HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Srinivas, S. P. Articles by Bonanno, J. A. Search for Related Content PubMed PubMed Citation Articles by Srinivas, S. P. Articles by Bonanno, J. A.

Lysosomal Ca²⁺ Stores in Bovine Corneal Endothelium

From School of Optometry, Indiana University, Bloomington, Indiana.

PURPOSE. Acidic organelles, including Golgi bodies and lysosomes, are known to operate as Ca²⁺ storage sites in many cell types. This study demonstrates the presence of Ca²⁺ stores in lysosomes of bovine corneal endothelial cells (BCECs) and examines their interaction with Ins(1,4,5)P₃-sensitive Ca²⁺ stores.

METHODS. Glycyl-L-phenylalanine-ß-naphthylamide (GPN) was used to release Ca²⁺ from lysosomes by inducing their selective osmotic swelling. Ca²⁺ released into the cytoplasm was measured with fura-2 or fura-PE3 fluorescent dyes. Fluorescence of acridine orange (AO), which selectively sequesters into acidic organelles, was used to establish swelling of lysosomes in response to GPN.

RESULTS. Exposure to GPN (100–200 µM) in cultured BCECs produced an increase in free cytosolic Ca²⁺ ([Ca²⁺]_i) equivalent to approximately 79% of the peak response to uridine triphosphate (UTP), a P2Y agonist (n = 19). The endothelium of the freshly isolated cornea also produced [Ca²⁺]_i transients similar to those in cultured BCECs; however, the peak [Ca²⁺]_i increase was smaller (43% of the peak response to UTP; n = 13). In cultured BCECs, the response to UTP was unaffected by pretreatment with GPN with extracellular calcium ([Ca²⁺]_o) at 0 and 1.2 mM (n = 10). Neither pretreatment with thapsigargin (5 µM) nor with U73122 (a phospholipase C inhibitor; 10 µM) blocked the peak GPN response (n = 6). Exposure to 20 µM monensin produced a [Ca²⁺]_i increase with [Ca²⁺]_o at 0 and 1.2 mM and also reduced the subsequent peak response to GPN (n = 6).

CONCLUSIONS. GPN-sensitive lysosomal Ca²⁺ stores, distinct from Ins(1,4,5)P₃-sensitive Ca²⁺ stores, are found in both cultured cells and fresh tissue. These stores are susceptible to depletion by the loss of the pH gradient across lysosomes and P2 agonists. The latter occurs through mechanisms independent of phospholipase C (PLC) activation or Ins(1,4,5)P₃. The GPN stores also induce [Ca²⁺]_o influx in response to their depletion.

This article has been cited by other articles:

				K. H. Ahmed and B. Pelster Ionic determinants of pH of acidic compartments under hypertonic conditions in trout hepatocytes J. Exp. Biol., October 15, 2008; 211(20): 3306 - 3314. [Abstract] [Full Text] [PDF]

				J. J. Jennings Jr., J.-h. Zhu, Y. Rbaibi, X. Luo, C. T. Chu, and K. Kiselyov Mitochondrial Aberrations in Mucolipidosis Type IV J. Biol. Chem., December 22, 2006; 281(51): 39041 - 39050. [Abstract] [Full Text] [PDF]

				S. P. Srinivas, M. Satpathy, Y. Guo, and V. Anandan Histamine-Induced Phosphorylation of the Regulatory Light Chain of Myosin II Disrupts the Barrier Integrity of Corneal Endothelial Cells. Invest. Ophthalmol. Vis. Sci., September 1, 2006; 47(9): 4011 - 4018. [Abstract] [Full Text] [PDF]

				A. A. Soyombo, S. Tjon-Kon-Sang, Y. Rbaibi, E. Bashllari, J. Bisceglia, S. Muallem, and K. Kiselyov TRP-ML1 Regulates Lysosomal pH and Acidic Lysosomal Lipid Hydrolytic Activity J. Biol. Chem., March 17, 2006; 281(11): 7294 - 7301. [Abstract] [Full Text] [PDF]

				P. Gomes, S. P. Srinivas, W. Van Driessche, J. Vereecke, and B. Himpens ATP Release through Connexin Hemichannels in Corneal Endothelial Cells Invest. Ophthalmol. Vis. Sci., April 1, 2005; 46(4): 1208 - 1218. [Abstract] [Full Text] [PDF]

				P. Gomes, S. P. Srinivas, J. Vereecke, and B. Himpens ATP-Dependent Paracrine Intercellular Communication in Cultured Bovine Corneal Endothelial Cells Invest. Ophthalmol. Vis. Sci., January 1, 2005; 46(1): 104 - 113. [Abstract] [Full Text] [PDF]

				J. E. Fries, T. H. Wheeler-Schilling, E. Guenther, and K. Kohler Expression of P2Y1, P2Y2, P2Y4, and P2Y6 Receptor Subtypes in the Rat Retina Invest. Ophthalmol. Vis. Sci., October 1, 2004; 45(10): 3410 - 3417. [Abstract] [Full Text] [PDF]