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S Villanueva

Publications and source records attributed to S Villanueva.

At least 19 recordsLinked to original sources
PubMed2001

Protein synthesis at the synapse: developmental changes, subcellular localization and regional distribution of polypeptides synthesized in isolated dendritic fragments.

This study evaluated local protein synthesis in subcellular fractions of pinched-off dendrites (synaptodendrosomes) from different brain regions and at different developmental ages. Synaptodendrosomes were labeled with [35S]methionine and newly synthesized proteins were characterized by SDS-PAGE and phosphorimaging. The same set of approximately 30 cycloheximide-sensitive labeled bands was observed in synaptodendrosomes isolated from different brain regions, although the relative enrichment of some individual bands varied. Labeling of several major proteins was developmentally regulated, revealing three different patterns of variation. Subcellular fraction studies revealed that at least 10 labeled bands were enriched in synaptic junctions.

Animals↗
PubMed2001

Glycoprotein synthesis at the synapse: fractionation of polypeptides synthesized within isolated dendritic fragments by concanavalin A affinity chromatography.

The synthesis of glycosylated proteins at postsynaptic sites was evaluated by combining metabolic labeling of isolated pinched-off dendritic fragments (synaptodendrosomes) with glycoprotein isolation by Con A affinity chromatography. Three major labeled proteins were detected (apparent molecular weights of 128, 42 and 19 kDa) along with seven minor polypeptides. Treatment of the glycoprotein fraction with N-glycosidase F led to shift in the apparent molecular weight of the bands. Also, label incorporation into glycoprotein species was blocked by tunicamycin. Thus, the three prominent polypeptides and most of the minor components of this fraction corresponded to bona fide N-glycoproteins. Incubation of synaptodendrosomes with cycloheximide also inhibited label incorporation into the isolated glycoproteins, indicating that the labeling resulted from local de novo synthesis. Subcellular fractionation revealed that the labeled glycoproteins were present in soluble and particulate fractions, mainly microsomes and synaptic membranes, and one of the species (42 kDa) appeared in the incubation medium, indicating secretion. In addition, these glycoproteins were dissimilarly distributed in several brain regions, and were expressed differentially during development, reaching their highest level of synthesis during the period of synaptogenesis. These results provide evidence for local dendritic synthesis of particular glycoprotein components of the synapse.

Amidohydrolases↗
PubMed2001

[Blood concentrations of calcium and magnesium in women with severe pre-eclampsia].

Abnormalities in the calcium and magnesium metabolism may participate in the pathogenesis of preeclampsia. Serum levels of magnesium, calcium, sodium, potassium and chlorine were measured in 15 pregnant women with severe preeclampsia and 20 uncomplicated pregnant women. Serum sodium, potassium and chlorine concentrations were similar in both groups. Nevertheless, serum magnesium levels were significantly elevated in severe preeclamptic women relative to women with uncomplicated pregnancy. The serum calcium concentration and calcium/magnesium ratio were not differences in the preeclamptic and the healthy pregnant woman.

Adolescent↗
PubMed2000

Effects of nitric oxide gas on cat carotid body chemosensory response to hypoxia.

It has been proposed that nitric oxide (NO) is an inhibitory modulator of carotid body (CB) chemoreception to hypoxia. However, the effects of NO gas on carotid chemoreception have not been tested yet. The role played by NO has been revealed by the use of pharmacological tools (i.e., NO donors and NO synthase inhibitors). Here, we studied the effects of NO gas (25 ppm in N(2)) on the chemosensory response to hypoxia (PO(2) approximately 30 Torr) in the cat CB perfused in vitro. During steady hypoxic chemoreceptor excitation, bolus injections or perfusion of Tyrode equilibrated with NO reduced the increased frequency of carotid chemosensory discharges (f(x)). Perfusion for 2 min of Tyrode equilibrated with NO also reduced the rate of the rise of the chemosensory response, as well as the maximal amplitude, as compared with the normal chemosensory response to hypoxia. Present results provide direct evidence that NO gas is an inhibitory modulator of CB hypoxic chemoreception.

Animals↗
PubMed2000

Dual effects of nitric oxide on cat carotid body chemoreception.

We studied the effects of nitric oxide (NO) released by NO donors on cat carotid body (CB) chemosensory activity during normoxia and hypoxia. CBs excised from pentobarbital sodium-anaesthetized cats were perfused with Tyrode at 38 degrees C and pH 7.40. The frequency of chemosensory discharges (f(x)) was recorded from the carotid sinus nerve, and changes of NO concentration were measured by a chronoamperometric technique, with NO-selective carbon-fiber microelectrodes inserted in the CB. During steady chemosensory excitation induced by hypoxia, bolus injections of NO (DeltaNO = 0. 5-12 microM), released by S-nitroso-N-acetylpenicillamine (SNAP) and 6-(2-hydroxy-1-methyl-nitrosohydrazino)-N-methyl-1-hexanamine++ + (NOC-9), transiently reduced f(x) in a dose-dependent manner. However, during normoxia, the same concentration of NO (DeltaNO = 0. 5-13 microM) released by the NO donors increased f(x) in a dose-dependent manner. The present results show a dual effect of NO on CB chemoreception that is dependent on the PO(2) levels. During hypoxia, NO is predominantly an inhibitor of chemoreception, whereas, in normoxia, NO increased f(x). The mechanisms by which NO produces chemosensory excitation during normoxia remain to be determined.

Animals↗
PubMed1999

[Effects of tibolone on lipid and glucose metabolism as well as insulin secretion in postmenopausal women].

Tibolone is a synthetic steroid with progestogenic, androgenic and estrogenic properties. In postmenopausal women it has been shown that improves climateric complaints and prevents osteoporosis, without impact on endometrial thickness and uterine fibroids volume. Tibolone administration decreases cholesterol and triglycerides. Although, the effects of hormonal replacement on insulin sensitivity and glucose metabolism has not been well established. The effect of tibolone 2.5 mg/day for 3 months was investigated in 10 healthy postmenopausal women. The results show that tibolone decreases tryglicerides and cholesterol levels along with an increase in fasting insulin levels. The oral glucose tolerance test was unaffected. This study suggests that tibolone reduces markers of cardiovascular disease.

Anabolic Agents↗
PubMed1997

Effect of glutamate receptor phosphorylation by endogenous protein kinases on electrical activity of isolated postsynaptic densities of rat cortex and hippocampus.

Postsynaptic densities (PSDs) were isolated from rat brain cortex and hippocampus, purified and incorporated into giant (5-80 microns in diameter) liposomes. Gigaohm seals were obtained with a patch-clamp pipette, and a giant liposome PSD-containing membrane patch, was excised and recorded. The PSD was always oriented in an inside-out configuration. This allowed receptor agonists or antagonists to be added from the interior of the recording pipette, and also the addition of different substances, such as ATP, calcium, calmodulin and others to the 'intracellular' side of the PSD, i.e. to the bath. alpha-Amino-3-hydroxy-5-methylisoxazole propionic acid (AMPA) receptor agonists such as quisqualate or AMPA induced in the PSD a complex pattern of electrical activity, that was blocked by 10 microM 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), but not by 2-aminophosphonovalerate (APV). The currents generated by 0.5-1 microM quisqualate were increased by about 100% when the PSDs were phosphorylated. Similar findings were obtained when the agonist was 0.2-2 microM kainate. These currents were also blocked by a non-N-methyl-D-aspartate (NMDA) receptor antagonist but not by APV, and were increased by about 70% by phosphorylation of the PSDs. Addition of 5-10 microM NMDA plus 1 microM glycine to the 'extracellular' side of the PSD, led to a characteristic pattern of activity, with the opening of multiple receptor ion channels. This was entirely blocked by 10 microM APV. Addition of extracellular Mg2+ (1-2 mM) induced a voltage-dependent block of the currents. Phosphorylation of the PSD led to an increase of Mg(2+)-blocked current of about 80%. The effect of phosphorylation on ion channel activity showed a markedly different requirement for calcium and for calmodulin among the AMPA, kainate and NMDA types of glutamate receptors, thus suggesting that each receptor type is coupled at the synapse with a unique complement of protein phosphokinases.

2-Amino-5-phosphonovalerate↗
PubMed1997

Petroleum pollution in the Gulf of Mexico and Caribbean Sea.

In 1976, IOC-UNESCO and UNEP convened a meeting in Port of Spain to analyze the marine pollution problems in the region, noting that petroleum pollution was of regionwide concern and recommended initiating a research and monitoring program to determine the severity of the problem and monitor its effects. The Wider Caribbean is potentially one of the largest oil-producing areas in the world. Major production sites include Louisiana and Texas in the U.S.; the Bay of Campeche, Mexico; Lake Maracaibo, Venezuela; and the Gulf of Paria, Trinidad. All these are classified as high-risk production accident zones. Main sources of petroleum pollution in the Wider Caribbean are production, exploitation, transportation, urban and municipal discharges, refining and chemical wastes, normal loading and unloading operations, and accidental spills. About 5 million barrels of crude oil are transported daily in the Caribbean, thus generating an intense tanker traffic. It has been estimated that oil discharges from tank washings within the Wider Caribbean could be as high as 7 million barrels/yr. The results of the Caribbean Pollution Regional Program (CARIPOL) conducted between 1980 and 1987 pointed out that significant levels of petroleum pollution exist throughout the Wider Caribbean, including serious tar contamination of windward exposed beaches, high levels of floating tar within the major current systems, and very high levels of dissolved and dispersed hydrocarbons in surface waters. Major adverse effects of this type of pollution include: high tar levels on many beaches that either prevent their recreational use or require very expensive cleanup operations, distress and death for marine life, and responses in the enzyme systems of marine organisms that have been correlated with declines in reproductive success. Finally, the presence of polycyclic aromatic hydrocarbons (PAHs) in tissues of important economic species has been reported, creating a risk for public health because of the potential carcinogenic effects.

Accidents, Occupational↗
PubMed1993

Recordings of glutamate receptor channels in isolated postsynaptic densities.

We have isolated highly purified rat brain postsynaptic densities (PSDs), that are known to contain glutamate receptors of the AMPA and NMDA types. These PSDs were incorporated into liposomes, and grown, by a cycle of partial de- and rehydration in 5% ethylene glycol, into giant (5-100 microns in diameter) liposomes. These giant liposomes were then made to form Gigaohm (10-20 G omega) seals with conventional patch-clamp electrodes, which, when withdrawn, retain an excised patch in an inside-out configuration. When 5-10 microM L-glutamate (or 10 microM NMDA) plus 1 microM glycine were present inside the patch pipette, but not in the external fluid, a highly complex pattern of currents was seen in about 55% of the cases. This was characterized by very fast kinetics, conductances as high as 460 pS and multiple lower levels of 45, 80, 120, 230 and 340 pS. These currents, when evoked by NMDA plus glycine, were entirely suppressed by the NMDA antagonist 2-amino-5-phosphonovalerate, APV. However, those activated by L-glutamate plus glycine still appeared in the presence of APV in about 18% of the cases, but with lower conductance levels. Current kinetics similar to the latter ones were also induced by the AMPA receptor agonist quisqualate (10 microM) in 16% of the cases. This indicated that both NMDA and AMPA receptors were present, in a functionally well preserved state, in isolated postsynaptic densities. Indirect evidence also suggested that in our experiments, in which 212 seals were studied, only a single postsynaptic density was present in the patches in which channel activity was found.(ABSTRACT TRUNCATED AT 250 WORDS)

2-Amino-5-phosphonovalerate↗
PubMed1993

The chemical nature of the main central excitatory transmitter: a critical appraisal based upon release studies and synaptic vesicle localization.

The chemical nature of the central transmitter responsible for fast excitatory events and other related phenomena is analysed against the historical background that has progressively clarified the structure and function of central synapses. One of the problems posed by research in this field has been whether one or more of the numerous excitatory substances endogenous to the brain is responsible for fast excitatory synaptic transmission, or if such a substance is, or was, a previously unknown one. The second question is related to the presence in the CNS of three main receptor types related to fast excitatory transmission, the so-called alpha-amino-3-hydroxy-5-methylisoxazole propionic acid, kainate and N-methyl-D-aspartate receptors. This implies the possibility that each receptor type might have its own endogenous agonist, as has sometimes been suggested. To answer such questions, an analysis was done of how different endogenous substances, including L-glutamate, L-aspartate, L-cysteate, L-homocysteate, L-cysteine sulfinate, L-homocysteine sulfinate, N-acetyl-L-aspartyl glutamate, quinolinate, L-sulfoserine, S-sulfo-L-cysteine, as well as possible unknown compounds, were able to fulfil the more important criteria for transmitter identification, namely identity of action, induced release, and presence in synaptic vesicles. The conclusion of this analysis is that glutamate is clearly the main central excitatory transmitter, because it acts on all three of the excitatory receptors, it is released by exocytosis and, above all, it is present in synaptic vesicles in a very high concentration, comparable to the estimated number of acetylcholine molecules in a quantum, i.e. 6000 molecules. Regarding a possible transmitter role for aspartate, for which a large body of evidence has been presented, it seems, when this evidence is carefully scrutinized, that it is either inconclusive, or else negative. This suggests that aspartate is not a classical central excitatory transmitter. From this analysis, it is suggested that the terms alpha-amino-3-hydroxy-5-methylisoxazole propionic acid, kainate and N-methyl-D-aspartate receptors, should be changed to that of glutamate receptors, and, more specifically, to GLUA, GLUK and GLUN receptors, respectively. When subtypes are described, a Roman numeral may be added, as in GLUNI, GLUNII, and so on.

Animals↗
PubMed1993

Use of play with clay to treat children with intractable encopresis.

We used play with modeling clay to treat six children, aged 4 to 12 years, with a history of intractable constipation with encopresis for a mean of 5.4 (2 to 8) years, refractory to treatment; biofeedback therapy had not been tried. Clay was chosen because, as a brown, messy material, it was a metaphor for feces and could let the child express either his disgust or aggressivity, or let him build symbolic structures. No interpretation was made during treatment. Four children had no symptoms during 2 months of therapy and no relapse during 1 year of follow-up, one child improved significantly, and one child failed to respond but withdrew from treatment after only three sessions. Modeling clay may be a cheap and effective treatment modality for refractory constipation with encopresis.

Aluminum Silicates↗
PubMed1991

Immunophenotyping of lymphocytes in liver tissue of patients with chronic liver diseases by flow cytometry.

Immunological factors are important in the pathogenesis of a spectrum of hepatobiliary diseases. To characterize the nature of specific immunological responses in liver disease, we determined lymphocyte changes in liver tissue and in blood using flow cytometry. A total of 113 liver biopsy specimens was collected from patients with the following diseases: 19 chronic hepatitis B; 39 chronic non-A, non-B hepatitis; 27 alcoholic liver disease; 10 hepatic malignancy; 8 autoimmune hepatitis; 6 fatty liver and 4 primary biliary cirrhosis. The lymphocytes were isolated from the liver biopsy specimens by mechanical and enzymatic methods. The lymphocyte yield was 7,901 +/- 575 cells/mg of liver tissue. The viability of lymphocytes was 97.7% +/- 0.3%. Lymphocytes were stained with four pairs of two-color mixed fluorescein-conjugated monoclonal antibodies, including T4-T8 (CD4/CD8), T11-B1 (CD2-CD20), NKH1-T8 (CD56-CD8), IL-2R1-T11 (CD25-CD2), and the ratios were determined by an Epics Profile flow cytometer. Immunophenotyping of lymphocytes in whole blood samples was simultaneously analyzed. Variability in lymphocyte yield and different patterns of lymphocyte subsets were found in the liver biopsy specimens. The yields of lymphocytes from patients with chronic non-A, non-B and autoimmune hepatitis were highest, and the lowest yield was from patients with fatty liver. Patients with primary biliary cirrhosis, fatty liver and hepatic malignancy had relatively high ratios of CD4/CD8, CD56/CD8 and CD25/CD2; whereas patients with chronic hepatitis B, autoimmune hepatitis and non-A, non-B hepatitis had lower ratios of CD4/CD8, CD56/CD8 and CD25/CD2. No difference in lymphocyte ratios between the patients with cirrhotic and noncirrhotic alcoholic liver disease was found.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Separation↗
PubMed1990

A study in rat brain cortex synaptic vesicles of endogenous ligands for N-methyl-D-aspartate receptors.

The presence of endogenous ligands for the N-methyl-D-aspartate receptor was looked for in highly purified rat brain cortex synaptic vesicles, the contents of which were extracted and fractionated by gel filtration on Sephadex G-10, or by three different high-voltage electrophoresis procedures. The presence of endogenous ligands was detected by their ability to compete with 50 nM L-[3H]glutamate for binding to whole rat brain N-methyl-D-aspartate receptors. The receptor preparations used were those present in purified postsynaptic densities, in which the quisqualate receptors were blocked by 10 microM quisqualate. Synaptic vesicles had a high content of N-methyl-D-aspartate receptor ligands, which on fractionation always coincided with glutamate or aspartate. A variable and very small amount of a highly acidic endogenous ligand was also found. The latter substance did not coincide in the electrophoresis with homocysteic, cysteic, quinolinic, cysteine sulphinic or homocysteine sulphinic acids, or with N-acetyl-aspartyl-glutamic acid, S-sulphocysteine or sulphoserine. We also found that a single centrifugation, in 0.25 M sucrose, 25 mM Tris-citrate, pH 7.1, of purified synaptic vesicles, at 135,000 gmax for 45 min, led to a 51% loss of endogenous glutamate, but did not change their aspartate content. Thus, in uncentrifuged vesicles the glutamate/aspartate ratio was 9.4, while in centrifuged ones the ratio was 3.9 ATP markedly enhanced L-[3H]glutamate uptake into synaptic vesicles, but did not change the binding of L-[3H]aspartate. Differences in labelled aspartate and glutamate efflux from the vesicles were also found.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗
PubMed1988

Veratridine-induced release of endogenous glutamate from rat brain cortex slices: a reappraisal of the role of calcium.

The efflux of endogenous glutamate from thin slices of rat brain cortex superfused in vitro with artificial cerebrospinal fluid (ACSF) was studied. Initially, glutamate efflux was very high (2.5 nmol/mg protein/min), possibly because of the cutting procedure, but declined sharply, and at 30 min of superfusion was 25 pmol/mg protein/min. In ACSF without added calcium, spontaneous glutamate efflux was always higher than that in calcium-containing medium, e.g. at 30 min it was 75 pmol/mg protein/min. Addition of 10 microM veratridine for 2 min, between 30 and 32 min of superfusion, led, in ACSF with calcium, to an increase in glutamate efflux of 288%, when the maximum efflux following veratridine is compared to the glutamate efflux that immediately preceded the application of this drug (from 25 to 97 pmol/mg protein/min), while in ACSF without added calcium, veratridine induced an increase of only 117% (from 75 to 163 pmol/mg protein/min). These results are interpreted as due to the dual effect of veratridine. In calcium-containing ACSF, veratridine increases sodium influx which depolarizes the neurons and opens voltage-sensitive calcium channels. The increased intraneuronal calcium induces glutamate release from synaptic vesicles, while increased intracellular sodium enhances the release of soluble cytoplasmic glutamate by the reverse operation of the plasma membrane, sodium-dependent glutamate carrier. In ACSF without calcium, the release of vesicular glutamate is suppressed, while the sodium-dependent mechanism remains. This appears as if veratridine-induced glutamate efflux were only partially calcium-dependent.

Animals↗
PubMed1988

Endogenous ligands for the quisqualate receptor: presence in rat brain cortex synaptic vesicles.

The presence in highly purified rat brain cortex synaptic vesicles of endogenous ligands for rat brain quisqualate receptors was investigated. The vesicles were extracted, and their contents fractionated by high voltage electrophoresis. Endogenous ligands were detected by a radioreceptor assay in which such ligands competed with 50 nM L-[3H]glutamate for binding to quisqualate receptors present in rat brain postsynaptic densities (PSDs). Binding of L-[3H]glutamate to N-methyl-D-aspartate (NMDA) receptors, also present in PSDs, was blocked by 100 microM NMDA. We found that the endogenous ligands present in brain cortex synaptic vesicles for quisqualate receptors, were glutamate and aspartate, in a molar ratio of about two to one. The quisqualate receptor had an affinity 130-fold higher for glutamate (Kd 0.3 microM) than for aspartate, and the latter amino acid also showed a marked negative cooperative for binding (Hill number 0.29, against 0.67 for glutamate). These findings suggest that glutamate is the natural transmitter that activates quisqualate receptors at some central excitatory synapses, and also that aspartate may be a classical transmitter, the receptor for which still remains to be shown.

Animals↗