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Errors affecting the estimation of the ionic product of water from the potentiometric titration of weak acids have been analysed. A Fortran programme has been developed in order to simulate experimental titration data sets. This programme allows one to isolate experimental error sources and analyse their effect upon the final fitted equilibrium constant. pKw dependence with titration final added volume and the pK value of the weak acid, predicted with the simulation programme are coincident with experimental data, suggesting that obtaining reliable pKw values entails using experimental data of titrations up to a sufficiently large hydroxyl concentration which depends on the value of pK.

Introduction: The mechanisms used by the cerebral cortex to modulate the cutaneous information at prethalamic level have been scarcely studied. This article reviews experimental evidence leading to a better understanding of this issue at the level of the cuneate nucleus (Burdach nucleus)., Development: The primary afferents and the corticocuneate fibers make synaptic contact with cuneothalamic neurons and with inhibitory interneurons in the middle cuneate nucleus. By stimulating the skin at different places while recording the cuneothalamic intracellular activity in anaesthetized animals with the cortex intact, with the cortex pharmacologically inactivated, or in absence of a cerebral cortex it was possible to ascertain the functional role of the corticocuneate fibers. The primary afferents activated by stimulating a particular zone of the skin induce monosynaptic excitation on a group of cuneothalamic cells at the same time at which inhibit, through intranuclear interneurons, neighboring cuneothalamic cells with unmatched receptive fields. Similarly, the corticocuneate cells receiving information from the stimulated skin further increase the excitation of the cuneothalamic neurons with matched receptive fields while inhibiting others with unmatched fields. The cortex exaggerates an excited center surrounded by an inhibited periphery thus increasing the tactile discrimination both spatially and temporally which is essential for exploratory and manipulative purposes.

Reaction of cyclometalated halide-bridged Pd(II) complexes 1-4 with the tertiary triphosphine ligand (Ph2PCH2CH2)2PPh (triphos) yielded complexes [((Ph2PCH2CH2)2PPh-P,P,P)Pd(N(Cy)=(H)C)C6H2(C(H)=N(Cy))Pd((Ph2PCH2CH2)2PPh-P,P,P)][ClO4]2 5, [Pd(C6H4-N=NC6H5)((Ph2PCH2CH2)2PPh-P,P,P)][ClO4] 6, and [Pd(R-C6H3C(H)=NCy)((Ph2PCH2CH2)2PPh-P,P,P)][ClO4] (7; R = 4-CHO, 8; 3-CHO). Spectroscopic and analytic data suggest five-coordination on the palladium atom, which, for complexes 5, 6, and 7, was confirmed by X-ray crystallography. The geometry around palladium may be view as a distorted trigonal bipyramid, with the palladium, nitrogen, and terminal phosphorus atoms in the equatorial plane. Compound 5 is the first doubly cyclometalated palladium(II) compound with two pentacoordinated metal centers. The structure of 6 comprises two discrete cations with slightly different geometries, showing the importance of crystal packing forces in order to determine the coordination arrangement.

Previous research on electrodermal conditioning suggests that the conditioned diminution of the unconditioned response (UR) has an associative basis. The aim of this experiment was to test whether this phenomenon also occurs in heart rate (HR) classical conditioning. For this purpose, a differential classical conditioning was performed. The conditioned stimuli (CSs) were geometrical shapes (the CS+ was a square and the CS- was a triangle) displayed on a computer screen and a burst of white noise was used as unconditioned stimulus (US). For analysis of the conditioned response (CR) components, an interval between CS+ and US of 8 seconds was used. After the acquisition phase, participants were tested using trials with the US preceded either by a CS+, a CS-, or a neutral stimulus (a circle). The results showed conditioned diminution of the UR and suggest that the second heart rate deceleration component (D2) is responsible for the occurrence of this phenomenon.

A series of 1H-pyrazolo[3,4-d]pyrimidines (3--6) substituted at positions 1 (R(1)=Ph, H, tert-butyl and ribosetribenzoate), 4 (R(2)=chlorine, nitrogen and oxygen nucleophiles), and 6 (dimethylamino) have been synthesized and their effect on the release of histamine from rat peritoneal mast cells measured. After chemical stimulation, (polymer 48/80), several compounds (i.e. 3b, 4a, 4b, 4d, 4g, 5a), produce inhibition two to three times higher (40--60%) than DSCG but this action is lower after preincubation. 4b (R(1)=Ph, R(2)=NHCH(2)Ph; 50--70% inhibition) and 5a (R(1)=H, R(2)=OMe; 50--55% inhibition) are the most active ones in both experiments. With ovoalbumin as stimulus, several pyrazolopyrimidines show inhibition similar to DSCG, the most active compounds being 6a--d (IC(50)=12--16 microM; R(1)=ribosetribenzoate, R(2)=methoxy and amino). Compounds 4e (R(1)=t-butyl, R(2)=OMe) and 4g (R(1)=t-butyl, R(2)=piperidino) are inducers of the release of histamine (60 and 150% increase). Compounds 4b and 4c showed cytotoxic activity (IC(50)=1 microg/mL) to HT-29 human colon cancer cells.

The DNA content of 10 species of bivalve molluscs from British Columbia coast was determined by image analysis, and the karyotypes of the horse clam Tressus capax, the bent-nose macoma Macoma nasuta, and the nuttall's mahogany clam Nuttallia nuttallii are described here for the first time. We also have analyzed the location of rDNA loci using a 28S-5.8S-18S probe in four of these species: Mytilus californianus, M. trossulus, Macoma nasuta and N. nuttallii. Results obtained report new data about cytogenetic characteristics of bivalve molluscs.

Hap1 and Rox1 are transcriptional regulators that bind regulatory sites in the promoters of oxygen-regulated genes in Saccharomyces cerevisiae. Hap1 is a heme-responsive activator of genes induced in aerobic conditions and Rox1 is a repressor of hypoxic genes in aerobic conditions. We have studied transcriptional regulation of a pool of 203 open reading frames (ORFs) from chromosomes IV, VII, and XIV in wild-type, hap1, and rox1 mutant genetic backgrounds in an attempt to extend the family of oxygen and heme regulated genes. Only three ORFs are significantly repressed by Rox1 but they cannot be considered as typical hypoxic genes because they are not overexpressed during hypoxia.

Yeasts do not form a homogeneous group as far as energy-yielding metabolism is concerned and the fate of pyruvate, a glycolytic intermediate, determines the type of energy metabolism. Kluyveromyces lactis has become an alternative to the traditional yeast Saccharomyces cerevisiae owing to its industrial applications as well as to studies on mitochondrial respiration. In this review we summarize the current knowdeledge about the K. lactis respirofermentative metabolism, taking into account the respiratory capacity of this yeast and the molecular mechanisms controlling its regulation, giving an up-to-date picture.

This research tested a multicomponent self-help manual that contained nicotine fading and some behavioral techniques. It also evaluated the incremental effects of using higher amounts of therapist contact on quitting rates. 114 smokers were randomly assigned to one of four treatment groups: (1) A 10-session multicomponent program (n = 25); (2) a 5-session multicomponent program (n = 31); (3) a 5-session multicomponent plus a self-help manual program (n = 25); and (4) a self-help-manual-only program (n = 33). A control group (n = 48) did not receive any treatment. The distinctive characteristic of the treatments was the different amount of therapist contact. Common components of the programs were a refundable deposit, self-monitoring, information on smoking, stimulus control, CO feedback, nicotine fading, and strategies to avoid withdrawal symptoms. End-of-treatment quit rates ranged from 36% (self-help manual) to 68% (10-session multicomponent program). At 12-month follow-up there were significant differences between groups, and the most effective group was the 5-session plus manual group, with an abstinence rate of 48%.

Background: The role of enterovirus infection in the pathogenesis of dilated cardiomyopathy (DCM) remains unclear. The objective of this study was to determine the prevalence of enterovirus in hearts explanted from patients with DCM and to compare it with enterovirus prevalence in hearts explanted from patients with other etiologies and in healthy donor hearts., Methods: A total of 138 cardiac samples were analyzed, 70 from heart donors and 68 from transplant recipients (22 with DCM). A highly sensitive enterovirus-specific nested RT-PCR was used to test for enterovirus., Results: All tests were negative except for one positive result that was attributed to carryover because sequencing of the amplification product showed it to be identical to the positive control., Conclusions: In this study the sample of explanted hearts nested RT-PCR showed no evidence of the presence of enteroviral RNA. This suggests that if enterovirus had a role in the genesis of DCM, it does not require or lead to the persistence of the virus in myocardial tissue.

A series of 8-cyanopyrido[3',2':4,5]thieno[3,2-d]-1,2,3-triazines, substituted at C-4 and C-7, were synthesized and evaluated as nitric oxide and prostaglandin E(2) inhibitors in murine peritoneal macrophages stimulated with bacterial endotoxin. Several compounds exhibited considerable activity, compounds 10 and 13 being the most interesting ones with IC(50) values of 11.2 and 3.4 microM on nitrites and 0.9 and 0.6 microM on prostaglandin E(2) production, respectively. None of the examples of pyridothienotriazines that were active at 10 microM showed any effect on inducible nitric oxide synthase, cyclooxygenase-2, and cyclooxygenase-1 enzymes, suggesting that they act by modifiying the level of expression of these inducible enzymes.

Sequence analysis of the Kluyveromyces lactis HIS4 (KlHIS4) gene promoter reveals relevant differences in comparison to the Saccharomyces cerevisiae HIS4 homologous gene. Among them are the absence of a Rap1 binding site and the presence of only three putative Gcn4 binding consensus sites instead of the five described in the S. cerevisiae promoter. Since these factors are implicated in the general control, we investigated the transcriptional regulation of the KlHIS4 gene under conditions of amino acid starvation and discovered that the mechanisms previously described for S. cerevisiae HIS4 regulation and related to general control are not functional in K. lactis. The expression analysis of the KlHIS4 gene under phosphate starvation or high adenine supply shows that factors, such as Bas1 or Bas2, involved in the basal control may also operate in a different way in K. lactis. Interestingly, and also in contrast to the HIS4 regulation in S. cerevisiae, we found domains for Nit2-like and yeast-Ap1-like binding sequences. Northern analyses showed transcriptional activation under ammonia starvation and oxidative stress.

The purpose of this experiment was to study whether interstimulus interval (ISI) modification differentially affects the amplitude of the unconditioned response (UR amp.). Seventy-five volunteer subjects received discrimination training with an interval between conditioned stimulus (CS) and unconditioned stimulus (US; aversive white noise) of 8 s. After the discrimination training phase, subjects exhibiting discriminative control were randomized into three groups that differed according to the ISI, of 1, 5 or 8 s. The subjects of each group were then tested with five presentations of CS + /US. The results demonstrate that the UR showed a greater amplitude when an ISI of 8 s was used than with ISIs of 1 or 5 s. No significant differences were found between the UR amp. of the 1- and 5-s ISI groups. These findings are discussed as a result of a hypothetical experimentally induced blend of the anticipatory CRs with the UR.

The purpose of this experiment was to study whether conditioned diminution of the unconditioned response (UR) is a phenomenon with an associative basis. Discriminative electrodermal conditioning was used with an interval between the conditioned stimulus (CS) and unconditioned stimulus (US; aversive white-noise) of 8 s. Fifty-nine volunteer subjects received discrimination training in which one CS was reinforced (CS+ /US) and a second CS was non-reinforced (CS-). After this discriminative training phase, participants were tested using intermixed trials in which a US was preceded by either a CS+, a CS-, or a neutral stimulus (NS). The results indicated that the skin conductance response amplitude of the UR was lower when the US was preceded by the CS+ than when the US was preceded by the CS- or the NS. However, NS/US presentations elicited URs of greater amplitude than those of the CS- /US presentations. The results can be explained in terms of orienting reflex reinstatement. In addition, it is argued that conditioned diminution has an associative basis.

Six open reading frames (ORFs) from chromosome, VII, YGL131c, YGL129c, YGL128c, YGL125w, YGL124c and YGL121c, were disrupted by deletion cassettes with short flanking regions homologous to the target locus (SFH). YGL129c is necessary for growth in non-fermentable carbon sources, YGL128c for growth at low or high temperatures and YGL125w is implicated in the biosynthesis of methionine. With regard to the other ORFs, basic phenotypic analyses did not reveal any significant clues about their function.

The synthesis of a series of pyridothienopyrimidines and their evaluation as inhibitors or inducers of the release of histamine from rat mast cells is reported. The activity was measured after immunological stimulation with ovoalbumin and chemical stimulation with polymer 48/80 and the drugs adryamicin and vinorelbine. The experiments were carried out with and without preincubation of the stimulus with the cells before addition of the drug. Several pyridothienopyrimidines show inhibitory IC50 values in the range 2-25 microM, indicating they are up to 100 times more potent than cromoglycate (DSCG) and 10 times greater than Ketotifen. Compound 9l is a potent inhibitor in all the conditions tested and shows IC50 = 9-25 microM. Pyridothienopyrimidines 4l and 9e are very strong inducers of histamine release in the immunological (4l, 170-230%) and chemical (9e, 100-150%) assays, respectively. Compounds 4l and 9i are cytotoxic in vitro (IC50 = 0.1-0.2 microgram/mL) against P-388, A-549, HT-29, and MEL-28 tumor cell lines.

Functional analysis of the KlCYC1 promoter reveals that sequences located upstream to those already published [Freire-Picos, M. A., Rodríguez-Torres, A. M., Ramil, E., Cerdán, M. E., Breuning, K. D., Hollenberg, C. P. & Zitomer, R. S. (1993) Sequence of a cytochrome c from Kluyveromyces lactis and its upstream region, Yeast 9, 201-204] and extending from positions -780 to -371 are important for maintaining high levels of expression, although this region contains both negative and positive elements. A consensus sequence for interaction with KlCpf1p is present at position -492, into the negative site, and specific protein binding to KlCpf1p has been demonstrated. Deletion of the sequences from positions -413 to -338 diminishes KlCYC1 transcription; protein binding to two sequences included in this activator region is detected and several points of evidence indicate that the complex observed is different from the Hap2/3/4/5p complex. Binding of KlCpf1p and the activator complex to the promoter is constitutive in different carbon sources. Although the promoter contains CCAAT boxes, directed mutagenesis has revealed that they are not related to the moderate de-repression observed in glycerol media.

We examined the variation with ionic strength (I, adjusted with KCl, KNO(3), KBr, NaCl or NaClO(4)) of the formal potential (E(const)) for glass electrodes exhibiting a Nernstian response (i.e. E(cell)=E(const)-slog[H(+)]). For this purpose, we investigated the different factors included in the formal potential, so we obtained reported values for the liquid junction potential as a function of ionic strength and determined the logarithm of the activity coefficient for the proton in various saline media, using Pitzer equations.

The study of changes in ERPs provoked by negative hypnotic hallucinations has thus far yielded contradictory results. Most previous studies have failed to separate specific changes in the processing of the hallucinated stimuli from non-specific changes in arousal due to hypnosis. The present study addresses this issue with the combination of two experimental effects in the task designed to test the hallucination: the noise-compatibility effect and the Simon effect. A choice reaction task was used in which targets appearing at either side of a screen were defined by color and were accompanied by irrelevant noise, and a negative hallucination suggestion was given for noise stimuli. Four highly hypnotizable subjects performed the task on two separate occasions, with and without the suggestion. Increases in reaction times and P300 latencies were found as a function of noise and spatial stimulus-response (SR) incompatibility (Simon effect). Suggestion decreased the response times for all types of trials non-significantly, and it decreased significantly those of left-hand responses. On the other hand, suggestion reduced the increase in P300 latencies in noise-incompatible trials, but did not influence the Simon effect. This result indicates a specific effect of suggestion in the processing of hallucinated stimuli, which is consistent with the hallucinatory experience reported by the subjects.

The Kluyveromyces lactis HIS4 gene was cloned by complementation of a Saccharomyces cerevisiae his4 mutant. Sequence analysis revealed a 2388 bp open reading frame encoding a single polypeptide predicted to encompass three distinct enzymatic activities (phosphoribosyl-AMP cyclohydrolase, phosphoribosyl-ATP pyrophosphohydrolase and histidinol dehydrogenase). This structural organization is strikingly similar to that of the His4 proteins from S. cerevisiae and Pichia pastoris. Transcript analysis detected a single mRNA species of 2.5 kb.

The sequence and characterization of the KlGSK-3 gene in chromosome VI [corrected] from Kluyveromyces lactis is presented. The deduced amino-acid sequence predicts a protein of 415 amino acids and an Mr of 47 kDa. A computer search reveals significant homology to serine/threonine protein kinases closely related to members of the GSK-3 subfamily. The Klgsk-3::URA3 disrupted strain is unable to grow in glucose at 37 C but KlGSK-3 is not essential for vegetative growth at 30 C or 14 C. The KlGSK-3 gene presents the highest homology with the Saccharomyces cerevisiae MDS1 gene. Expression studies show an increase of mRNA levels caused both by carbon starvation and when diploids are shifted from rich to sporulation media. The data reported show that KlGSK-3, like MCK1 from S. cerevisiae, is related to glycogen storage.

To characterize honey types, a citric acid determination may be useful. A citric acid determination on honey was carried out with previous polyvinylpolypyrrolidone (PVPP) clarification followed by the Boehringer-Mannheim GmbH enzymatic test. The sample solution was prepared from 2 g of honey in 100 mL of Milli-Q water. A volume of 10 mL of this sample was clarified with PVPP stirring for 1 min and filtered. The enzymatic determination was measured spectrophotometrically at 340 nm, using citrate lyase, L-malate dehydrogenase, and L-lactate dehydrogenase. With these conditions, there were no observed interference effects. The proposed method improves precision [coefficient of variation (CV) between 0.26% and 1.60%] and recovery (between 98.0% and 100.9%) on the direct enzymatic analysis (% CV between 1.02 and 2.66 and recovery between 84.0% and 115.6%). Furthermore, the cost was reduced 70% using a microtest. The method was applied to 20 honeys of Galicia (northwestern Spain), and the results ranged between 44.2 and 827.0 mg of citric acid/kg of honey (mean = 192.9 mg/kg).

Background: To determinate the viral variables (viremia and genotype) and patients variables (route and years of infection, histology, etc) related with response to the treatment with alpha intefferon in patients with chronic hepatitis C., Subjects and Methods: We have studied 50 patients with chronic hepatitis C that received an intefferon treatment during a year. In all them it was accomplished a hepatic biopsy before of the beginning of treatment and it was studied the viral load by quantitative PCR (Monitor, Roche) at the beginning of treatment, at four weeks and at the end of treatment. It was determined the HCV genotype by method INNO-LIPA (Boehringer Inghelheim)., Results: 8 patients (16%) achieved a complete sustained response during all the follow-up period (13.8 + 1.6 months); 8 patients responded but relapsed when the intefferon was interrupted and 34 patients not responded. The responders had lower viremia (p < 0.032) and HCV RNA negative at the fourth treatment week (p < 0.0082). The genotype 1b was the most frequent, the one which was presenting a smaller percentage of response and it was associated with a higher viremia. Cirrhotic patients were the worse responders. Ex-lDAs were better responders than the others groups, however this difference did not show statistics meaning., Conclusions: A low pretreatment HCV-RNA level seems to be indicative of a sustained response to IFN, Whereas a high level seems to be indicative of a non sustained response to IFN. Negativization of HCV-RNA at week four is a good indicator of response to intefferon. Genotype 1b is the most frequent and the worse responder. To have 1b genotype, high pretreatment HCV-RNA level and to be viremic at four treatment seems to be indicative of non response.

The octavolateral area of lampreys, which receives primary fibers from the octaval and lateral line nerves, is involved in the premotor organization of body movements through secondary projections to the reticular formation. Here, the typology of neurons of the three octavolateral nuclei (ventral, medial, and dorsal) that putatively project to the middle and posterior rhombencephalic reticular nuclei were studied by retrograde transport of horseradish peroxidase (HRP) applied to these reticular nuclei. Several types of neurons were labeled in the ventral nucleus, both ipsilateral and contralateral to the site of HRP application. Some of these neurons showed a rather simple morphology (octavomotor neurons, monopolar cells), but most had more- or less-branched dendrites that were associated with one, or several, fields of terminal fibers in the octavolateral area. Unlike those of the ventral nucleus, labeled neurons of the medial nucleus were homogeneous in appearance (mostly pear-shaped). The dorsal nucleus was scarcely developed in larvae, as judged from the very simple and small labeled cells. The presence of terminal or "en-passant" boutons of secondary octavolateral fibers in the reticular area and the commissural nature of these fibers were also investigated by means of application of HRP or indocarbocyanine dye to the octavolateral nuclei. In addition, neurons of other alar plate nuclei that were labeled by the HRP application to the reticular nuclei (trigeminal descending root nucleus and solitary nucleus) were also characterized. The functional significance of these results is discussed.

The KlHEM1 gene from Kluyveromyces lactis encodes a functional 5-aminolevulinate synthase (deltaALA synthase), as confirmed by complementation of a hem1 mutant Saccharomyces cerevisiae strain, homology search, and detection of a 2.3 kb transcript. The gene is highly homologous to the ScHEM1 gene, and the sequence of the promoter region contains a complex combination of putative regulatory signals. Some of them are related to phospholipid biosynthesis, glycolytic metabolism, and regulation by carbon source. Transcription of KlHEM1 increased significantly in response to limited oxygen, and only slightly with the change from repressed (glucose) to derepressed conditions (glycerol). The deltaALA synthase from K. lactis contains, in the amino-terminal region, two heme-responsive elements that are not present in the protein from Saccharomyces cerevisiae.

2-Guanadino-3-cyanopyridines 8-33 and pyrido[2,3-d]-pyrimidines 35-52 were synthesized by nucleophilic displacement and cyclization of the chloroamidines 6a-d easily obtained by reaction of 2-aminocyanopyridines 5a-d with phosgene iminium chloride and their action on the release of histamine by mast cells examined under immunological and chemical stimulus, with and without pre-incubation. Several 2-guanadino-3-cyanopyridines and pyrido[2,3-d]-pyrimidines are shown to be inhibitors of the release of histamine when stimulated with ovoalbumin as antigen or with polymer 48/80 as chemical stimulus. Guanadino-3-cyanopyridine 30 and pyrido[2,3-d]-pyrimidine 49 are the more active of all, inhibiting the release of histamine in all the conditions tested (30-60% inhibition). Guanadinocyanopyridines 15, 17, and 19 are very potent stimulators of the release of histamine (150-300%) while pyrido[2,3-d]-pyrimidines are mostly inactive. Compounds 28 and 14 present moderate in vitro cytotoxic activity against P-388, A-549, HT-29, and MEL-28 cell lines.

The neuronal system of the saccus vasculosus of two species of trout was studied with immunocytochemical methods and carboindocyanine-dye (DiI) tract-tracing. The cerebrospinal-fluid-contacting neurons of the saccus were immunoreactive for gamma-aminobutyric acid (GABA), glutamic acid decarboxylase (GAD), and neuropeptide Y (NPY). Immunostaining of alternate sections of the saccus vasculosus of fry with anti-GAD and anti-NPY indicated that these substances were colocalized. The tractus sacci vasculosi and the neuropil of the nucleus sacci vasculosi were also immunoreactive to these substances. The GABA, GAD, and neuropeptide Y immunoreactivity of the saccus vasculosus system appeared early in trout ontogeny. After applying DiI to various levels of the tractus sacci vasculosi of adult trout, we observed massive bilateral saccular projections to the nucleus sacci vasculosi and could follow the course of the sacco-thalamic tract. This tract extended in the subependymal region of the thalamus rostral to the nucleus sacci vasculosi and split into two small tracts that reached the subhabenular-preoptic region. Sacco-thalamic fibers formed extensive periependymal plexuses along their trajectory. Interestingly, no clear evidence of the existence of a saccopetal system was obtained. On the basis of these results, we postulate that the saccus vasculosus system modulates the function of centers of the posterior tubercle and periventricular thalamus.

Experimental designs in clinical investigation are discussed in this article. Guideline examples have been used in the area of Cardiology using always the same one only one whenever possible. We have looked for a different perspective from what is generally used in the discussion of the general characteristics of experimental designs, and more specifically of clinical trials and we deal with the aspects of clinical trials which are usually ignored due to their marginal character. We also discuss those characteristics which differentiate clinical trials in respect to other designs and types of questions which are answered by clinical trials. And we finally discuss various aspects such as randomization and its various types (simple, block, stratified, pre-randomized) and variable types of evaluating the answers, masking and the problems in its maintenance, with certain kinds of designs, sample size, etc. There is a brief mention of two particular cases: factorial and cross over designs are both discussed, mentioning their strong and weak points. Likewise, we discuss community trials as another experimental design and examples are provided. Finally, we discuss aspects of criteria: such as, When to stop the trials? or Who are the results applicable to?, and we suggest points to take into consideration when these decisions are made.

Iontophoretic application of L-arginine (L-Arg) resulted in a profound decrease in visually elicited and spontaneous activity in 22 of 77 (29%) cells in area 17 of the anaesthetized/paralysed cat. Duration was long, and cells did not recover pre-application activity levels, indicating permanent decline. This effect was obtained without change in the extracellularly recorded wave-form, demonstrating that this did not result from depolarization block. In the remaining 55 cells, application of L-Arg alone, at levels capable of eliciting inhibition as described above, was without effect. In 29 cells, L-Arg application was able to reverse the effect of inhibition of nitric oxide (NO) production. Populations of cells showing the depressive effect described above and those affected by NO modulation levels were mutually exclusive.

A simple model is presented to illustrate how an underlying generalized polygenic basis of hybrid sterility is expected to lead to spurious localizations of factors with major effects, when a conventional experiment of recombination mapping is carried out. The model shows that a major gene will be detected at roughly the same distance from each of the chromosome markers used in the experiment. These expectations are contrasted with the results from several experiments on hybrid male sterility in Drosophila, which claimed to have mapped single sterility factors. It is concluded that, except for one report, all the evidence presented so far on the genetic basis of hybrid male sterility in Drosophila is in fact compatible with the generalized polygenic model.

The observation of segregation ratios of sterile and fertile males in offspring samples from backcrossed hybrid females is, in principle, a valid method to unveil the genetic basis of hybrid male sterility in Drosophila. When the female parent is heterozygous (hybrid) for a sterility factor with major effects, equal proportions of fertile and sterile sons are expected in her offspring. However, intact (not recombined) chromosome segments of considerable length are expected to give segregation ratios that can not be easily differentiated from the 1:1 ratio expected from a single factor. When the phenotypic character under analysis can be determined by combinations of minor factors from the donor species spanning a certain chromosome length, very large offspring samples may be needed to test this alternative hypothesis against the null hypothesis of a single major factor. This is particularly the case of hybrid male sterility determinants in Drosophila.

In hybrids between the sibling species D. buzzatii and D. koepferae, both sexes are more or less equally viable in the F1. However, backcross males to D. buzzatii are frequently inviable, apparently because of interspecific genetic incompatibilities that are cryptic in the F1. We have performed a genetic dissection of the effects of the X chromosome from D. koepferae. We found only two cytological regions, termed hmi-1 and hmi-2, altogether representing 9% of the whole chromosome, which when introgressed into D. buzzatii cause inviability of hybrid males. Observation of the pattern of asynapsis of polytene chromosomes (incomplete pairing, marking introgressed material) in females and segregation analyses were the technique used to infer the X chromosome regions responsible for this hybrid male inviability. The comparison of these results with those previously obtained with the same technique for hybrid male sterility in this same species pair indicate that in the X chromosome of D. koepferae there are at least seven times more regions that produce hybrid male sterility than hybrid male inviability. We have also found that the inviability brought about by the introgression of hmi-1 is suppressed by the cointrogression of two autosomal sections from D. koepferae. Apparently, these three regions conform to a system of species-specific complementary factors involved in an X-autosome interaction that, when disrupted in backcross hybrids by recombination with the genome of its sibling D. buzzatii, brings about hybrid male inviability.

The Simon effect was studied using ERPs and the lateralized readiness potential (LRP). In Experiment 1, visual stimuli requiring left or right hand responses were presented at either side of fixation. In Experiment 2, the empty side was occupied by a distractor. Spatial S-R incompatibility delayed P300 latency in both experiments, as predicted by perceptual interference accounts of the Simon effect. However, the LRP results were not conclusive. The lateralized presentation of visual stimuli (Experiment 1) or targets (Experiment 2) produced a posterior ERP hemispheric asymmetry that spread to central electrodes and obscured the early development of the LRP. Experiment 3 avoided the problem by presenting the stimuli in the vertical meridian, and the LRP showed signs of incorrect response preparation in incompatible trials, in agreement with response-interference models. Both perceptual and response interference may be present in the Simon effect.

Flow cytometry (FCM) was used to determine changes in cellular volume, transmembrane potential, mitochondrial membrane potential and intracellular pH in the marine microalga Phaeodactylum tricornutum immediately (5 to 10 seconds) after the addition of selected concentrations of copper. An acute increase in the forward scatter signal of this diatom was detected after the addition of 10 mg 1-1 of copper. Stress produced by the copper addition resulted in various physiological alterations that can be easily and quickly detected by FCM: (i) the hyperpolarization of the cell membrane, as a result of an immediate increase in the cytoplasmic membrane potential, (ii) the increase of the mitochondrial membrane potential, being maximum at the higher copper concentration assayed, and (iii) the increase in the intracellular pH with the highest copper concentration assayed (10 mg 1-1).

A bibliographic review on honey pollution with pesticides is presented. This paper reviews the methods set up for determining pesticide residues in honey samples as well as the pesticide residue levels found in European countries.

In the present work we obtained the experimental pKs of the amino acid alpha-alanine in NaCl at 25 degrees and different ionic strengths. The equilibrium constants have been potentiometrically determined with a commercial glass electrode and the results analysed through three models: two directly based on the Pitzer and Scatchard approaches to the ion specific interaction theory and the other based on a simpler modification of the Debye-Hückel equation. The three models fit the data reasonably well and the extrapolated pK values obtained show a good agreement. The goodness of the ridge regression method cannot be probed in this case.

A bibliographic review on the pollution of honey with acaricides is presented. This paper reviews methods for determining amitraz, bromopropylate, coumaphos, cymiazole, fluvalinate, malathion and phenothiazine residues in honey samples, as well as multiresidue methods. Acaricide residue levels found in European countries are also reviewed.

Transcriptional regulation of the KlCYC1 gene from the aerobic respiratory yeast Kluyveromyces lactis has been studied. The KlCYC1 gene produces two transcripts of different sizes, in contrast with the single transcripts found for CYC1 and CYC7 from Saccharomyces cerevisiae, and for the CYC gene from Schwanniomyces occidentalis. Both KlCYC1 transcripts respond in the same way to the regulatory signals studied here. The transcription of KlCYC1 is regulated by oxygen and this control is mediated by heme. The KlCYC1 gene is also subject to catabolite repression. Heterologous expression in S. cerevisiae mutants reveals that the factors HAP1 and HAP2 take part in the regulatory mechanism.

The availability of the KICYC1 sequence was used to establish homologies with other cytochrome c genes from yeasts and the fungus Neurospora crassa. In terms of nucleotide composition, the cytochrome c gene from Kluyveromyces lactis showed a higher homology with Schwanniomyces occidentalis than with Saccharomyces cerevisiae, and this point is discussed in regard to the differences found in the codon usage of these yeasts. The deduced amino acidic composition of the protein facilitated comparison of its sequence with other cytochrome c protein sequences and new assignments of phylogenetic relationships. In this context Kluyveromyces lactis was most closely related to Candida krusei.

We have shown that application of an inhibitor of the enzyme nitric oxide synthase (NOS) effectively suppresses the visual responses of relay cells in the dorsal lateral geniculate nucleus (dLGN) of the anaesthetized paralysed cat. Such suppression seems to result from a specific reduction in transmission via N-methyl-D-aspartic acid (NMDA) receptors, since iontophoretic application of the inhibitor of NOS selectively and in a dose-dependent manner decreased the responses to exogenously applied NMDA. Responses to other exogenously applied amino acid agonists, such as quisqualate (Quis), kainate (Kain) and alpha-amino-3-hydroxy-5-5-methyl-4-isoxazole-propionic acid (AMPA) were largely unaffected. Furthermore, the excitatory action of acetylcholine (ACh), normally co-localized with NOS in axonal terminals within the dLGN arising from the brainstem, was also unaffected. Unlike some other actions of nitric oxide (NO), this role seems not to involve an increase in production of cyclic guanosine-3',5'-mono-phosphate (cGMP), since application of the membrane permeable cGMP analogue 8-bromo-cGMP did not alter the suppressive effect of NOS inhibitors on either visual or NMDA evoked responses. We conclude that the normal function of NO at this level of the visual system is permissive, allowing full expression of NMDA mediated visually elicited information.

Different responses of the marine diatom Phaeodactylum tricornutum (Bohlin) to toxic copper concentrations were investigated. Besides the classical variables applied to toxicity studies in microalgae, such as growth or chlorophyll a content, other variables analyzed by flow cytometry were used. Toxic effects due to copper concentration were observed. Cell density reached in the stationary phase was reduced to 50% in cultures with 20 mg Cu/l, with respect to control cultures without copper. Cell light scatter properties (related to cell volume and intracellular granularity) and chlorophyll a fluorescence of microalgal cells were determined by flow cytometry analysis at the beginning of growth, 1 h after copper exposure, and when cultures reached the stationary phase (72 h). After 1 h of exposure to metal, no differences were observed, but when cultures reached the stationary phase, a gradual increase in the variables analyzed by flow cytometry was observed as the copper concentration increased. The increase in chlorophyll a fluorescence detected by flow cytometry was not correlated with an increase in the cell content of this photopigment, thus indicating an inhibitory effect of copper on photosystem II.

Codon usage (CU) in Kluyveromyces lactis has been studied. Comparison of CU in highly and lowly expressed genes reveals the existence of 21 optimal codons; 18 of them are also optimal in other yeasts like Saccharomyces cerevisiae or Candida albicans. Codon bias index (CBI) values have been recalculated with reference to the assignment of optimal codons in K. lactis and compared to those previously reported in the literature taking as reference the optimal codons from S. cerevisiae. A new index, the intrinsic codon deviation index (ICDI), is proposed to estimate codon bias of genes from species in which optimal codons are not known; its correlation with other index values, like CBI or effective number of codons (Nc), is high. A comparative analysis of CU in six cytochrome-c-encoding genes (CYC) from five yeasts is also presented and the differences found in the codon bias of these genes are discussed in relation to the metabolic type to which the corresponding yeasts belong. Codon bias in the CYC from K. lactis and S. cerevisiae is correlated to mRNA levels.

HRP tracing techniques and silver staining methods were used to investigate the central projections of the octaval nerve of the larval sea lamprey (Petromyzon marinus). We compare our findings with those reported for the adult and for other anamniotes. The general pattern of afferent projections (to the ventral octavolateral area of the medulla oblongata and to the cerebellar plate) is similar to that in the adult, although we did not observe projections to the trigeminal, reticular or contralateral medullary areas. We describe the pattern of connections of the octaval afferents with the different octaval nuclei (i.e. subdivisions of the ventral octavolateral area), and report the presence of a small number of efferent vestibular neurons.

The telencephalon of the small-spotted dogfish, Scyliorhinus canicula L., was examined by Nissl and Golgi-aldehyde techniques. On the basis of differences in perikaryal and dendritic morphology and size, several cell types were distinguished in pallial and subpallial regions, most of them reported here for the first time in elasmobranchs. In the pallium, the pallium dorsalis is the richest in cell types (eight types of neurons), whereas the neuron population of the pallium medialis is the most homogeneous. Dendrites of most neuron types in the pallium are smooth or sparsely thorny. Interestingly, the pallium dorsalis and pallium lateralis contain a type of primitive pyramidal cell characterized by the dense appearance of its thorny dendrites. In the subpallium, the area superficialis basalis contains a heterogeneous population (six types of neurons): large radial cells are the most characteristic cell type. Dendrites of these cell types are smooth or sparsely thorny. The cell populations of the nucleus N are roughly similar to those of the area superficialis basalis, but they lack the large radial cells characteristic of this area. The area centralis subpallialis and striatum consist of populations of small neurons. The regio septalis contains a rather homogeneous population of small cells. The populations in the nucleus entopeduncularis and the nucleus interstitialis of the basal forebrain bundle are the least varied and consist of large radial cells and bipolar cells similar to those of the area superficialis basalis. This investigation reveals important differences in cytoarchitecture that should be useful in the interpretation of immunocytochemical, tracing, and electrophysiological studies of the telencephalon of elasmobranchs.

The complete sequence of a cytochrome c gene from Kluyveromyces lactis including its upstream region is reported. Sequence of the translated open reading frame is discussed in terms of cytochrome c structural requirements. Putative regulatory signals in the upstream region are described and compared with reported sequences which modulate the expression of respiratory-related yeast genes.

The appearance of G- and C-banding patterns on metaphase chromosomes from the Don cell line (Cricetulus griseus) was accompanied by extraction of chromosomal DNA. A microdensitometric technique was employed to measure the loss of DNA produced by Hin dIII, Hae III and Eco RI endonuclease digestion on fixed chromosomes, from 0.5 to 24 h of treatment. Agarose gel electrophoresis was carried out to estimate the size of the DNA fragments extracted after digestion by Alu I and the restriction endonucleases already mentioned. Results obtained show that Alu I and Hae III, which possess 4 base pair recognition sequences, caused higher DNA extraction than enzymes with recognition targets of 6 base pairs (Hin dIII and Eco RI). Incubation buffers induced G-banding patterns which were accompanied by DNA extraction, although this loss was lower than that produced by endonuclease treatments.

The different responses of metaphase chromosomes to restriction endonuclease activity depend on the stage of chromosomal condensation. Results obtained established that this factor plays a remarkable role in obtaining high resolution banding patterns which facilitated visualization of a large number of sub-bands. It is evident that various restriction endonucleases, which are able to induce a coincident G-like banding pattern on Don cell line chromosomes, act in very different ways.