cytochalasin-d and benzamil

Sodium influx is tightly regulated in the cells of blood origin. Amiloride-insensitive sodium channels were identified as one of the main sodium-transporting pathways in leukemia cells. To date, all known regulatory pathways of these channels are coupled with intracellular actin cytoskeleton dynamics. Here, to search for physiological mechanisms controlling epithelial Na

Topics: Actin Cytoskeleton; Actins; Amiloride; Cell Membrane; Cell Membrane Permeability; Cytochalasin D; Epithelial Sodium Channel Blockers; Epithelial Sodium Channels; Humans; K562 Cells; Leukemia, Myeloid; Membrane Potentials; Microscopy, Fluorescence; Models, Biological; Patch-Clamp Techniques; Phalloidine; Sodium; Trypsin; Trypsin Inhibitors

Single-channel recording techniques were used to characterize mechanosensitive channels in identified (1.1'-dioctadecyl-3,3,3', 3'-tetramethylindocarbocyanine methanesulfonate labeled) colon sensory neurons dissociated from adult S1 dorsal root ganglia. Channels were found in 30% (7/23) of patches in a cell-attached configuration and in 43% (48/111) of excised inside-out patches. Channels were highly selective for K(+), had a slope conductance of 54 pS in symmetrical solutions, and were blocked by tetraethylammonium, amiloride, and benzamil. Channels were also seen under Ca(2+)-free conditions. Gadolinium (Gd(3+)), a known blocker of mechanosensitive ion channels, did not block channel activity. Tetrodotoxin and 4-aminopyridine were also ineffective. The cytoskeletal disrupters colchicine and cytochalasin D reduced the percentage of patches containing mechanosensitive channels. These results indicate that rat colon sensory neurons contain K(+)-selective mechanosensitive channels that may modulate the membrane excitability induced by colonic distension.

Topics: 4-Aminopyridine; Amiloride; Animals; Calcium; Colchicine; Colon; Cytochalasin D; Cytoskeleton; Diuretics; Gadolinium; Ganglia, Spinal; Ion Channel Gating; Male; Membrane Potentials; Neurons, Afferent; Nucleic Acid Synthesis Inhibitors; Physical Stimulation; Potassium; Potassium Channels; Rats; Rats, Sprague-Dawley; Sodium; Stress, Mechanical; Suction; Tetraethylammonium

The trophectoderm of the mouse blastocyst is a fluid transporting epithelium that is responsible for generating a fluid-filled cavity called the blastocoel. Vectorial transport of ions from the medium into the blastocoel generates an osmotic gradient that drives fluid across this epithelium. We report here that substitution of Na+ or Cl-, but not K+, in the medium halves the rate of blastocoel expansion in the mouse blastocyst. Entrance of Na+ into the trophectoderm may involve several routes, since both blastocoel expansion and 22Na+ uptake are decreased in the presence of the highly specific Na+/H+ exchanger inhibitor, 5-(N-ethyl-N-isopropyl)amiloride, and to a lesser extent with the amiloride-sensitive Na+-channel blocker, benzamil. Uptake of 22Na+ manifests saturation kinetics as a function of extracellular Na+ concentration, whereas uptake of 36Cl- is linear. Furthermore, neither 4,4-diisothiocyanostilbene-2,2-disulfonic acid, which is an inhibitor of the Cl-/HCO3- exchanger, nor 2-(3,4-dichlorobenzyl)-5-nitrobenzoic acid, which is a Cl- -channel blocker, affect either blastocoel expansion or 36Cl- uptake. These results suggest that Na+ entry into the mouse blastocyst is carrier-mediated and probably involves several routes that include the Na+/H+ exchanger and possibly the Na+-channel. Chloride entry, however, may not be carrier-mediated and may occur through a paracellular route, i.e., between the trophectodermal cells.

Topics: 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; Adenosine Triphosphate; Amiloride; Animals; Biological Transport; Blastocyst; Carrier Proteins; Chloride Channels; Chloride-Bicarbonate Antiporters; Chlorides; Cytochalasin D; Cytochalasins; Ectoderm; Epithelium; Kinetics; Membrane Proteins; Mice; Nitrobenzoates; Potassium; Sodium; Sodium Channels; Sodium-Hydrogen Exchangers