Odorant stimulation of secretory and neural processes in the salamander olfactory mucosa

Topical application of the odorants guaiacol (10(-3) mol/l, 1-30 min) and 2-isobutyl-3-methoxypyrazine (IBMP, 10(-5)-10(-3) mol/l, 15 min) caused time- and concentration-dependent reductions in the secretory granule content of acinar cells of the superficial Bowman's glands (sBG) and moderate to extensive vacuolation in acinar cells of sBG and deep olfactory glands (dG). Topical application of 9.8 mg/ml scopolamine 10 min before 10(-4) mol/l IBMP significantly reduced the amount of secretory granule depletion from sBG compared to that seen with IBMP alone and resulted in less extensive vacuolation in sBG and dG acinar cells. The i.p. injection of 42 mg/kg propranolol 10 min before topical application of 10(-4) mol/l IBMP had no effect on the action of IBMP. Guaiacol and IBMP also had time- and concentration-dependent effects on the secretory activity of sustentacular cells in the olfactory epithelium. The protrusion of secretory material into the mucociliary matrix that covers the epithelial surface and vacuolation within the secretory material resulted from odorant application. Scopolamine and propranolol had no effects on the action of IBMP on sustentacular cell secretory activity. When applied in the vapor phase, guaiacol elicited action potentials recorded from individual olfactory receptor neurons; the impulse frequency was concentration-dependent and showed tonic and phasic components when the duration of stimulation was varied. Low to moderate concentrations of IBMP delivered in the vapor phase evoked monophasic negative slow voltage transients recorded from the surface of the olfactory mucosa. The amplitudes of these transients increased with increasing stimulus concentrations. Higher concentrations or longer stimulus durations evoked longer-latency positive-voltage generating processes and negative afterpotentials. The properties of the electrophysiological responses to both odorants were characteristic of responses evoked by a wide variety of 'typical' odorants.     

Sugar-activated ion transport in canine lingual epithelium. Implications for sugar taste transduction

There is good evidence indicating that ion-transport pathways in the apical regions of lingual epithelial cells, including taste bud cells, may play a role in salt taste reception. In this article, we present evidence that, in the case of the dog, there also exists a sugar-activated ion-transport pathway that is linked to sugar taste transduction. Evidence was drawn from two parallel lines of experiments: (a) ion-transport studies on the isolated canine lingual epithelium, and (b) recordings from the canine chorda tympani. The results in vitro showed that both mono- and disaccharides in the mucosal bath stimulate a dose-dependent increase in the short-circuit current over the concentration range coincident with mammalian sugar taste responses. Transepithelial current evoked by glucose, fructose, or sucrose in either 30 mM NaCl or in Krebs-Henseleit buffer (K-H) was partially blocked by amiloride. Among current carriers activated by saccharides, the current response was greater with Na than with K. Ion flux measurements in K-H during stimulation with 3-O-methylglucose showed that the sugar-evoked current was due to an increase in the Na influx. Ouabain or amiloride reduced the sugar-evoked Na influx without effect on sugar transport as measured with tritiated 3-O-methylglucose. Amiloride inhibited the canine chorda tympani response to 0.5 M NaCl by 70-80% and the response to 0.5 M KCl by approximately 40%. This agreed with the percent inhibition by amiloride of the short-circuit current supported in vitro by NaCl and KCl. Amiloride also partially inhibited the chorda tympani responses to sucrose and to fructose. The results indicate that in the dog: (a) the ion transporter subserving Na taste also subserves part of the response to K, and (b) a sugar-activated, Na-preferring ion-transport system is one mechanism mediating sugar taste transduction. Results in the literature indicate a similar sweet taste mechanism for humans.     

The biochemical identification of fibronectin in the sea urchin embryo

We report the biochemical identification of fibronectin in the basal lamina of the sea urchin embryo. A. punctulata gastrula stage embryos were solubilized in Triton X-100 and the insoluble basal laminae extracted by incubation in buffer containing 8M urea, 2% 2-mercaptoethanol and 2% SDS. Extracted proteins were separated by SDS-PAGE, electrophoretically transferred to nitrocellulose filters and probed with monospecific antibodies directed against human plasma fibronectin (pFN). Incubation in 125I-labelled secondary antibody revealed a single band which co-migrates with human pFN at an apparent molecular weight of 220,000. This is the first direct biochemical demonstration of a fibronectin-like molecule in the sea urchin embryo which cross reacts with antibodies to vertebrate fibronectin.     

The identity of the current carriers in canine lingual epithelium in vitro

Ion transport across the lingual epithelium has been implicated as an early event in gustatory transduction. The fluxes of isotopically labelled Na+ and Cl- were measured across isolated canine dorsal lingual epithelium under short-circuit conditions. The epithelium actively absorbs Na+ and to a lesser extent actively secretes Cl-. Under symmetrical conditions with Krebs-Henseleit buffer on both sides, (1) Na+ absorption accounts for 46% of the short-circuit current (Isc); (2) there are two transcellular Na+ pathways, one amiloride-sensitive and one amiloride-insensitive; (3) ouabain, added to the serosal solution, inhibits both Isc and active Na+ absorption. When hyperosmotic (0.25 M) NaCl is placed in the mucosal bath, both Isc and Na+ absorption increase; net Na+ absorption is at least as much as Isc. Ion substitution studies indicate that the tissue may transport a variety of larger ions, though not as effectively as Na+ and Cl-. Thus we have shown that the lingual epithelium, like other epithelia of the gastrointestinal tract, actively transports ions. However, it is unusual both in its response to hyperosmotic solutions and in the variety of ions that support a transepithelial short-circuit current. Since sodium ion transport under hyperosmotic conditions has been shown to correlate well with the gustatory neural response, the variety of ions transported may likewise indicate a wider role for transport in taste transduction.     

Phylogeny of Greya (Lepidoptera: Prodoxidae), based on nucleotide sequence variation in mitochondrial cytochrome oxidase I and II: congruence with morphological data

The phylogeny of Greya Busck (Lepidoptera: Prodoxidae) was inferred from nucleotide sequence variation across a 765-bp region in the cytochrome oxidase I and II genes of the mitochondrial genome. Most parsimonious relationships of 25 haplotypes from 16 Greya species and two outgroup genera (Tetragma and Prodoxus) showed substantial congruence with the species relationships indicated by morphological variation. Differences between mitochondrial and morphological trees were found primarily in the positions of two species, G. variabilis and G. pectinifera, and in the branching order of the three major species groups in the genus. Conflicts between the data sets were examined by comparing levels of homoplasy in characters supporting alternative hypotheses. The phylogeny of Greya species suggests that host-plant association at the family level and larval feeding mode are conservative characters. Transition/transversion ratios estimated by reconstruction of nucleotide substitutions on the phylogeny had a range of 2.0-9.3, when different subsets of the phylogeny were used. The decline of this ratio with the increase in maximum sequence divergence among taxa indicates that transitions are masked by transversions along deeper internodes or long branches of the phylogeny. Among transitions, substitutions of A-->G and T-->C outnumbered their reciprocal substitutions by 2-6 times, presumably because of the approximately 4:1 (77%) A+T-bias in nucleotide base composition. Of all transversions, 73%-80% were A<-->T substitutions, 85% of which occurred at third positions of codons; these estimates did not decrease with an increase in maximum sequence divergence of taxa included in the analysis. The high frequency of A<-->T substitutions is either a reflection or an explanation of the 92% A+T bias at third codon positions.      

Cellular Concentrations and Uniformity of Cell-Type Accumulation of Two Lea Proteins in Cotton Embryos

The levels and cell-type distribution of late embryogenesis abundant (Lea) proteins D-7 and D-113 have been determined in mature cotton embryos by immunochemical methods. The two proteins were expressed in and purified from Escherichia coli and utilized for antibody production in rabbits. The antiserum to each protein was found to interact with all members of each protein family in cotton extracts by protein gel blotting. Using these antibodies in quantitative "rocket" immunoelectrophoreses, D-7 proteins were found to accumulate to ~8 x 1015 molecules per embryo, which is equivalent to ~109 molecules per "average cell." D-113 proteins accumulate to ~1016 molecules per embryo, which equates to ~1.3 x 109 molecules per average cell. These values calculate to concentrations of about 226 and 283 [mu]M, respectively, in the cell aqueous phase immediately prior to seed desiccation. In immunocytochemical studies using the fluorophor rhodamine linked to the secondary antibody, both proteins appeared to be evenly present in the cytosol of all cell types present in the embryo, including both cotyledon and axis epidermal cells. Thus, their function does not appear related to unique functions of specific cell or tissue types. The very high molar concentrations of the two proteins, coupled with their unusual predicted structure and their cytosol location, would seem to reduce the number of their conceivable functions.