1. Povider:Dr.nourabadi

2. Provider: Davood Nourabadi Ph.D of medical physiology-IUMS Povider:Dr.nourabadi

3. RefrenceRefrence 1.Brenner & Rector’s the kidney,Maarten W. Taal …,9 th ed,2012. 2.Bern&Levy physiology,6 th ed,2008. 3.Medical Physiology, Boron& Boulpaep, 2 th ed, 2012. 4.Seldin.and.Giebisch's.The.Kidney.5 th ed,2013. Povider:Dr.nourabadi

4. Ca 2+ Homeostasis  Tight control of blood calcium levels  Many vital physiologic functions: Muscle contraction Signaling Neuronal excitation Bone formation Coagulation Cell division and growth Apoptosis Neurotransmitter release Povider:Dr.nourabadi

5. .. Contd..  Disturbances in the Ca 2+ balance: Seizures Rickets Heart failure  Collaborative action for calcium balance: Intestine (regulation of absorption in duodenum&colon) Bone (regulation of metabolism) Kidney (regulation of excretion) Povider:Dr.nourabadi

6.  99%is stored in bone.  Approximately 1% in ICF  0.1% in the ECF Total [Ca 2+ ] in plasma is 10 mg/dL (2.5 mM or 5 mEq/L).  Distribution of Calcium in Normal Human Plasma: (Filterable)  Diffusible (Filterable) Ionized (Ca 2+ ) Complexed to HCO 3 –, citrate, sulfate, phosphate.  Nondiffusible Bound to albumin Bound to globulin Povider:Dr.nourabadi

9. ►8 g/daily of Ca 2+ is filtered at the glomerulus ►Less than 2% is excreted into the urine ►Passive paracellular and active transcellular reabsorption in the epithelial cell layers Povider:Dr.nourabadi

10. HANDLING OF Ca 2+ ALONG NEPHRON Povider:Dr.nourabadi

12. Proximal Tubule(PTs) ◙Including the proximal convoluted tubule (PCT) and proximal straight tubule (PST) ◙65% of the filtered Ca 2+ is reabsorbed ◙Transport is passive and follows the local Na + and water reabsorption (solvent drag) ◙Not provide an independent regulation of Ca 2+ reabsorption Povider:Dr.nourabadi

14. Loop of Henle ◙Thin descending thin ascending no ◙Thin descending and thin ascending limbs of the loop of Henle virtually no Ca 2+ is reabsorbed. ◙Thick ascending ◙Thick ascending limb of the loop of Henle (TAL) is permeable to Ca 2+ Povider:Dr.nourabadi

15. Thick Ascending Limb (TAL) of Henle ◙Accounts for approximately 20% of the total Ca 2+ reabsorption.  50%paracellular-50%transcellular ◙Claudin-16claudin-19 ◙Claudin-16 (paracellin-1) and claudin-19 together form paracellular pores and determine the Ca 2+ and Mg 2+ selectivity of the paracellular junctions. Povider:Dr.nourabadi

17. Distal Convoluted Tubule (DCT) and Connecting Tubule (CNT) ◙Ca 2+ reabsorption takes place against its electrochemical gradient=transport is active ◙Tight junctions are nearly impermeable to Ca 2+ initiallater not ◙Relative contribution of the initial (DCT1) and later (DCT2) segments of the DCT and of the CNT to active Ca 2+ reabsorption is not entirely clear. ◙DCT2&CNT expressed: Na + -K + -ATPase NCX1 PMCA1b TRPV5 Calbindin-D28K Povider:Dr.nourabadi

18. Contd... ◙Transepithelial transport of Ca 2+ is a 3 step procedure: 1)Apical Entry of Ca 2+ via TRPV5 2)Intracellular Carrier Calbindin-D28K 3)Basolateral Extrusion Systems: NCX1 and PMCA1b (Energy- consuming step). Povider:Dr.nourabadi

19. Model of active Ca 2+ reabsorption in DCT2 & CNT Povider:Dr.nourabadi

20. Collecting Duct ◙Cortical part of the collecting duct (CCD) accounts for small amount (3%) of Ca 2+ reabsorption. ◙Net transport occurs against the electrochemical gradient for Ca 2+ (transport is active) Povider:Dr.nourabadi

22. Passive Ca 2+ Reabsorption  Paracellular component of epithelial Ca 2+ transport.  Takes place in the PTs and TAL. Povider:Dr.nourabadi

23. Active Ca 2+ Reabsorption  Transcellular component of epithelial Ca 2+ transport.  Is the main target site for specific regulation of Ca 2+ (re)absorption by various calciotropic hormones.  Takes place in principal cells of the DCT and CNT Povider:Dr.nourabadi

24. Characteristics of Ca 2+ Channels in the Kidney osmosensitivity TRPV4 is candidate for a molecular sensor that confers osmosensitivity Povider:Dr.nourabadi

26. TRPV5() TRPV5(E Ca C 1 ) A member of the TRP channel superfamily Intracellular N and C-terminal tails Homotetramers Six transmembrane segments (S1-S6) Pore region=S5-S6 N-terminal tail contains multiple ankyrin repeats First extracellular loop between S1-S2 contains an asparagine (complex glycosylation for regulation of channel activity) C-terminal tail contains three potential PKC,PKA,PKG sites Gatekeeper function of TRPV5 in active Ca 2+ reabsorption Increased [Ca 2+ ] i to inhibit the activity of TRPV5 TRPV6 (E Ca C 2 ) a homologous channel (75%) in proximal intestine Povider:Dr.nourabadi

27. Epithelial Ca 2+ channel TRPV5 Povider:Dr.nourabadi

29. Characteristics of TRPV5  Constitutive activity at low intracellular Ca 2+ concentrations and physiologic membrane potentials  Shows strong inward rectification  its 100 times larger selectivity for Ca 2+ than for Na +  Mg 2+ competes with Ca 2+ for entry into the cytoplasm  A constitutively open channel, because no stimulus or ligand was required  Localized in intracellular vesicles dynamin-clathrin  Reinternalized via dynamin- and clathrin-dependent processes  In contact with the extracellular environment, TRPV5 channels exhibit closed and open states (open probability)  Extracellular H + blocks channel activity Povider:Dr.nourabadi

32. Contd...Contd... ► Protein-Protein Interactions of TRPV5 ► Protein-Protein Interactions of TRPV5:  S100A10–Annexin 2  facilitates the translocation of TRPV5 toward the plasma membrane  80K-H  acts as novel Ca 2+ sensor controlling TRPV5 channel activity  Rab11a  direct role is in the trafficking of the Ca 2+ channel toward the plasma membrane Povider:Dr.nourabadi

34. Calbindin-D28KCalbindin-D28K  In principal cells of the DCT2 and CNT  Three pairs of EF hands that form the structural basis of its high Ca 2+ affinity binding capacity  Slow Ca 2+ -binding kinetics of calbindin-D28K Povider:Dr.nourabadi

35. PMCA(1b)PMCA(1b)  High-affinity Ca 2+ efflux pump  Maintain the resting Ca 2+ concentration in virtually all cells  Highest Ca 2+ -ATPase activity in kidney was reported in the DCT 30%  Capacity of PMCA in the CNT seems to be insufficient to keep pace with the absorptive flux of Ca 2+, because it can transport only approximately 30% of the total Ca 2+ efflux Povider:Dr.nourabadi

36. NCX1NCX1  Prerequisite for transepithelial Ca 2+ transport  Widely expressed protein that can be found in several tissues: Heart Brain Skeletal muscle Kidney(distal part of the nephron-CNT)  70%  70% of Ca 2+ efflux in CNT Povider:Dr.nourabadi

37. Regulation of Active Ca 2+ Reabsorption  By Calciotropic Factors  Vitamin D  Parathyroid Hormone  Estrogens  Dietary Ca 2+ Intake  Acidosis and Alkalosis  Klotho  Tissue Kallikrein  By Immunosuppressives and Diuretics  Calcineurin Inhibitors  Furosemide  Thiazide  By Calciotropic Factors  Vitamin D  Parathyroid Hormone  Estrogens  Dietary Ca 2+ Intake  Acidosis and Alkalosis  Klotho  Tissue Kallikrein  By Immunosuppressives and Diuretics  Calcineurin Inhibitors  Furosemide  Thiazide Povider:Dr.nourabadi

38. Vitamin D  Synthesized in the PTs by 1α-hydroxylase  Genomic transcriptional mechanisms with a nuclear vitamin D receptor (VDR)  VDR-RXR complex  Intestine and kidney are the main target organs ↑  Effects (↑):  TRPV5  Calbindins (9K&28K)  NCX1  [Ca 2+ ] p  Bone formation Povider:Dr.nourabadi

39. PTH  A key role in maintaining Ca 2+ in ECF by CaSR (most important regulator)  Stimulates the activity of 1α-hydroxylase in the PT  PTH receptor mRNA in rat kidney (NOT thin limb of the loop of Henle&CD)  Directly stimulates active Ca 2+ reabsorption in the distal part of the nephron  In the TAL, PTH increases the transepithelial driving force for Ca 2+ reabsorption, enhancing paracellular Ca 2+ transport. Povider:Dr.nourabadi

40. PTH (contd..) ►Mechanisms of PTH action for the effect in the DCT:  Membrane insertion of apical Ca 2+ channels  Opening of basolateral CLC (cellular hyperpolarization)  Modulation of PMCA activity Povider:Dr.nourabadi

42. EstrogensEstrogens  Estrogen deficiency after menopause results in bone loss= increase in plasma and urinary Ca 2+  Estrogen receptors (ER  ) also reside in proximal and distal tubules within the nephron (independent of Vit-D)  Upregulates the expression of TRPV5 in kidney (independent of Vit-D) Povider:Dr.nourabadi

43. Dietary Ca 2+ Intake vitamin D–independent  Dietary Ca 2+ can affect active Ca 2+ (re)absorption via vitamin D–independent modulation of the expression of Ca 2+ transport proteins.  Hypercalcemia also activates the CaSR in the TAL, which subsequently inhibits the activity of Na +- K + -2Cl – cotransporter 2 (NKCC2) and therefore passive paracellular Ca 2+ transport Povider:Dr.nourabadi

44. Acidosis and Alkalosis  Chronic metabolic acidosis is associated with increased renal Ca 2+ excretion protons  Extracellular protons inhibit TRPV5 channel activity pH sensorpH sensitivity is mainly mediated by glutamate at position 522 and may act as the “pH sensor” of TRPV5. Povider:Dr.nourabadi

45. Klotho  Antiaging hormone  Secreted and activated by cleavage of the N-terminal extracellular domain  Exhibits β-glucuronidase activity  Expressed in the DCT/CNT and the parathyroid gland  Klotho may work by affecting the extracellular glycosylation status of the TRPV5  Increased expression of Na + -K + -ATPase Povider:Dr.nourabadi

47. Tissue Kallikrein  A serine protease that is expressed mainly in the DCT and CNT  Involved in the regulation of water, sodium, and potassium metabolism  Enhances active Ca 2+ reabsorption through an increased TRPV5 plasma membrane expression via the BK-activated phospholipase/DAG/PKC pathway Povider:Dr.nourabadi

48. Calcineurin Inhibitors  Tacrolimus Reduced calbindin-D28K levels Downregulation of the renal mRNA and protein expression of TRPV5  Cyclosporine High bone turnover Osteopenia Hypercalciuria Suppresses the expression of calbindin-D28K Povider:Dr.nourabadi

50. Furosemide  Inhibit renal Ca 2+ reabsorption  Inhibits the NKCC2 transporter of the TAL Reduction in NaCl reabsorption and K + recycling across the apical membrane  Increase the expression level of TRPV5 and calbindin- D28K Povider:Dr.nourabadi

51. Thiazide  Decreasing Na + reabsorption while increasing Ca 2+ reabsorption  Two hypotheses exist with respect to the Ca 2+ -sparing effect of thiazides 1)Renal salt and water loss=> contraction of ECF V => triggers a compensatory increase in proximal Na + reabsorption 2) Stimulates Ca 2+ reabsorption in the DCT Povider:Dr.nourabadi

52. THANK YOU Povider:Dr.nourabadi