Recent biophysical investigations of vertebrate olfactory signal transduction have revealed that Ca2+-gated Cl- channels are activated during odorant detection in the chemosensory membrane of olfactory sensory neurons (OSNs). To understand the role of these channels in chemoelectrical signal transduction, it is necessary to know the Cl--equilibrium potential that determines direction and size of Cl- fluxes across the chemosensory membrane. We have measured Cl-, Na+, and K+ concentrations in ultrathin cryosections of rat olfactory epithelium, as well as relative element contents in isolated microsamples of olfactory mucus, using energy-dispersive x-ray microanalysis. Determination of the Cl- concentrations in dendritic knobs and olfactory mucus yielded an estimate of the Cl--equilibrium potential ECl in situ. With Cl- concentrations of 69 mM in dendritic knobs and 55 mM in olfactory mucus, we obtained an ECl value of +6 +/- 12 mV. This indicates that Ca2+-gated Cl- channels in olfactory cilia conduct inward currents in vivo carried by Cl- efflux into the mucus. Our results show that rat OSNs are among the few known types of neurons that maintain an elevated level of cytosolic Cl-. In these cells, activation of Cl- channels leads to depolarization of the membrane voltage and can induce electrical excitation. The depolarizing Cl- current in mammalian OSNs appears to contribute a major fraction to the receptor current and may sustain olfactory function in sweet-water animals.

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Parabolic refractive x-ray lenses with short focal distance can generate intensive hard x-ray microbeams with lateral extensions in the 100 nm range even at a short distance from a synchrotron radiation source. We have fabricated planar parabolic lenses made of silicon that have a focal distance in the range of a few millimeters at hard x-ray energies. In a crossed geometry, two lenses were used to generate a microbeam with a lateral size of 380 nm by 210 nm at 25 keV in a distance of 42 m from the synchrotron radiation source. Using diamond as the lens material, microbeams with a lateral size down to 20 nm and below are conceivable in the energy range from 10 to 100 keV.

Summary• Translocation is shown of phosphorus, nitrogen, potassium and magnesium to a P-deficient host from ectomycorrhizal fungal hyphae.• Mycorrhizal (with Paxillus involutus ) and nonmycorrhizal P-deficient spruce ( P. abies ) seedlings were grown in a two-compartment sand-culture system. Hyphal translocation of nutrients from the inner compartment (penetrated only by hyphae) to the host was measured using mass balance (for N, P and K) or stable isotope ( 15 N and 25 Mg) methods.• Addition of P to the hyphal compartment strongly stimulated hyphal growth, and this also increased both seedling P status and growth. Hyphae translocated nonlimiting elements in addition to P, contributing 52, 17, 5 and 3-4%, respectively, to total P, N, K or Mg plant uptake. The potential role of the ectomycorrhizal mycelium in K acquisition was demonstrated. Translocation to mycorrhizal seedings of N, K and Mg was strongly reduced when hyphal P-fluxes ceased; this translocation of nonlimiting nutrients depended on simultaneous translocation of P.• The ectomycorrhizal mycelium has an active role in P acquisition from sources not available to roots. Nutrient fluxes within fungal hyphae are interdependent and strong coupling of N, K and Mg fluxes with long-distance P translocation in the mycorrhizal mycelium occurs.

Yoshikami and Hagins first suggested that calcium is sequestered within membranous disks in the outer segments of vertebrate rods and that the bleaching of visual pigment molecules by light causes the release of Ca from the disks. Once released, the Ca was postulated to bind to Na+ channels or carrier molecules in the plasma membrane to produce the electrical response. This theory, termed the 'calcium hypothesis', is supported by much evidence but remains controversial, largely because of the difficulty in measuring calcium in rods and of demonstrating light-induced release. Here we describe direct measurements of total rod Ca using a new microprobe method, called laser micro-mass analysis, or LAMMA . Using this technique, we show that rods contain large amounts of Ca concentrated in their outer segments. Physiological levels of illumination produce a graded efflux of rod Ca content, amounting to about 10(4) ions per rhodopsin molecule bleached in dim light. As light does not change the rate of Ca influx, the total Ca content of the rod decreases. In bright light, as much as half the total Ca leaves the rod during only 1 min of illumination.

The entry of calcium and magnesium from external sources into mycorrhizal roots of 3-year-old Norway spruce trees (Picea abies [L.] Karst.) was monitored. Roots of intact plants were exposed for various periods of time, ranging from 2 min to 48 h, to nutrient solutions which contained the stable-isotope tracers 25Mg and 44Ca. After labelling, samples of roots were excised from the plants, shock-frozen, cryosubstituted and embedded. The resulting isotope composition in this material was analysed by a laser-microprobe-mass-analyser (LAMMA) at relevant positions within cross-sections of the roots. For both elements, we determined (i) the fractions of the isotopes originating from the plant prior to labelling, and (ii) the fraction of isotopes originating from the corresponding tracer that penetrated into the root. Both divalent cations rapidly penetrated across the cortical apoplast and reached the endodermis. After 2 min of exposure to the labelling solution, an initial transient gradient of the tracers could be observed within the root cortex. Subsequently, calcium as well as magnesium equilibrated between the apoplast of the entire cortex and the external tracer with a half-time, t1/2, of about 3 min. In contrast, the kinetics of radial movement into the vascular stele showed a delay with a t1/2 of 100-120 min. We take this as strong evidence that there exists a free apoplastic path for divalent cations in the cortex and that the endodermis is a major barrier to the further passage of Mg and Ca into the xylem. While 25Mg in the labelling solution exchanged rapidly with Mg in the cortical apoplast, the exchange across the plasma membrane with Mg present in the protoplasm of the same cortical cells was almost 2 orders of magnitude slower. The kinetics of Ca and Mg entry at + 6 degrees C were similar to those obtained at a root temperature of +22 degrees C.

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The apoplastic permeability of the fungal sheath of two different ectomycorrhizal associations of Pinus sylvestris L. was analysed by laser microprobe mass analysis (LAMMA) and energy-dispersive X-ray spectroscopy (EDXS) after stable isotope labelling with 25Mg, 41K and 44Ca. Entry of 25Mg and 44Ca into the outer cortical apoplast of non-mycorrhizal roots was detected after 4 min of labelling. After a longer exposure time the endodermis with its Casparian band acted as an efficient apoplastic diffusion barrier for the radial movement of 25Mg and 44Ca into the stele. A fraction of approximately one-third of the apoplastic cations of the root cortex could not be exchanged against the external label even after longer exposure times. The ectomycorrhizal sheath of the two fungal species used, Pisolithus tinctorius (Pers.) Coker & Couch and Suillus bovinus (L. ex Fr.) Kuntze, does not completely inhibit the apoplastic movement of ions into the mycorrhizal root cortex, but retarded the penetration of isotopes into the cortical apoplast. In roots inoculated with S. bovinus, a clear labelling of the cortical apoplast could first be detected after 24 h of exposure to the stable isotope solution. At this time the labelling of the cortical apoplast in these mycorrhizal roots was higher than those of non-mycorrhizal roots and, with EDXS, changes in the element composition of the apoplast were detected. The results indicated that possibly hydrophobins localized in the fungal cell wall might be involved in the increased hydrophobicity of mycorrhizal roots and the lower permeability of the ectomycorrhizal sheath.