An approach to the focal presentation of chemical stimuli

An olfactometer is described that presents temporally-discretepulses of stimuli to individual chemosensory structures. Thedevice is based on standard pressure injection techniques inwhich pulses of compressed air eject nanoliter volumes of upto six stimulants from small diameter glass micropipettes. Thedevice should be readily adaptable to different chemoreceptorpreparations.     

Stimulation of the hypothalamic-pituitary-adrenal axis and inhibition of growth hormone release via increased central noradrenaline neuronal activity by urethane anaesthesia in the rat: blockade by clonidine

Computerized gas chromatography-mass spectrometry was used to measure precisely the hypothalamic levels of noradrenaline (NA), dopamine and serotonin together with those of their major neuronal metabolites 3,4-dihydroxyphenylethyleneglycol (DHPG), 3,4-dihydroxyphenylacetic acid and 5-hydroxyindoleacetic acid in normal male rats 45 min after stimulation of hypothalamic-pituitary-adrenal function by urethane (1.3 g/kg) administration. Urethane treatment resulted in a significant elevation of central noradrenergic neuronal activity (NNA) as assessed from marked rises in hypothalamic DHPG concentrations and the ratio (DHPG/NA). At the same time there was significant stimulation of ACTH and corticosterone release and inhibition of growth hormone release. These hormonal and central effects of urethane (but not anesthesia) were inhibited when the alpha 2-agonist clonidine (150 micrograms/kg) was co-administered. Urethane had no major effect on hypothalamic dopamine or serotonin status. We propose that the release of ACTH and the suppression of growth hormone release following urethane anaesthesia is a result of activation of central NNA and suggest that the hormonal responses are mediated via hypothalamic noradrenergic facilitation of corticotrophin releasing factor and somatostatin release to the anterior pituitary.     

Experience of the use of probiotics for Salmonellae control in poultry

The Editors of Letters in Applied Microbiology will, at their discretion, publish invited and submitted 'Opinions' on subjects in the general area of Applied Microbiology. They will not be subjected to the normal refereeing procedures and reprints will not be provided. The 'Opinions' will not necessarily represent the views of the Society of Applied Bacteriology or of the Editors. The Editors may invite or readers may submit 'Responses' to published 'Opinions' , provided that they are not merely polemics, and these will also be published at the discretion of the Editors. 'Responses' will be treated precisely like 'Opinions' . They should clearly indicate, in their first sentence, the 'Opinion' to which they are a response.     

Mutations in the FAD-binding fold of alcohol oxidase from Hansenula polymorpha.

Alcohol oxidase of methylotrophic yeast is an FAD-containing enzyme. When in its active form, the enzyme is an octamer and located in the peroxisomes. To study the importance of FAD-binding on the activity, octamerization and intracellular localization of the enzyme, alcohol oxidase of Hansenula polymorpha was mutated in its presumed nucleotide-binding domain, which is formed by the N-terminal sequence. Whereas mutations of a glutamic acid residue (E42) reduced the stability of the octamer, it hardly affected enzyme activity and expression. However, replacements of three conserved glycines (G13, G15 and G18) and a conserved glutamic acid (E39) within the fold had severe effects. The mutations not only resulted in loss of enzyme activity but in reduced protein levels as well, probably due to decreased stability of the mutant alcohol oxidase. However, octamerization of the protein still occurred. The existence of inactive octameric proteins provides information about the formation pathway of this octameric flavoprotein.     

A trace display and editing program for data from fluorescence based sequencing machines.

'Ted' (Trace editor) is a graphical editor for sequence and trace data from automated fluorescence sequencing machines. It provides facilities for viewing sequence and trace data (in top or bottom strand orientation), for editing the base sequence, for automated or manual trimming of the head (vector) and tail (uncertain data) from the sequence, for vertical and horizontal trace scaling, for keeping a history of sequence editing, and for output of the edited sequence. Ted has been used extensively in the C.elegans genome sequencing project, both as a stand-alone program and integrated into the Staden sequence assembly package, and has greatly aided in the efficiency and accuracy of sequence editing. It runs in the X windows environment on Sun workstations and is available from the authors. Ted currently supports sequence and trace data from the ABI 373A and Pharmacia A.L.F. sequencers.     

Role of the nose in resistive load detection

Previous resistive load detection (RLD) studies have ignored the nose, the usual route of breathing. Weber's law predicts the delta R50 (the added load detectable on 50% of presentations) to be a fixed percent of the background resistance (R0) and thus the delta R50/R0 ratio (the Weber fraction) is constant. We have noted the nose to be sensitive to added load, we wondered if the nose might play a role in RLD. To determine whether this was true and to characterize the effects of changes in R0 in the range of normal nasal resistance (RN), we determined R0 and delta R50 using standard techniques under the following conditions: nose vs. decongested nose, nose vs. nose with added external R0 (3.0 and 8.0 cmH2O X l-1 X s), nose vs. anesthetized nose, nose vs. mouth, and mouth vs. mouth with added load (3 cmH2O X l-1 X s). We found that decongestant decreased RN [4.3 +/- 0.6 (SE) to 3.1 +/- 0.5 cmH2O X l-1 X s, P less than 0.05] and delta R50 (1.7 +/- 0.5 to 1.1 +/- 0.3 cmH2O X l-1 X s, P less than 0.05). When an external load of 3 cmH2O X l-1 X s was added to the nose, delta R50 did not change significantly (1.4 +/- 0.2 to 1.1 +/- 0.2 cmH2O X l-1 X s), but the Weber fraction decreased (0.28 +/- 0.05 to 0.15 +/- 0.03, P less than 0.02).(ABSTRACT TRUNCATED AT 250 WORDS)     

Ectonucleotidase Activities Associated with the Olfactory Organ of the Spiny Lobster

The olfactory system of the Florida spiny lobster, Panulirus argus, has olfactory receptors that are excited by the purine nucleotides AMP, ADP, and ATP. These receptors reside on chemosensory neurons that are contained within aesthetasc sensilla on the lateral filaments of the antennules. Also associated with the lobster's olfactory system are ectonucleotidase activities that dephosphorylate excitatory nucleotides, resulting in the production of the nonstimulatory nucleoside adenosine. Our studies of the 5'-ectonucleotidase, ecto-ADPase, and ecto-ATPase activities of this olfactory system showed that each activity was characterized by Michaelis-Menten kinetics; Michaelis constants ranged from 6.9 to 33.5 microM, and maximum velocities ranged from 2.5 to 28.8 fmol/sensillum/s. Evidence that AMP dephosphorylation may serve as an inactivation process was shown by the close correlation between the kinetics of 5'-ectonucleotidase activity and the periodicity of olfactory sampling. Decreased magnesium ion concentration or increased calcium ion concentration resulted in increased ecto-ATPase activity; this activity was insensitive to vanadate ion. Ectonucleotidase activities may have multiple effects on the detection of exogenous nucleotides by a chemosensory system. These effects can be either direct, such as the conversion of an odorant to an inactive compound, or indirect, such as the conversion of an odorant to another compound that can activate or inhibit either receptors or enzymes associated with the system.     

Ligand-regulated transport of the Menkes copper P-type ATPase efflux pump from the Golgi apparatus to the plasma membrane: a novel mechanism of regulated trafficking.

The Menkes P-type ATPase (MNK), encoded by the Menkes gene (MNK; ATP7A), is a transmembrane copper-translocating pump which is defective in the human disorder of copper metabolism, Menkes disease. Recent evidence that the MNK P-type ATPase has a role in copper efflux has come from studies using copper-resistant variants of cultured Chinese hamster ovary (CHO) cells. These variants have MNK gene amplification and consequently overexpress MNK, the extents of which correlate with the degree of elevated copper efflux. Here, we report on the localization of MNK in these copper-resistant CHO cells when cultured in different levels of copper. Immunofluorescence studies demonstrated that MNK is predominantly localized to the Golgi apparatus of cells in basal medium. In elevated copper conditions there was a rapid trafficking of MNK from the Golgi to the plasma membrane. This shift in steady-state distribution of MNK was reversible and not dependent on new protein synthesis. In media containing basal copper, MNK accumulated in cytoplasmic vesicles after treatment of cells with a variety of agents that inhibit endosomal recycling. We suggest that MNK continuously recycles between the Golgi and the plasma membrane and elevated copper shifts the steady-state distribution from the Golgi to the plasma membrane. These data reveal a novel system of regulated protein trafficking which ultimately leads to the efflux of an essential yet potentially toxic ligand, where the ligand itself appears directly and specifically to stimulate the trafficking of its own transporter.