Cation channel blocked by extracellular Ca2+ in the apical membrane of the chick embryonic ectoderm

Abstract

In the chick embryo (20 h incubation, gastrula stage), the apical membrane of the ectodermal cells shows a high density of a non-selective cation channel which is blocked by very low extracellular Ca2+ concentrations. Properties of this channel were studied at the single-channel level using the patch-clamp technique in the cell-attached mode. With 1 mmol/l Ca2+ in the pipette, only outward current was present and the channel conductance measured at +120 mV was 25.5 pS. In the absence of Ca2+, also inward current through the channel was observed. The conductances measured at -50 mV were 49.5 pS with Na+ as the charge carrier, 72.5 pS with K+, 49.1 pS with Cs+, and 18.5 pS with Li+. The conductance measured at +80 mV was around 23 pS in all four cases. The reversal potential was similar (around 25 mV) for all four ions, which indicates a poor selectivity of the channel. In the absence of Ca2+ and the presence of 1 mmol/l ethylenebis(oxonitrilo)tetraacetate (EGTA), the kinetics of the channel were characterized by bursts of the order of seconds. During a burst, the channel flickered between one open and one closed level. The open time was constant between -30 mV and -80 mV, while the closed time decreased with hyperpolarization. The open time varied according to the permeant ion (K+ < Na+ = Cs+ < Li+). Extracellular Ca2+ blocked the inward current in a voltage-dependent manner. The Kd values, 1 mumol/l at -30 mV and 3.2 mumol/l at -80 mV, indicate that Ca2+ ions exit the channel toward the intracellular side. A weak voltage dependency of the association rate constant suggests that the Ca(2+)-binding site is close to the outside mouth. Extracellular Ca2+ was much less efficient at blocking the outward current (Kd about 1 mmol/l at 80 mV). Tetracaine, but not uraniumdioxide, decreased the opening probability of the channel. The embryonic channel shows similarities with the Ca(2+)-blockable, poorly selective channel described in the epithelium of toad urinary bladder.

Extracellular divalent cations block a cation non‐selective conductance unrelated to calcium channels in rat cardiac muscle

“…In amphibian epithelia, a channel with characteristics similar to those of the conductance described here has been detected using noise analysis and patch-clamp techniques (Van Driessche & Zeiske, 1985;Li et al 1994). The epithelial channel was permeable to a variety of monovalent cations (K¤, Rb¤, Cs¤, Tl¤, Li¤, NHÚ¤) and could be blocked by Ca¥, Ba¥, Cd¥ or Mg¥ as well as by the trivalent LaŤ (see Van Driessche et al 1988).…”

Permeation properties of a Ca(2+)-blockable monovalent cation channel in the ectoderm of the chick embryo: pore size and multioccupancy probed with organic cations and Ca2+.

“…In cell-attached patches, millimolar concentrations of Ca ~+ in the extracellular solution blocked the inward current completely. In these almost physiological conditions, the CMC current was strongly outwardly rectifying (Li et al, 1994). In the absence of extracellular Ca 2+, the single-channel inward current was larger than the outward current ( Fig.…”

“…1 A). The current of the active periods was interrupted by shortclosures of unknown origin (Li et al, 1994). Histograms of both active and silent times collected from several cell-attached patch experiments have monoexponential distributions ( Fig.…”

“…The pipette solutions included usually 5 mM EDTA and the appropriate amount of cation chloride. The Ca2+concentradons were calculated as described in Li et al, 1994. The osmolarity of the low-Na + solutions (5--70 mM) used to estimate the channel affinity for Na § and the surface potential of the Ca2+-blockable monovalent cation (CMC) channel was maintained at ~300 mOsm with the appropriate amount of sucrose.…”

“…The reversal potentials can be used to determine the permeabilities of the organic cations relative to intracellular K + (see Methods). As (Li et al, 1994) are shown for comparison. The dotted line represents the theoretical curve expected for a pore radius of 2.9 and a K + radius of 2.0.…”