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S J Kleene

Publications and source records attributed to S J Kleene.

25 records · Page 2Linked to original sources
PubMed1987

Comparative study of immature and mature olfactory receptor cells in adult frogs.

We describe a method to produce frog olfactory epithelium with a population of developmentally synchronized receptor cells suitable for electrophysiological studies. The epithelium is ablated with ZnSO4 and allowed to regenerate for 10 days. Generation of new cells is then blocked by continuous treatment with hydroxyurea. Since new receptor cells are generated beginning on the 6th day after ablation, it is reasonable to assume that receptor cells in the preparation originated between the 6th and 10th days following ablation. The age range of these cells would be no more than 5 days. The cells develop normally and in relative synchrony. Olfactory epithelia with developmentally synchronized receptor cells were used for a physiological study of response properties as a function of developmental age. Developmental stages of receptor cells were confirmed by fine-structure analysis. There were no differences in the polarities or shapes of electro-olfactograms (EOGs) recorded from olfactory epithelia composed of immature or mature receptor cells. However, amplitudes of the main negative components in EOGs recorded from epithelia composed of mature receptor cells were generally higher than those from epithelia composed of immature cells. The majority of immature olfactory receptor cells had low spontaneous activity or none at all. Mature olfactory receptor cells had a variety of frequencies of spontaneous activity ranging from less than one to more than 70 spikes/min. Extracellular single unit activity recordings showed that, contrary to what has been observed in rat embryos, olfactory receptor cells in regenerating epithelial of adult frogs do not go through a stage in which they respond to all odorants.

Animals↗
PubMed1986

Suppression of mitotic activity and synchronization of cell development in olfactory epithelium.

Prolonged continuous application of hydroxyurea to frog olfactory epithelium suppresses mitotic activity in this tissue without side effects. After ZnSO4-induced coagulation necrosis of the olfactory epithelium, application of hydroxyurea prevents the usual epithelial regeneration. A method for obtaining frog olfactory epithelium with a developmentally synchronized neuronal population is introduced. The epithelium is first ablated by perfusion of the nasal cavity with ZnSO4. Then the epithelium is allowed to regenerate for 6 days, after which continuous introduction into the nasal cavity of hydroxyurea suppresses mitotic activity and thus the generation of new cells in the tissue. Cells formed during the 6 days prior to hydroxyurea treatment continue to develop. We found that the olfactory receptor cells are generated beginning on the 6th day after ablation, and so the epithelia contain a highly synchronized population of neurons all generated within about 24 h.

Animals↗
PubMed1983

Dissociation of frog olfactory epithelium.

We report a method for producing cell suspensions from frog olfactory epithelium. The tissue is incubated for 45 min at room temperature in a solution which causes dissociation of the epithelium. The solution is an isotonic saline buffered to maintain a pH of 10.3 and a free Ca2+ ion concentration of 10(-6) M. The method uses no degradative enzymes, except for a brief DNase treatment. The resulting cell suspension contains single olfactory receptor neurons, sustentacular cells, glandular cells, and respiratory epithelial cells. The cells are viable as judged by vital staining, ciliary motility, and synthesis of RNA. Some types of cells lose their normal columnar shapes and become rounded in the suspension.

Animals↗
PubMed1981

Dissociation of frog olfactory epithelium with N-ethylmaleimide.

Treatment of frog olfactory epithelium with 8 mM N-ethylmaleimide for 2 min results in extensive dissociation of the epithelium. The resulting cell suspension contains single olfactory receptor neurons, sustentacular cells, respiratory epithelial cells, and cells of Bowman's glands. The cells in suspension exhibit the same morphologies seen in histological sections of intact epithelium.

Animals↗
PubMed1979

Attractants and repellents influence methylation and demethylation of methyl-accepting chemotaxis proteins in an extract of Escherichia coli.

During bacterial chemotaxis, attractants and repellents alter the methylation levels of the methyl-accepting chemotaxis proteins (MCPs). These methylation levels represent a balance between two enzymatic processes: methylation and demethylation. In vivo experiments previously have shown that chemoeffectors influence the demethylation process; effects on the methylation system have not been reported. Here we show that in a cell-free extract of Escherichia coli both methylation and demethylation of the MCPs are affected by attractants and repellents. Attractants enhance methylation and inhibit demethylation. Repellents inhibit methylation and stimulate demethylation. The cell-free system provides an opportunity for further study of the mechanisms by which attractants and repellents influence the levels of methylation of the MCPs.

Bacterial Proteins↗
PubMed1977

Isolation of glutamic acid methyl ester from an Escherichia coli membrane protein involved in chemotaxis.

We have isolated glutamic acid 5-methyl ester from an Escherichia coli protein that is involved in chemotaxis. The bacteria were first incubated with [methyl-3H]methionine under conditions which are known to result in methylation of the protein. The protein, isolated by gel electrophoresis, was then digested by successive treatment with three proteolytic enzymes. One of the products was [methyl-3H]glutamic acid 5-methyl ester, identified by comparison with an authentic sample in the following studies: (a) chromatography on an automatic amino acid analyzer, (b) chromatography on paper in two solvent systems, (c) chromatography on paper of the N-acetyl derivatives, and (d) stability of the ester bond to various pH conditions. No aspartic acid 4-methyl ester was found in the enzymatic digest. Treatment of the methylated protein with alkali released the radioactivity as [3H]methanol, which was identified by gas chromatography and by preparation of the 3,5-dinitrobenzoate.

Amino Acids↗