Kallidin applied to the human nasal mucosa produces algesic response not blocked by capsaicin desensitization.

Various kinins (dissolved in 50 microliters) were applied to the nasal mucosa of healthy human volunteers to test the algesic and proinflammatory effects of these peptides in an intact human tissue. [des-Arg9]-bradykinin (0.5 mumol) was found to be inactive, while bradykinin (0.05-0.5 mumol) and especially kallidin (0.005-0.5 mumol) induced: (a) a mild painful sensation described as burning and pricking (latency 30 s, duration 3-5 min), (b) perception of pulsatility and obstruction in the nasal cavity (onset 1 min, duration 6-8 min). Substance P (0.5 mumol) and neurokinin A (0.5 mumol) produced slight obstruction and weak pulsatile sensation but not pain. Capsaicin (0.05 nmol) produced pain and secretion of fluid, but not pulsatile sensation. The effects of kallidin were not affected by repeated (to induce desensitization) applications of capsaicin (0.5 mumol). Likewise, ipratropium bromide (80 mg in 100 microliters) did not affect responses to kallidin. In an intact human tissue, kallidin produces various effects, including an algesic response, that are apparently independent from activation of B1 receptors and from desensitization of capsaicin-sensitive primary afferents.     

Cell surface contact mediates neuronal recognition and synapse formation between two identified leech neurons

An early event in the formation of the serotonergic synapse by the Retzius (R) onto the pressure-sensitive (P) neurons of the leech is the elimination of an extrasynaptic response to transmitter from sites of contact on the postsynaptic cell. This event during synapse formation is cell-specific in that it is elicited in vitro by contact with the presynaptic R cell but not with other neurons. In the study reported here, we investigated the nature of this interaction between R and P neurons. The loss of the extrasynaptic response of the P cell was elicited by contact with R cells fixed in a mild paraformaldehyde solution, but not by R cells treated with the proteolytic enzyme trypsin prior to fixation. As well, a variety of lectins were assayed for their ability to interfere with synapse formation. The transmitter responses of P cells plated on lectin-coated substrates were unaffected. However, exposure of the R cell to the lectin wheat germ agglutinin (WGA), but not to other lectins, prior to pairing prevented the loss of the extrasynaptic response in contacted P cells and blocked the formation of the R? P synapse in culture. We conclude that recognition by the P cell of the R cell during synapse formation may be mediated by an R cell-specific surface protein which binds wheat germ agglutinin. 1994 John Wiley & Sons, Inc.     

Sensitive fluorescence in situ hybridization signal detection in maize using directly labeled probes produced by high concentration DNA polymerase nick translation

The signal produced by fluorescence in situ hybridization (FISH) often is inconsistent among cells and sensitivity is low. Small DNA targets on the chromatin are difficult to detect. We report here an improved nick translation procedure for Texas red and Alexa Fluor 488 direct labeling of FISH probes. Brighter probes can be obtained by adding excess DNA polymerase I. Using such probes, a 30 kb yeast transgene, and the rp1, rp3 and zein multigene clusters were clearly detected.     

Identification, cloning, and characterization of rcsF, a new regulator gene for exopolysaccharide synthesis that suppresses the division mutation ftsZ84 in Escherichia coli K-12.

A new gene, designated rcsF, was located adjacent to drpA at the 5.2-min position of the genetic map of Escherichia coli. The deduced amino acid sequence encoded by the rcsF gene indicates a small protein of 133 amino acid residues with a calculated pI of 10.8 that is rich in proline, serine, alanine, and cysteine residues. When overexpressed as a result of its presence on a multicopy plasmid, rcsF confers a mucoid phenotype and restores colony formation to ftsZ84 mutant cells on L agar medium containing no added NaCl. These two phenotypes are not observed in rcsB mutant cells. Ion mutant cells harboring an rcsF mutation accumulate considerably lower levels of exopolysaccharides, whereas the presence of a multicopy rcsF plasmid not only increases capsule synthesis but also confers a mucoid phenotype at 37 degrees C, a temperature at which ion mutant cells are known not to form mucoid colonies. RcsF does not stimulate the expression of rcsB, indicating that it exerts its action through the RcsB protein, possibly by phosphorylation. It is also shown that RcsF stimulation of capsule synthesis is RcsA-dependent, whereas colony formation of ftsZ84 mutant cells can be restored by RcsF in the absence of RcsA.     

In‐line monitoring of amino acids in mammalian cell cultures using raman spectroscopy and multivariate chemometrics models

The application of PAT for in‐line monitoring of biopharmaceutical manufacturing operations has a central role in developing more robust and consistent processes. Various spectroscopic techniques have been applied for collecting real‐time data from cell culture processes. Among these, Raman spectroscopy has been shown to have advantages over other spectroscopic techniques, especially in aqueous culture solutions. Measurements of several process parameters such as glucose, lactate, glutamine, glutamate, ammonium, osmolality and VCD using Raman‐based chemometrics models have been reported in literature. The application of Raman spectroscopy, coupled with calibration models for amino acid measurement in cell cultures, has been assessed. The developed models cover four amino acids important for cell growth and production: tyrosine, tryptophan, phenylalanine and methionine. The chemometrics models based on Raman spectroscopy data demonstrate the significant potential for the quantification of tyrosine, tryptophan and phenylalanine. The model for methionine would have to be further refined to improve quantification.     

Synapse formation and function: Insights from identified leech neurons in culture

Identified leech neurons in culture are providing novel insights to the signals underlying synapse formation and function. Identified neurons from the central nervous system of the leech can be removed individually and plated in culture, where they retain their characteristic physiological properties, grow neurites, and form specific synapses that are directly accessible by a variety of approaches. Synapses between cultured neurons can be chemical or electrical (either rectifying or not) or may not form, depending on the neuronal identities. Furthermore, the characteristics of these synapses depend on the regions of the cells that come into contact. The formation and physiology of synapses between the Retzius cell and its partners have been well characterized. Retzius cells form purely chemical, inhibitory synapses with pressuresensitive (P) cells where serotonin (5-HT) is the transmitter. Retzius cells synthesize 5-HT, which is stored in vesicles that recycle after 5-HT is secreted on stimulation. The release of 5-HT is quantal, calcium-dependent, and shows activity-dependent facilitation and depression. Anterograde and retrograde signals during synapse formation modify calcium currents, responses to 5-HT, and neurite outgrowth. The nature of these synaptogenic signals is being elucidated. For example, contact specifically with Retzius cells induces a localized selection of transmitter responses in postsynaptic P cells. This effect is signaled by tyrosine phosphorylation prior to synapse formation. © 1995 John Wiley & Sons, Inc.     

Growth kinetics of Dioscorea deltoidea and Catharanthus roseus in batch culture

Sugar-limited batch growth of Dioscorea deltoidea and Catharanthus roseus plant cell cultures was studied in a 14-L stirred tank fermentor. With dissolved oxygen concentration monitored and maintained at nonlimiting levels, growth rates and ratios of dry weight to fresh weight were found to be strongly influenced by sugar concentration. Linear relationships between respiration rate and growth rate were observed, and respiration rate was found to drop to a maintenance level after sugar had been fully depleted from the medium. Diosgenin biosynthesis by D. deltoidea was shown to be independent of growth rate. Ajmalicine biosynthesis in C. roseus was negligible during sugar-limited growth but was induced by inoculation into an 80-g/L glucose solution.