcium reabsorption (30).

Alterations in urine pH can result in hypercalciuria in the DCT segment; chronic metabolic acidosis results in hypercalciuria, whereas alkalinization results in decreased urinary calcium excretion (1). The urine calcium excretion varies with the serum bicarbonate concentration (31). This is known to be due to changes in renal tubular calcium absorption rather than changes in the filtered load (32–34). The likely mechanism is the pH effects on the TRPV5 calcium channel in the DCT (18, 35, 36).

Volume expansion decreases tubular absorption of sodium, chloride, and calcium with opposite changes occurring during volume contraction (29). The major site of this effect is thought to be the proximal tubule (29). During volume expansion, there is an increase in GFR and filtered load of calcium (4, 37). The decrease in proximal tubular calcium absorption is proportional to the decrease in sodium and water absorption such that the luminal calcium concentration remains unchanged (29). The absolute amount of calcium absorbed in the loop of Henle during volume expansion is increased above control (29).

In the proximal tubule, osmotic diuretics (i.e., mannitol) block paracellular water reabsorption from the decreased calcium absorption through a solvent drag mechanism (38). Acetazolamide also blocks calcium absorption by blocking water absorption. This effect is mediated by the creation of a luminal disequilibrium pH that inhibits bicarbonate absorption mediated by NHE3 (39). SGLT2 inhibitors block sodium-glucose co-transport (and possibly NHE3) in the proximal tubule, resulting in a glucose-induced osmotic diuresis and impaired bicarbonate transport (40). In a similar fashion, calcium absorption is impaired.

In the TAL, diuretics decrease calcium absorption by competing for the chloride site on the Na-K-2Cl cotransporter (41). Inhibiting NKCC2 sodium chloride reabsorption inhibits the back leak of potassium via ROMK and impairs the generation of the lumen-positive potential needed to drive paracellular calcium absorption. In neonates, chronic use of loop diuretics can be deleterious leading to the development of nephrocalcinosis (42).

In the DCT and connecting tubule, the direct effect of thiazides in the DCT is to decrease NCC transport and calcium absorption (43, 44). However, in the whole kidney, thiazide diuretics significantly decrease renal calcium excretion due to enhanced proximal tubule calcium absorption as a result of hypovolemia (44). There are accompanying changes in distal calcium delivery that modulate calcium transport in the DCT (45, 46). Postulated mechanisms for this effect include increased entry of luminal calcium via TRPV5, enhanced basolateral extrusion via the Na-Ca exchanger, and decreased levels of calcium transporters (47, 48). If thiazide therapy increases plasma calcium above 12 mg/dl or if hypercalcemia persists, primary hyperparathyroidism or another hypercalcemic state should be suspected (49).

In the cortical collecting duct, amiloride increases calcium reabsorption and reduces calcium excretion (38, 50). The mechanism by which amiloride reduces calcium excretion is not well understood. The effect of amiloride may involve both the connecting tubule in which sodium entry occurs via both ENaC Na⁺ channels and NCC cotransporters (46) and the initial cortical collecting tubule where cellular Na⁺ entry occurs only via ENaC (46, 51, 52). By hyperpolarizing the apical membrane amiloride promotes calcium influx via TRPV5 channels (50). Little is known of the effect of spironolactone and eplerenone on urinary calcium excretion in normal subjects. Spironolactone or adrenalectomy can reduce hypercalciuria (53).

FGF23 plays an extensive role in renal phosphate handling (5, 27, 54, 55). FGF23 secreted in response to hyperphosphatemia binds to FGF23R/Klotho receptors in the parathyroid and the PCT. The downstream signaling effects are achieved via calcineurin (CN) and mitogen-activated protein -kinase (MAP-K) pathways (27), due to the regulation of 1-alpha-hydroxylase and PTH by FGF23 (54). FGF23 exerts its effects via mitogen-activated kinase (MAPK) signaling, influencing the intracellular sodium hydrogen exchange regulatory factor-1 (NHERF-1) (55).

The actions of phosphatonins on calcium are intertwined with FGF23’s regulatory action on vitamin D metabolism (27, 56). In the PCT, FGF23 decreases 1-alpha-hydroxylase thereby decreasing the level of active vitamin D3. It also increases 24-hydroxylase which results in active D₃ hydroxylation and deactivation (27, 56). In doing so, FGF23 results in decreased vitamin D signaling with the vitamin D receptor (27, 56). This results in decreased calcium absorption systemically but does not result in hypocalcemia (27, 56). FGF23 also results in increased PTH signaling form the parathyroid gland. In aggregate, FGF23 stimulates PTH-mediated effects on phosphate secretion while inhibiting vitamin D3 production (27, 56) ( Supplemental Figure A ).

PTH and FGF23 modulate phosphate reabsorption through inhibition of apical sodium-phosphate (NaPi) cotransporters which reabsorb phosphate (28). They do so through protein kinase A and C regulation. Specifically, the apical transporters NaPi2a, NaPi2c, and Pit-2 are endocytosed resulting in decreased function of these phosphate transporters in the PCT brush border (5) (see Supplemental Figure A ).

In patients with chronic kidney disease (CKD), FGF23 levels rise spontaneously in response to hyperphosphatemia (26). Secondary hyperparathyroidism occurs due to rising uremic toxins and increasing phosphate (26); an effect that is CKD stage dependent (26). There is associated decreased renal calcium excretion and decreased intestinal calcium absorption. In end-stage renal disease (ESRD), these effects are more pronounced (27).

Vitamin D-resistant ricket disorders are caused by mutations in vitamin D synthesis and cytochrome P450 proteins (57). One such mutation is in 1-alpha hydroxylase (CYP27B1-chromosome 12) that is the liver enzyme for production of 1-hydroxy-vitamin D₃. Hypocalcemia and hypophosphatemia may sometimes be present. Elevated PTH and alkaline phosphatase levels are present seen due to high levels of skeletal turnover are usually seen. This autosomal recessive disorder results in a phenotype of osteomalacia, short stature, dental caries, and genu varum.

Another cause of vitamin D-resistant rickets is due to vitamin D receptor mutations encoded on chromosome 12q13.11. The inheritance pattern is autosomal recessive, and the biochemistry profile is similar to other cases of vitamin D-resistant rickets (58). X-linked hypophosphatemic rickets is encoded in the PHEX gene and does not affect calcium metabolism (59).

Disorders resulting in low serum calcium include autosomal dominant hypocalcemia (types type 1 and 2) (60). Type 1 is typically caused by mutations in CaSR, where a constitutively active CaSR results in decreased levels of PTH and vitamin D3. Constitutive activation of CaSR (type 1; autosomal dominant) results in reduced calcium absorption and hypocalcemia (60). Type 2 autosomal dominant hypocalcemia is typically caused by mutations in the GNA11 gene (encoded in chromosome 19p13.3), which produces G protein alpha subunit 11 that regulates CaSR (61). Hypocalcemia is due to decreased intestinal calcium absorption. FHH (familial hypercalcemia with hypocalciuria) results from inactivating mutations in CaSR. This disorder results in a higher-than-normal constitutive expression of PTH resulting in a mimic of hyperparathyroidism. Clinically, a modest elevation of serum calcium and PTH are present associated with an abnormally low urine calcium excretion rate, as opposed to the normal or high rate of calcium excretion in primary hyperparathyroidism (60). In cases of 2 nonfunctional CaSR alleles or mutations that result in severely reduced CaSR activity, the phenotype is more severe and is referred to as neonatal severe hyperparathyroidism.

Familial hypoparathyroidism (autosomal recessive) can result from parathyroid hormone loss-of-function mutations (encoded on chromosome 3p21) which can also occur resulting in poor or absent hormonal signaling (62). Predictably, inactivating mutations result in poor GI calcium absorption and hypocalcemia. Genetic syndromes of magnesium wasting (TRPM6, SLCA4 mutations) can mimic inactivating PTH mutations, because PTH is unable to function properly without magnesium as a cofactor (62).

Familial hyperparathyroidism (autosomal dominant) can conversely result from gain-of-function mutations in PTH, encoded as above (63). The clinical presentation is associated with high levels of serum calcium and low levels of serum phosphate without increased PTH. Hypercalciuria is expected in familial hyperparathyroidism cases, in contrast to FHH (63).

Patients with PTH-resistant hypocalcemia may have several mutations in transcription factor proteins including glial cells missing protein (GCM2), T box-1 mutations (TBX-1), SRY Box 3 (SOX3), GATA-binding protein 3 (GATA3), and tubulin-specific chaperone E. These mutations confer a resistance to PTH and vitamin D3 by affecting vitamin D3 receptor and PTH receptor expression. Transmission is autosomal recessive, and clinically the phenotype is severe hypocalcemia with high PTH levels (PTH resistance) (11).

These disorders result from mutations in MEN1 and CDC73/HPRT2 genes which map to chromosome 11q13 (64). These diseases are transmitted in an autosomal dominant manner and clinically present with hypercalcemia from parathyroid malignancies overproducing PTH. They differ from FHH based on the clinical pattern of urinary hypocalciuria (as opposed to normal or hypercalciuria in MEN1) and the risk of endocrine malignancy with MEN1 mutations.

William’s syndrome (autosomal dominant) is another genetic cause of hypercalcemia. It is caused by mutations in elastin and actin binding (LIM) kinase. Both of these proteins have been localized to genes in 7q11.23 (65). The phenotype of William’s syndrome includes hypercalcemia and behavioral abnormalities with a diminished fear response and loss of caution when approaching strangers (66).

Bartter’s syndrome causes hypercalciuria and a hypokalemic metabolic alkalosis without a change in the serum calcium. There are various types of Barrter’s syndrome (Types 1-6) involving mutations in genes encoding NKCC2, ROMK, ClC-Ka, ClC-Kb, Barttin, CaSR, and MAG-D2 (67). The effect of loop diuretics on the TAL often mimics the findings in these disorders (67–69).

Gitelman’s syndrome, an autosomal recessive disorder, is caused by mutations in NCC in the DCT. Like Bartter’s syndrome, patients with Gitelman’s syndrome have metabolic alkalosis and hypokalemia. Unlike most types of Bartter's syndrome, patients have hypocalciuria and their phenotype mimics the action of thiazide diuretics (70). Decreased renal calcium excretion is thought to be due to enhanced proximal tubule calcium absorption as a result of hypovolemia (44) and possibly via enhanced calcium reabsorption distally at the thiazide-sensitive site in the distal tubule and connecting segment (45, 46).

Gordon’s syndrome is an autosomal dominant disorder that clinically presents with hypertension, hyperkalemia, and hypercalciuria (71). The four most common mutations are WNT-signal transduction kinase 1 (WNK1), WNT-signal transduction kinase 4 (WNK4), Kelch-like protein 3 (KLHL3), and Cullin-3 (CUL3) (71). WNK1 and WNK 4 are kinases that negatively regulate the NCC transporter. KLHL3 and CUL3 are components of the ubiquitin degradation proteosome which degrade the WNK1 and 4 protein products (71). Loss-of-function mutations result in increased sodium reabsorption via NCC activity, decreased potassium excretion, hypertension, and hyperkalemia. Renal calcium excretion can be increased in Gordon’s syndrome and may lead to nephrocalcinosis (71).

EAST syndrome is an abbreviation for a clinical syndrome of epilepsy, ataxia, sensorineural deafness, and tubulopathy (72). The syndrome is also called SESAME syndrome (seizures, sensorineural deafness, ataxia, mental disability, and electrolyte imbalance) (73). Inactivating mutations in the basolateral inward rectifying Kir 4.1 potassium channel in the DCT cause the syndrome (72). Kir 4.1 is also expressed in neuronal tissue accounting for the complex phenotype. Patients present with hypokalemia, metabolic alkalosis, hypomagnesemia, and hypocalciuria. See Supplemental Table A .

Mutations in claudin 16 and 19 (familial hypomagnesemia with hypercalciuria and nephrocalcinosis, FHHNC), result in hypercalciuria, nephrocalcinosis, nephrolithiasis and renal failure (74). See Table 1 .