Biomedical subjects

M J Delgado

Publications and source records attributed to M J Delgado.

At least 19 recordsLinked to original sources

PubMed2006

Feeding entrainment of locomotor activity rhythms, digestive enzymes and neuroendocrine factors in goldfish.

The existence of food anticipatory activity (FAA) in animals subjected to daily feeding schedules seems to be mediated by a feeding-entrainable oscillator (FEO). Such an FEO may help in anticipating meal time and so optimizing food acquisition and nutrient utilization. In this study we investigated the existence of FAA and whether digestive enzymes, plasma cortisol, hypothalamic NPY and gastrointestinal tract (GIT) and plasma melatonin were entrained by periodic feeding in goldfish. We observed that periodically fed goldfish showed FAA in locomotor activity as well as in amylase and NPY. Alkaline protease and GIT melatonin were higher after feeding, whereas plasma cortisol levels were reduced. Plasma melatonin remained unmodified before and after meal time. These results suggested that scheduled feeding entrained both behavioral and certain physiological patterns in goldfish, FAA being of adaptive value to anticipate a meal and prepare the digestive physiology of fish.

Activity Cycles↗

PubMed2006

Effects of prenatal or postnatal ethanol consumption on zinc intestinal absorption and excretion in rats.

AIMS: The effect of ethanol consumption, either during the pregnancy or lactation period, on the altered metabolism of zinc is not well-defined; consequently, this study was performed to analyze the effect of chronic ethanol exposure on milk consumption, serum, milk, duodenal absorption, fecal and urinary excretion of zinc in dams and offspring during either gestation or lactation in the rat. A complementary study was performed regarding pregnancy outcome. We evaluated testosterone values, the offspring born/litter and several indices such as fertility, viable gestations and the survival index. METHODS: To study the effect of chronic alcoholism during gestation or lactation separately, at birth control newborns were cross-fostered to ethanol dams (ED), and the offspring issued from the ethanol treated mothers were cross-fostered to control dams (CD). Thus, three experimental groups of offspring were formed: (i) control offspring receiving no treatment (CO); (ii) offspring exposed to ethanol only during gestation (GO); and (iii) offspring exposed to ethanol only during lactation (LO). All the results were compared with offspring pair-fed groups (PFO) born of the pair-fed dams (PFD). RESULTS: Duodenal absorption of zinc increased significantly in LO offspring when the substrate concentrations in the perfusion medium were 25, 75, and 150 microM. A higher faecal excretion in GO pups compared with those with LO exposure and control groups (CO and PFO). The urine excretion of zinc was higher for LO offspring with respect to the other three experimental groups (CO, GO, and PFO). CONCLUSIONS: Maternal adaptation resulted in zinc retention, adequate to meet the demands of pup's growth in the face of a lower diet intake. The zinc status in pups is regulated by a higher absorption of zinc and intestinal conservation of endogenous fecal zinc after postnatal ethanol consumption. The increase in urinary zinc excretion could be responsible for decreased serum zinc. However, we found an increase in serum zinc probably due to an increase in the zinc absorption values.

Animals↗

PubMed2006

The Bradyrhizobium japonicum napEDABC genes are controlled by the FixLJ-FixK(2)-NnrR regulatory cascade.

Nitrate respiration by the N(2)-fixing symbiotic bacteria Bradyrhizobium japonicum USDA110 is mediated by a Nap (periplasmic nitrate reductase) encoded by the napEDABC genes. Expression of a transcriptional fusion of the nap promoter region to the reporter gene lacZ, P(napE)-lacZ, was very low in aerobically grown cells of USDA110, but expression was induced approx. 3-fold when the cells were cultured under microaerobic conditions, and 12-fold when nitrate was added to the microaerobic incubation medium. The P(napE)-lacZ fusion was not expressed in the fixL 7403, fixJ 7360 and fixK(2) 9043 mutant strains. Microaerobic induction of the P(napE)-lacZ fusion was retained in the nnrR 8678 mutant, but no increase in beta-galactosidase activity was observed upon nitrate addition. Western-blot and Methyl Viologen-dependent nitrate reductase activity assays showed that synthesis and activity of the catalytic NapA subunit in USDA110 was similar to that in the napC 0906 and nirK GRK308 mutant strains incubated microaerobically with nitrate. These results suggest that nitrate and nitrite, which are not reduced by the napC 0906 and nirK GRK308 mutant cells respectively, induced the synthesis and activity of NapA; conversely, formation of endogenous NO was not required for induction of Nap expression.

Bacterial Proteins↗

PubMed2006

The role of Bradyrhizobium japonicum nitric oxide reductase in nitric oxide detoxification in soya bean root nodules.

The identification of nitric oxide-bound leghaemoglobin within soya bean nodules has led to the question of how Bradyrhizobium japonicum bacteroids overcome the toxicity of this nitric oxide. It has previously been shown that one candidate for nitric oxide detoxification, the respiratory nitric oxide reductase, is expressed in soya bean nodules from plants supplied with nitrate. In this paper, the role of this enzyme in nitric oxide detoxification is assessed and discussion is provided on other possible B. japonicum nitric oxide detoxification systems.

Bradyrhizobium↗

PubMed2006

Acute and chronic leptin reduces food intake and body weight in goldfish (Carassius auratus).

The purpose of the present study was to elucidate the possible role of leptin in food intake and body weight regulation in goldfish. We examined the effects of i.c.v. or i.p. acute leptin administration on food intake in food-deprived goldfish at different time intervals post-injection (0-2, 2-8 and 0-8 h). Food intake was reduced by i.p. administered leptin (1 microg) at 8 h post-injection, without statistically significant differences after i.c.v. treatment. The present study shows for the first time in a teleost that chronic (10 days) leptin treatment (i.p.) reduces food intake, body weight gain, specific growth rate and food efficiency ratio. Moreover, lipid and carbohydrate metabolism seems to be regulated by leptin in fish. Chronic leptin treatment increased lipid mobilization and carbohydrate storage as hepatic and muscle glycogen. Finally, leptin could mediate its actions on energy homeostasis in fish, at least in part, through interactions with hypothalamic catecholamines, since chronic leptin treatment reduced both hypothalamic noradrenergic and dopaminergic turnover without significant modifications in hypothalamic serotoninergic and neuropeptide Y (NPY) systems. In summary, our results suggest that leptin can regulate feeding behaviour and body weight homeostasis in fish.

Animals↗

PubMed2005

The endocannabinoid system in the brain of Carassius auratus and its possible role in the control of food intake.

Cannabinoid receptors and the endocannabinoids anandamide and 2-arachidonoylglycerol have been suggested to regulate food intake in several animal phyla. Orthologs of the mammalian cannabinoid CB(1) and CB(2) receptors have been identified in fish. We investigated the presence of this endocannabinoid system in the brain of the goldfish Carassius auratus and its role in food consumption. CB(1)-like immunoreactivity was distributed throughout the goldfish brain. The prosencephalon showed strong CB(1)-like immunoreactivity in the telencephalon and the inferior lobes of the posterior hypothalamus. Endocannabinoids were detected in all brain regions of C. auratus and an anandamide-hydrolysing enzymatic activity with features similar to those of mammalian fatty acid amide hydrolase was found. Food deprivation for 24 h was accompanied by a significant increase of anandamide, but not 2-arachidonoylglycerol, levels only in the telencephalon. Anandamide caused a dose-dependent effect on food intake within 2 h of intraperitoneal administration to satiated fish and significantly enhanced or reduced food intake at low (1 pg/g body weight) or intermediate (10 pg/g) doses, respectively, the highest dose tested (100 pg/g) being inactive. We suggest that endocannabinoids might variously contribute to adaptive responses to food shortage in fish.

Amidohydrolases↗

PubMed2005

Molybdate-dependent expression of the periplasmic nitrate reductase in Bradyrhizobium japonicum.

The napEDABC genes of Bradyrhizobium japonicum encode the periplasmic nitrate reductase, an Mo-containing enzyme which catalyses the reduction of nitrate to nitrite when oxygen concentrations are limiting. In this bacterium, another set of genes, modABC, code for a high affinity ABC-type Mo transport system. A B. japonicum modA mutant has been obtained that is not capable of growing anaerobically with nitrate and lacks nitrate reductase activity. Under nitrate respiring conditions, when Mo concentrations are limiting, the B. japonicum modA mutant lacked both the 90 kDa protein corresponding to the NapA component of the periplasmic nitrate reductase, and the membrane-bound 25 kDa c-type cytochrome NapC. Regulatory studies using a napE-lacZ fusion indicated that napE expression was highly reduced in the modA mutant background when the cells were incubated anaerobically with nitrate under Mo-deficient conditions.

Blotting, Western↗

PubMed2005

The complete denitrification pathway of the symbiotic, nitrogen-fixing bacterium Bradyrhizobium japonicum.

Denitrification is an alternative form of respiration in which bacteria sequentially reduce nitrate or nitrite to nitrogen gas by the intermediates nitric oxide and nitrous oxide when oxygen concentrations are limiting. In Bradyrhizobium japonicum, the N(2)-fixing microsymbiont of soya beans, denitrification depends on the napEDABC, nirK, norCBQD, and nosRZDFYLX gene clusters encoding nitrate-, nitrite-, nitric oxide- and nitrous oxide-reductase respectively. Mutational analysis of the B. japonicum nap genes has demonstrated that the periplasmic nitrate reductase is the only enzyme responsible for nitrate respiration in this bacterium. Regulatory studies using transcriptional lacZ fusions to the nirK, norCBQD and nosRZDFYLX promoter region indicated that microaerobic induction of these promoters is dependent on the fixLJ and fixK(2) genes whose products form the FixLJ-FixK(2) regulatory cascade. Besides FixK(2), another protein, nitrite and nitric oxide respiratory regulator, has been shown to be required for N-oxide regulation of the B. japonicum nirK and norCBQD genes. Thus nitrite and nitric oxide respiratory regulator adds to the FixLJ-FixK(2) cascade an additional control level which integrates the N-oxide signal that is critical for maximal induction of the B. japonicum denitrification genes. However, the identity of the signalling molecule and the sensing mechanism remains unknown.

Bradyrhizobium↗

PubMed2005

Function of the Rhizobium etli CFN42 nirK gene in nitrite metabolism.

Rhizobium etli CFN42 is not capable of growing anaerobically with nitrate but it grows with nitrite as a terminal electron acceptor. This bacterium contains the nirK gene encoding the copper-containing Nir (nitrite reductase), which is located on the cryptic plasmid pCFN42f. Mutational analysis has demonstrated that a nirK deficient mutant was not capable of growing under nitrite-respiring conditions. Moreover, microaerobic growth of this mutant was inhibited by the presence of nitrite. Nir activity and nitrite uptake were highly diminished in a nirK mutant, compared with the wild-type levels after incubation under anaerobic conditions. Our results suggest that the copper-containing Nir may have both a respiratory and a nitrite-detoxifying role in R. etli.

Genes, Bacterial↗

PubMed2004

Binding characteristics and daily rhythms of melatonin receptors are distinct in the retina and the brain areas of the European sea bass retina (Dicentrarchus labrax).

Melatonin is synthesized, with a circadian rhythm, in the pineal organ of vertebrates, high levels being produced during the scotophase and low levels during the photophase. The retina also produces melatonin, although in the case of the European sea bass, its secretion pattern appears to be inverted. In the study described here, radioreceptor assay techniques were used to characterize the melatonin binding sites, their regional distribution and their daily variations. Brain and retina membrane preparations were used in all the binding assays and 2-[125I]iodomelatonin ([125I]Mel) as radioligand at 25 degrees C. The specific binding of [125I]Mel was seen to be saturable, reversible, specific and of high affinity. In all the tissues assayed, the power of the ligands to inhibit [125I]Mel binding decreased in the following order: melatonin>>4-P-PDOT>luzindole> or =N-acetylserotonin, which points to the presence of Mel1-like receptors. The inhibition curves of 4-P-PDOT suggested the presence of two different binding sites in the brain areas, but only one type of site of low affinity in the neural retina. No daily variations in [125I]Mel binding capacity (Bmax) or affinity (Kd) were detected in the brain areas, while a clear rhythm in Kd melatonin receptor affinity and Bmax binding capacity was observed in the retina. Kd and Bmax retinal rhythms were out of phase with the lowest Kd and the highest Bmax occurring at scotophase. This result suggests that retinal melatonin is a paracrine factor able to control receptor desensitization during photophase when ocular melatonin is higher in this species.

Animals↗

PubMed2003

Changes in glucose, glycogen, thyroid activity and hypothalamic catecholamines in tench by starvation and refeeding.

The effects of short-term food deprivation (7 days) and refeeding (2 days) on different biochemical and neuroendocrine parameters were studied in tench. A 7-days fast resulted in a significant reduction of plasma glucose and glycogen hepatic content, supporting the key role of liver glycogen as energy depot for being consumed during fasting. The rapid recovery of normal values of blood glucose and glycogen stores by refeeding indicates a rapid replenishment of liver glycogen stores. The short-term starvation decreased circulating thyroid hormones (both T3 and T4) and T4 release from thyroid, supporting an interaction between nutritional state and thyroid function in tench. All these metabolic and hormonal changes were partial or totally reversed under refeeding conditions. An increase in hypothalamic content of norepinephrine and dopamine was found in fasted fish. This result might be a consequence of stress induced by starvation.

Analysis of Variance↗

PubMed2001

Characterization of the nirK gene encoding the respiratory, Cu-containing nitrite reductase of Bradyrhizobium japonicum.

The structural gene, nirK, for the respiratory Cu-containing nitrite reductase from Bradyrhizobium japonicum USDA110 has been isolated and sequenced. The deduced amino acid sequence exhibited a high degree of similarity to other Cu-containing nitrite reductases from various sources. The full-length protein included a signal peptide for protein export. Analysis of the sequence upstream from the structural nirK gene revealed the presence of an anaerobox located 83 base pairs from the putative translational start codon. Cells of strain GRK308, a nitrite reductase-deficient derivative of strain USDA110, were unable to grow when cultured under microaerobic conditions (1% O(2)) in the presence of either nitrate or nitrite. Maximal expression of a nirK-lacZ fusion in strain USDA110 required simultaneously both low level oxygen conditions and the presence of nitrate. Expression of beta-galactosidase activity was not detected in the B. japonicum fixL 7403, fixJ 7360 and fixK(2) 9043 mutants transformed with the nirK-lacZ fusion after incubation of the cells under oxygen-limiting conditions either with or without nitrate. Complementation of B. japonicum 9043 with the fixK(2) gene restored beta-galactosidase activity to levels similar to those found in the parental strain. These results suggest that nirK expression depends on the low-oxygen-responsive two-component regulatory system FixLJ and on the Fnr/FixK-like DNA binding protein FixK(2).

Amino Acid Sequence↗

PubMed2001

Day/night variations of dopamine ocular content during Xenopus laevis ontogeny.

Concentration of dopamine (DA) and its metabolite, 3,4-dihydroxyphenylacetic acid is quantified by high-pressure liquid chromatography with a coulometric detection system in the eye of Xenopus laevis through ontogeny and in adults at two times during photocycle (midday and midnight). Ocular dopaminergic activity remains low during pre- and prometamorphosis and significantly rises in postmetamorphic froglets. This increase is more pronounced at midnight than at midday. The dualism of DA content versus DA release in Xenopus ocular tissue is studied in an eyecup culture system. On a 24-h cycle of DA release from adult Xenopus eyecups the highest DA release by eyecups is produced during daytime, and significantly decreases in darkness. From these results it can be concluded that in spite of the early development of the retinal dopaminergic system in the ontogeny of Xenopus, the final maturation must occur during the metamorphic climax. Endogenous DA release is significantly inhibited by light offset, which explains the higher ocular DA content found at midnight as compared to midday in postmetamorphic froglets and adults.

3,4-Dihydroxyphenylacetic Acid↗

PubMed2001

Food intake inhibition by melatonin in goldfish (Carassius auratus).

Feeding regulation by monoamines, neuropeptides and certain hormones has been studied in fish, but a possible role of melatonin is unknown. The purpose of the present study was to investigate the effects of melatonin on food intake in goldfish. Fishes were housed in 12L:12D and injected with different doses of either melatonin or 2-iodomelatonin. Two routes of administration, intracerebroventricular and intraperitoneal injections, and two times of the daily photocycle, midday and midnight, were tested. Food intake was measured at 2, 5 and 8 h postinjection. Melatonin and its analog, 2-iodomelatonin intracerebroventricularly injected had no effect on food intake at any time. However, intraperitoneal injections of both indoleamines significantly reduced food intake at different postinjection times. The inhibitory effect of melatonin was blocked by intraperitoneal administration of its antagonist, luzindole. These results demonstrate the in vivo efficiency of luzindole as melatonin antagonist, and thus provide a useful experimental tool to investigate melatonin functions. In conclusion, both melatonin and its agonist 2-iodomelatonin administered peripherally, inhibit food intake in goldfish, and this inhibitory effect appears to be mediated via luzindole-sensitive melatonin receptors. Our results strongly suggest that melatonin is involved in the peripheral satiety mechanisms in goldfish.

Animals↗

PubMed2001

One of two hemN genes in Bradyrhizobium japonicum is functional during anaerobic growth and in symbiosis.

Previously, we screened the symbiotic gene region of the Bradyrhizobium japonicum chromosome for new NifA-dependent genes by competitive DNA-RNA hybridization (A. Nienaber, A. Huber, M. Göttfert, H. Hennecke, and H. M. Fischer, J. Bacteriol. 182:1472-1480, 2000). Here we report more details on one of the genes identified, a hemN-like gene (now called hemN(1)) whose product exhibits significant similarity to oxygen-independent coproporphyrinogen III dehydrogenases involved in heme biosynthesis in facultatively anaerobic bacteria. In the course of these studies, we discovered that B. japonicum possesses a second hemN-like gene (hemN(2)), which was then cloned by using hemN(1) as a probe. The hemN(2) gene maps outside of the symbiotic gene region; it is located 1.5 kb upstream of nirK, the gene for a Cu-containing nitrite reductase. The two deduced HemN proteins are similar in size (445 and 450 amino acids for HemN(1) and HemN(2), respectively) and share 53% identical (68% similar) amino acids. Expression of both hemN genes was monitored with the help of chromosomally integrated translational lacZ fusions. No significant expression of either gene was detected in aerobically grown cells, whereas both genes were strongly induced (> or = 20-fold) under microaerobic or anaerobic conditions. Induction was in both cases dependent on the transcriptional activator protein FixK(2). In addition, maximal anaerobic hemN(1) expression was partially dependent on NifA, which explains why this gene had been identified by the competitive DNA-RNA hybridization approach. Strains were constructed carrying null mutations either in individual hemN genes or simultaneously in both genes. All mutants showed normal growth in rich medium under aerobic conditions. Unlike the hemN(1) mutant, strains lacking a functional hemN(2) gene were unable to grow anaerobically under nitrate-respiring conditions and largely failed to fix nitrogen in symbiosis with the soybean host plant. Moreover, these mutants lacked several c-type cytochromes which are normally detectable by heme staining of proteins from anaerobically grown wild-type cells. Taken together, our results revealed that B. japonicum hemN(2), but not hemN(1), encodes a protein that is functional under the conditions tested, and this conclusion was further corroborated by the successful complementation of a Salmonella enterica serovar Typhimurium hemF hemN mutant with hemN(2) only.

Amino Acid Sequence↗

PubMed2000

Melatonin synthesis in the greenfrog retina in culture: I. Modulation by the light/dark cycle, forskolin and inhibitors of protein synthesis.

Melatonin is synthesized in the pineal gland and the retina of vertebrates. Retinal serotonin N-acetyltransferase (NAT) activity and melatonin show a daily rhythm with high levels during the dark phase of the photocycle. In some vertebrates, these retinal NAT and melatonin rhythms are maintained in vitro. The aim of present work is to develop an eyecup culture system for the greenfrog (Rana perezi), suitable to analyze the mechanisms of regulation of melatonin synthesis by simultaneous determination of NAT activity and melatonin release. The R. perezi eyecups released melatonin to the culture medium in a rhythmic manner at least over a 27-h period under photocycle conditions. NAT activity and melatonin rhythms were similar to that observed in vivo under natural environmental conditions. Rana perezi retina exhibits a pronounced photosensitivity in vitro. Forskolin increased up to 2-fold the NAT activity and 4-fold the melatonin production at any lighting conditions. The addition of the translation inhibitor, cycloheximide, to the medium reduced significantly both nocturnal NAT activity and melatonin release, suggesting that de novo protein synthesis is produced daily during darkness. Actinomycin D, a transcription inhibitor, needs a longer time of action, because pre-existing mRNA must be depleted before the inhibition of melatonin release can be observed. The eyecup culture system is highly sensitive to light and chemical factors, which makes it particularly suitable as a model for the neurochemical analysis of melatonin biosynthesis in the retina of Rana perezi.

Animals↗

PubMed2000

Melatonin synthesis in the greenfrog retina in culture: II. Dopaminergic and adrenergic control.

Serotonin N-acetyltransferase (NAT) activity and melatonin show a daily rhythm with high levels at night. Although the rhythmic properties of NAT and melatonin are similar in pineal gland and retina, great differences in the light perception and transmission mechanisms exist. We have analyzed the effects of adrenergic and dopaminergic agents on greenfrog (Rana perezi) eyecup culture, in order to identify the receptors involved in the regulation of retinal melatonin synthesis. A D2-like receptor is directly involved in the regulation of NAT activity and melatonin release in R. perezi retina. Quinpirole mimics the effect of light, reducing the darkness-stimulated NAT activity and melatonin release, while sulpiride antagonized these actions. Neither D1-agonist (SKF 38393) nor D1-antagonist (SCH 23390) had effect on NAT activity. However, a significant inhibition of darkness-evoked melatonin release was produced by SKF 38393 after 6 hours of culture. The beta- and antagonist1-agonists showed a clear inhibition. However, a direct effect of beta, alpha1 and D1-agonists on photoreceptors is unproven, being more probable that the adrenergic actions imply a non-photoreceptor retinal cell. In conclusion, eyecup culture of Rana perezi revealed a dopaminergic control of melatonin synthesis and a possible modulation of dopaminergic tone by adrenergic receptors. Melatonin release is a more sensitive parameter than NAT activity to the action of neuroactive agents, suggesting that melatonin synthesis can be regulated by more than one enzymatic step in Rana perezi.

Adrenergic Agents↗

PubMed2000

NPY receptors and opioidergic system are involved in NPY-induced feeding in goldfish.

The present study evaluated the effects of both intraperitoneal (i.p. ) and intracerebroventricular administration of selective Y(1) [(Leu(31), Pro(34))-NPY] and Y(2) [(Pro(13), Tyr(36))-NPY (13-36)] receptor agonists on food intake in satiated goldfish. Food intake (FI) was significantly increased by central administration of the Y(1) agonist (1 microg), but not by the Y(2) agonist, at 2 h postinjection. The feeding increase induced by (Leu(31), Pro(34))-NPY was in a similar magnitude to that obtained after ICV injection of the neuropeptide Y, and both feeding stimulations were reversed by the NPY (27-36), a general NPY antagonist. The i.p. administration of the agonists either did not significantly modify (Y(2) agonist) or decreased (Y(1) agonist) food intake in goldfish. These data indicate that it is the Y(1)-like (similar to Y(1) and/or Y(5)) receptor, and not Y(2), that is involved in the central modulation of the feeding behavior in goldfish. We also investigated the possible involvement of opioid peptides as mediators of the NPY stimulatory action on food intake in goldfish. The ICV administration of naloxone (10 microg), a general opioid antagonist, blocked the NPY-induced feeding in goldfish, suggesting that the opioidergic system is involved in feeding regulation by NPY.

Animals↗