Endogenous chloride channels of insect sf9 cells. Evidence for coordinated activity of small elementary channel units

The endogenous Cl- conductance of Spodoptera frugiperda (Sf9) cells was studied 20-35 h after plating out of either uninfected cells or cells infected by a baculovirus vector carrying the cloned beta-galactosidase gene (beta-Gal cells). With the cation Tris+ in the pipette and Na+ in the bath, the reversal potential of whole-cell currents was governed by the prevailing Cl- equilibrium potential and could be fitted by the Goldman-Hodgkin-Katz equation with similar permeabilities for uninfected and beta-Gal cells. In the frequency range 0.12 < f < 300 Hz, the power density spectrum of whole-cell Cl- currents could be fitted by three Lorentzians. Independent of membrane potential, >50% of the total variance of whole-cell current fluctuations was accounted for by the low frequency Lorentzian (fc = 0.40 +/- 0.03 Hz, n = 6). Single-Cl- channels showed complex gating kinetics with long lasting (seconds) openings interrupted by similar long closures. In the open state, channels exhibited fast burst-like closures. Since the patches normally contained more than a single channel, it was not possible to measure open and closed dwell-time distributions for comparing single-Cl- channel activity with the kinetic features of whole-cell currents. However, the power density spectrum of Cl- currents of cell-attached and excised outside-out patches contained both high and low frequency Lorentzian components, with the corner frequency of the slow component (fc = 0.40 +/- 0.02 Hz, n = 4) similar to that of whole-cell current fluctuations. Chloride channels exhibited multiple conductance states with similar Goldman-Hodgkin-Katz-type rectification. Single-channel permeabilities covered the range from approximately 0.6.10(-14) cm5/s to approximately 6.10(-14) cm3/s, corresponding to a limiting conductance (gamma 150/150) of approximately 3.5 pS and approximately 35 pS, respectively. All states reversed near the same membrane potential, and they exhibited similar halide ion selectivity, P1 > PCl approximately PBr. Accordingly, Cl- current amplitudes larger than current flow through the smallest channel unit resolved seem to result from simultaneous open/shut events of two or more channel units

Regulation of Cl- channels in normal and cystic fibrosis airway epithelial cells by extracellular ATP.

The rate of Cl- secretion by human airway epithelium is determined, in part, by apical cell membrane Cl- conductance. In cystic fibrosis airway epithelia, defective regulation of Cl- conductance decreases the capability to secrete Cl-. Here we report that extracytosolic ATP in the luminal bath of cultured human airway epithelia increased transepithelial Cl- secretion and apical membrane Cl- permeability. Single-channel studies in excised membrane patches revealed that ATP increased the open probability of outward rectifying Cl- channels. The latter effect occurs through a receptor mechanism that requires no identified soluble second messengers and is insensitive to probes of G protein function. These results demonstrate a mode of regulation of anion channels by binding ATP at the extracellular surface. Regulation of Cl- conductance by external ATP is preserved in cystic fibrosis airway epithelia

A Hubble Space Telescope Survey for Resolved Companions of Planetary-Nebula Nuclei

We report results of an HST "snapshot" survey aimed at finding resolved binary companions of the central stars of Galactic planetary nebulae (PNe). Using WF/PC and WFPC2, we searched the fields of 113 PNe for stars whose close proximity to the central star suggests a physical association. We find 10 binary nuclei that are very likely to be physically associated, and another six that are possible binary associations. By correcting for interstellar extinction and placing the central stars' companions on the main sequence, we derive distances to the objects, and thereby significantly increase the number of PNe with reliable distances. Comparison of our derived distances with those obtained from various statistical methods shows that all of the latter have systematically overestimated the distances, by factors ranging up to a factor of two or more. We show that this error is most likely due to the fact that the properties of our PNe with binary nuclei are systematically different from those of PNe used heretofore to calibrate statistical methods. Specifically, our PNe tend to have lower surface brightnesses at the same physical radius than the traditional calibration objects. This difference may arise from a selection effect: the PNe in our survey are typically nearby, old nebulae, whereas most of the objects that calibrate statistical techniques are low-latitude, high-surface-brightness, and more distant nebulae. As a result, the statistical methods that seem to work well with samples of distant PNe, e.g., those in the Galactic bulge or external galaxies, may not be applicable to the more diverse population of local PNe.Comment: 37 text pages, 17 table pages, 9 figures. Accepted by Astronomical Journal for June 1999 issu