Skip to Main Content
 

Global Search Box

 
 
 
 

Files

File List


NBR Thesis final.pdf (1.3 MB)

ETD Abstract Container

Abstract Header

K-Cl Cotransport: Role of KCC3 in cellular Potassium (K) homeostasis in KCC3- transfected HEK-293 cells

Ravilla, Nagendra Babu
http://rave.ohiolink.edu/etdc/view?acc_num=wright1377519708

Abstract Details

2013, Master of Science (MS), Wright State University, Pharmacology and Toxicology.
K-Cl cotransport (KCC) mediated by four protein isoforms, KCC1 to KCC4, plays a significant role in cell volume regulation, and in K and Cl homeostasis. In this study, we demonstrate the importance of KCC3 and its two threonine (T) phosphorylation sites T991 and T1048 in cellular K homeostasis in isogenic human embryonic kidney (HEK-293) cells transfected with KCC3 wild type (WT) or constitutively active double mutant KCC3 (AA) expressed under tetracycline or doxycycline control. In both WT and AA cells, under baseline conditions, the Na-K-2Cl cotransport (NKCC) (55 %- 60 %) was the major contributor of Rb influx followed by the Na/K pump (NKP) (35 % - 40 %) and KCC (10 % - 15 %) even though the absolute values of Rb influx were higher in AA cells. Doxycycline induction had no effect on either NKCC or KCC in WT cells, whereas NKCC was completely inhibited and KCC was increased by 25 fold in AA cells. In AA cells, intracellular K (Ki) decreased by 90 % upon doxycycline induction in Cl medium and 50 % of this loss was attenuated in Cl-free (sulfamate replacement) medium. Doxycycline-induced Cl-dependent Ki loss was sensitive to furosemide (2 mM) but insensitive to tetra ethyl ammonium (TEA) (2 mM) and to changes in external K and Na, whereas doxycycline-induced Cl-independent Ki loss was sensitive to 4-(2-butyl-6,7-dichloro-2-cyclopentylindan-1-on-5-yl)oxybutyric acid (DCPIB) (50 microM) but insensitive to TEA (2 mM), furosemide (2 mM) and extracellular K and Na.
Norma Adragna, Ph.D. (Advisor)
Peter Lauf, M.D. (Committee Member)
Kristopher Kahle, M.D., Ph.D. (Committee Member)
Mauricio Di Fulvio, Ph.D. (Committee Member)
53 p.
Biophysics; Cellular Biology; Molecular Biology; Pharmacology; Physiology
K-Cl cotransport, KCC3, KCC3 mutations, cellular K homeostasis, cell volume regulation

Recommended Citations

Citations

  • Ravilla, N. B. (2013). K-Cl Cotransport: Role of KCC3 in cellular Potassium (K) homeostasis in KCC3- transfected HEK-293 cells [Master's thesis, Wright State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=wright1377519708

    APA Style (7th edition)

  • Ravilla, Nagendra Babu. K-Cl Cotransport: Role of KCC3 in cellular Potassium (K) homeostasis in KCC3- transfected HEK-293 cells . 2013. Wright State University, Master's thesis. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=wright1377519708.

    MLA Style (8th edition)

  • Ravilla, Nagendra Babu. "K-Cl Cotransport: Role of KCC3 in cellular Potassium (K) homeostasis in KCC3- transfected HEK-293 cells ." Master's thesis, Wright State University, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=wright1377519708

    Chicago Manual of Style (17th edition)

wright1377519708
504
© 2013, all rights reserved.
This open access ETD is published by Wright State University and OhioLINK.