Cell volume regulation is critical to overall cellular homeostasis and plays a role in a variety of pathological situations such as coma, edema, stroke and diabetes. This proposal uses a combination of electrophysiological, fluorescent and confocal microscopy and molecular biology techniques to address the physiological role of anions, osmolytes and the cytoskeleton in volume regulation. The focus is on a volume-regulated anion conductance (VRAC), which is a member of a ubiquitous family of mechanosensitive ion channels. This conductance is found in a wide variety of cell types and has been implicated in the recovery from cell swelling in many cells, including lymphocytes, the model system used in these studies. This proposal addresses several hypotheses directed toward understanding the regulation of this volume-regulated anion conductance and the contribution of this conductance to cellular volume regulation by concurrent measurements of current amplitude and cell swelling in a hyposmotic environment. Specifically addressed is the hypothesis that volume regulated anion current activity is regulated by the intracellular composition and concentration of osmolytes. Current and volume measurements will be combined with molecular biological techniques to address the hypothesis that the activity of this current is also regulated by a small, cytosolic actin binding protein, termed ICln. Changes in the actin cytoskeleton which provide cells with structure have also been implicated in regulating cell volume and have been suggested as the mechanotransducer which activated the anion current. This hypothesis will be addressed with two proteins, BotC2 and gelsolin to alter the equilibrium between F- and G- actin and determining the effect on current activation and cell swelling using electrophysiological, molecular and fluorescent techniques. Successful completion of this proposal will provide a greater understanding of the underlying mechanisms of mechanotransduction in cells and their role in volume regulation under normal and pathological conditions.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
7R01DK046672-08
Application #
2905551
Study Section
Physiology Study Section (PHY)
Program Officer
Mckeon, Catherine T
Project Start
1992-09-30
Project End
2002-06-30
Budget Start
1999-07-01
Budget End
2002-06-30
Support Year
8
Fiscal Year
1999
Total Cost
Indirect Cost
Name
Rosalind Franklin University
Department
Physiology
Type
Schools of Medicine
DUNS #
069501252
City
North Chicago
State
IL
Country
United States
Zip Code
60064
Hoffman, M M; Garber, S S (2004) Volume-dependent glutamate permeation depends on transmembrane ionic strength and extracellular Cl-. J Membr Biol 197:193-202
Garber, Sarah S; Hoffman, Mary M (2002) Cl(-) and glutamate(-) competition for a volume-regulated anion channel. Biol Bull 203:194-5
Hubert, M D; Levitan, I; Hoffman, M M et al. (2000) Modulation of volume regulated anion current by I(Cln). Biochim Biophys Acta 1466:105-14
Verghese, C; Levitan, I; Nair, C et al. (1997) Impaired lymphocyte volume regulation in schizophrenic patients with polydipsia-hyponatremia. Biol Psychiatry 42:733-6
Chanson, M; White, M M; Garber, S S (1996) cAMP promotes gap junctional coupling in T84 cells. Am J Physiol 271:C533-9
Levitan, I; Garber, S S (1995) Voltage-dependent inactivation of volume-regulated Cl- current in human T84 colonic and B-cell myeloma cell lines. Pflugers Arch 431:297-9
Levitan, I; Almonte, C; Mollard, P et al. (1995) Modulation of a volume-regulated chloride current by F-actin. J Membr Biol 147:283-94