The proton extrusion mechanisms of Leishmania promastigotes were studied in terms of electrogenic movements of protons and anions (Cl and HCO). Changes in membrane potential (V) and intracellular pH (pH) were monitored fluorimetrically with the potential sensitive dye bis-oxonol and the pH-sensitive dye tetraacethoxymethyl 2`,7`-bis-(carboxyethyl)-5,6-carboxyfluorescein, respectively. In nominal bicarbonate-free medium (pH7.4, 28 °C), V and pH of Leishmania promastigotes were maintained at -113 ± 4 mV and 6.75 ± 0.02, respectively. In Cl free (gluconate-based) medium, cells underwent a time-dependent acidification (0.3 pH units) and a long term membrane hyperpolarization (7-10 mV), both of which were greatly enhanced in the presence of the anion blocker, 4,4`-diisothiocyanodihydrostilbene-2,2`-disulfonic acid (HDIDS). Cells in Cl-free medium underwent a marked depolarization upon treatment with the H-ATPase inhibitor dicyclohexylcarbodiimide (DCCD), but hyperpolarized after repletion with Cl. In Cl-depleted cells, replenishment of Cl led to a HDIDS-sensitive cytoplasmic alkalinization and a small initial hyperpolarization. Cells exposed either to DCCD or to the H uncoupler carbonylcyanide chlorophenylhydrazone caused a marked cytoplasmic acidification and membrane depolarization. In the presence of 25 mM HCO, promastigotes maintained an almost neutral cytosol, irrespective of H pump action or ionic composition of the medium. The present observations provide evidence for the operation of a DCCD-sensitive electrogenic H-ATPase which contributes to the maintenance of a highly hyperpolarized plasma membrane in Leishmania promastigotes. H pump activity required a parallel pathway of Cl ions in order to dissipate the pump generated electrical potential. In nominally CO-free media, the two electrogenic systems are implicated in the maintenance of cell pH and indirectly in electrochemically driven nutrient uptake. In physiological CO/HCO-containing media, the H pump and Cl channel play a role only secondary to that of HCO in pH homeostasis.

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

This article has been cited by other articles:

				A. Banerjee, J. Roychoudhury, and N. Ali Stearylamine-bearing cationic liposomes kill Leishmania parasites through surface exposed negatively charged phosphatidylserine J. Antimicrob. Chemother., January 1, 2008; 61(1): 103 - 110. [Abstract] [Full Text] [PDF]

				R. J. W. Allen and K. Kirk The Membrane Potential of the Intraerythrocytic Malaria Parasite Plasmodium falciparum J. Biol. Chem., March 19, 2004; 279(12): 11264 - 11272. [Abstract] [Full Text] [PDF]

				A. M. ALLAHVERDIYEV, S. UZUN, M. BAGIROVA, M. DURDU, and H. R. MEMISOGLU A SENSITIVE NEW MICROCULTURE METHOD FOR DIAGNOSIS OF CUTANEOUS LEISHMANIASIS Am J Trop Med Hyg, March 1, 2004; 70(3): 294 - 297. [Abstract] [Full Text] [PDF]

				A. Stein, G. Vaseduvan, N. S. Carter, B. Ullman, S. M. Landfear, and M. P. Kavanaugh Equilibrative Nucleoside Transporter Family Members from Leishmania donovani Are Electrogenic Proton Symporters J. Biol. Chem., September 12, 2003; 278(37): 35127 - 35134. [Abstract] [Full Text] [PDF]

				K. Kirk Membrane Transport in the Malaria-Infected Erythrocyte Physiol Rev, April 1, 2001; 81(2): 495 - 537. [Abstract] [Full Text] [PDF]

				A. Seyfang and S. M. Landfear Four Conserved Cytoplasmic Sequence Motifs Are Important for Transport Function of the Leishmania Inositol/H+ Symporter J. Biol. Chem., February 25, 2000; 275(8): 5687 - 5693. [Abstract] [Full Text] [PDF]

				G. Vasudevan, N. S. Carter, M. E. Drew, S. M. Beverley, M. A. Sanchez, A. Seyfang, B. Ullman, and S. M. Landfear Cloning of Leishmania nucleoside transporter genes by rescue of a transport-deficient mutant PNAS, August 18, 1998; 95(17): 9873 - 9878. [Abstract] [Full Text] [PDF]

				H. P. de Koning, C. J. Watson, and S. M. Jarvis Characterization of a Nucleoside/Proton Symporter in Procyclic Trypanosoma brucei brucei J. Biol. Chem., April 17, 1998; 273(16): 9486 - 9494. [Abstract] [Full Text] [PDF]

				B. Moreno, J. A. Urbina, E. Oldfield, B. N. Bailey, C. O. Rodrigues, and R. Docampo 31P NMR Spectroscopy of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major. EVIDENCE FOR HIGH LEVELS OF CONDENSED INORGANIC PHOSPHATES J. Biol. Chem., September 8, 2000; 275(37): 28356 - 28362. [Abstract] [Full Text] [PDF]