title: PTH modulation of NCC activity regulates TRPV5 Ca2+ reabsorption.
abstract: Since parathyroid hormone (PTH) is known to increase transient receptor potential vanilloid (TRPV)5 activity and decrease Na(+)-Cl(-) cotransporter (NCC) activity, we hypothesized that decreased NCC-mediated Na(+) reabsorption contributes to the enhanced TRPV5 Ca(2+) reabsorption seen with PTH. To test this, we used mDCT15 cells expressing functional TRPV5 and ruthenium red-sensitive (45)Ca(2+) uptake. PTH increased (45)Ca(2+) uptake to 8.8 ± 0.7 nmol·mg(-1)·min(-1) (n = 4, P < 0.01) and decreased NCC activity from 75.4 ± 2.7 to 20.3 ± 1.3 nmol·mg(-1)·min(-1) (n = 4, P < 0.01). Knockdown of Ras guanyl-releasing protein (RasGRP)1 had no baseline effect on (45)Ca(2+) uptake but significantly attenuated the response to PTH from a 45% increase (6.0 ± 0.2 to 8.7 ± 0.4 nmol·mg(-1)·min(-1)) in control cells to only 20% in knockdown cells (6.1 ± 0.1 to 7.3 ± 0.2 nmol·mg(-1)·min(-1), n = 4, P < 0.01). Inhibition of PKC and PKA resulted in further attenuation of the PTH effect. RasGRP1 knockdown decreased the magnitude of the TRPV5 response to PTH (7.9 ± 0.1 nmol·mg(-1)·min(-1) for knockdown compared with 9.1 ± 0.1 nmol·mg(-1)·min(-1) in control), and the addition of thiazide eliminated this effect (a nearly identical 9.0 ± 0.1 nmol·mg(-1)·min(-1)). This indicates that functionally active NCC is required for RasGRP1 knockdown to impact the PTH effect on TRPV5 activity. Knockdown of with no lysine kinase (WNK)4 resulted in an attenuation of the increase in PTH-mediated TRPV5 activity. TRPV5 activity increased by 36% compared with 45% in control (n = 4, P < 0.01 between PTH-treated groups). PKC blockade further attenuated the PTH effect, whereas combined PKC and PKA blockade in WNK4KD cells abolished the effect. We conclude that modulation of NCC activity contributes to the response to PTH, implying a role for hormonal modulation of NCC activity in distal Ca(2+) handling.
authors.affiliation: Division of Nephrology, Department of Medicine, Emory University, Atlanta, Georgia; Atlanta Veteran's Administration Medical Center, Decatur, Georgia;
authors.forename: Robert S
authors.initials: RS
authors.lastname: Hoover
authors.affiliation: Department of Integrative Biology and Pharmacology, The University of Texas Health Science Center, Houston, Texas; Institute of Cytology, Russian Academy of Sciences, St. Petersburg, Russian Federation; and.
authors.forename: Viktor
authors.initials: V
authors.lastname: Tomilin
authors.affiliation: Department of Medicine, University of Chicago, Chicago, Illinois.
authors.forename: Lauren
authors.initials: L
authors.lastname: Hanson
authors.affiliation: Department of Integrative Biology and Pharmacology, The University of Texas Health Science Center, Houston, Texas;
authors.forename: Oleh
authors.initials: O
authors.lastname: Pochynyuk
authors.affiliation: Department of Medicine, University of Chicago, Chicago, Illinois bko@uchicago.edu.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Ko
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism*; D002460:Cell Line; D007686:Kidney Tubules, Distal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D010281:Parathyroid Hormone / Q000494:pharmacology*; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism; D064486:Solute Carrier Family 12, Member 3 / Q000378:metabolism; D050916:TRPV Cation Channels / Q000378:metabolism*
chemical_list: D015220:Calcium Channels; D010281:Parathyroid Hormone; D018160:Receptors, Cytoplasmic and Nuclear; C578875:Slc12a3 protein, mouse; D064486:Solute Carrier Family 12, Member 3; D050916:TRPV Cation Channels; C492240:Trpv5 protein, mouse; C040985:phosphatidylinositol receptors; D002118:Calcium
keywords: parathyroid hormone; sodium-chloride cotransporter; thiazide; transient receptor potential vanilloid 5 channel; with no lysine kinase