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Biomedical subjects

P Calvo

Publications and source records attributed to P Calvo.

At least 37 records · Page 2Linked to original sources
PubMed1996

Identification of a long huntingtin mRNA transcript in mouse brain.

Two mRNA species of the Huntington disease (HD) gene that share identical protein coding sequences but differ in their 3' untranslated region have been identified in human. Although a similar situation has been suggested to occur in mouse, only one cDNA has been isolated to date. We report the isolation of a novel partial cDNA of the mouse HD gene that is identical in its protein coding sequence to the previously reported cDNA, although it differs in the distal portion of the 3' untranslated region. Northern blotting assays indicate that this mRNA transcript is preferentially expressed in brain, with highest levels in cerebellum, cerebral cortex and striatum.

Amino Acid Sequence↗
PubMed1996

Identification of four splice variants of the mouse stress-activated protein kinase JNK/SAPK alpha-isoform.

Four splice variants (alpha I-IV) of the stress-activated protein kinase JNK/SAPK alpha-isoform have been identified in the mouse. One of them (alpha I) contains an open reading frame of 1269 bp encoding a potential protein of 423 amino acids, whereas the second variant (alpha II) differs in a region encoding 31 amino acids located in subdomain IX. alpha III lacks this region and also differs in the terminal portion of the 3' untranslated region (3' UTR). A fourth variant (alpha IV) which lacks a region of 41 amino acids located in subdomain IX has also been identified. These splice variants are differentially expressed in mouse tissues: alpha I is the most abundant in brain areas, whereas alpha II is mainly expressed in extracerebral tissues, such as liver; alpha III and alpha IV are present in brain and other tissues although in lower amounts.

Amino Acid Sequence↗
PubMed1996

Effects of an acute dose of ethanol on dopaminergic and serotonergic systems from rat cerebral cortex and striatum.

Intraperitoneal injection 10 min before sacrifice of 1.5 g ethanol/kg weight produced an increase in rat striatal levels of homovanillic acid (HVA) (p < 0.05) but did not affect the striatal concentrations of dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA). A similar ethanol treatment led to decreases in 5-HT (p < 0.05) and 5-HIAA (p < 0.05) from cerebral cortex (prefrontal and anterior cingulate areas). The results point to several ethanol-linked alterations in central serotonergic and dopaminergic systems.

3,4-Dihydroxyphenylacetic Acid↗
PubMed1996

Comparative in vitro evaluation of several colloidal systems, nanoparticles, nanocapsules, and nanoemulsions, as ocular drug carriers.

Three different colloidal carriers, namely, nanoparticles and nanocapsules made of poly-epsilon-caprolactone and submicron emulsions, were designed, and their capacity for increasing the comeal penetration of drugs was investigated. The three systems differed in their inner structure and composition, but they had a similar size (200-250 nm) and a negative superficial charge (-16 to -42 mV). Indomethacin, which was used as a model drug, was dispersed at a molecular level within the colloidal systems, no chemical interaction between the polymer and the drug being detected. Release of the encapsulated indomethacin occurred very rapidly upon high dilution in a buffered medium and was independent of the composition of the system. The in vitro comeal penetration of the encapsulated indomethacin was more than 3-fold that of the commercial eye drops. This increased penetration was similar for the three formulations investigated, which therefore excludes the influence of the inner structure or chemical composition of the colloidal systems on the comeal penetration of indomethacin. Thus, it could be stated that the main factor responsible for the favorable comeal transport of indomethacin is the colloidal nature of these carriers rather than their inner structure or composition.

Animals↗
PubMed1996

Polyester nanocapsules as new topical ocular delivery systems for cyclosporin A.

PURPOSE: Nanocapsules composed of an oily core (Migliol 840) (MG) surrounded by a poly-epsilon-caprolactone (PECL) coat were evaluated as potential vehicles for the topical ocular administration of cyclosporin A (CyA). METHODS: A 2(3) experimental factorial design was applied to optimize the coating of the oily nanodroplets by a solvent displacement technique and to encapsulate a high dose of CyA. The variables investigated were: volume of oil (MG), amount of polymer (PECL), and volume of the organic solvent (acetone) used to dissolve the polymer. RESULTS: Nanocapsules had a mean size in the range of 210-270 nm, a negative zeta potential (between -55 and -60 mV) and a maximum loading capacity of 50% (CyA/PECL ratio). These highly loaded nanocapsules displayed a thick spongeous polymer coating around the oily nanodroplets. The corneal levels of CyA were up to 5 times higher for the encapsulated CyA than for the oily solution of CyA. In addition, these levels remained significantly higher than those of the control group (oily solution) for up to 3 days. Furthermore, the area-under-the-curve (AUC) values were significantly increased for the encapsulated CyA (319.98) with respect to the oily control (74.34). CONCLUSIONS: The CyA-loaded nanocapsules are shown to be interesting vehicles for the improvement of the ocular penetration of CyA.

Administration, Topical↗
PubMed1996

Improved ocular bioavailability of indomethacin by novel ocular drug carriers.

The ability of different drug carriers to improve the ocular bioavailability of drugs was investigated in the rabbit eye. The assayed drug carriers were suspensions of nanoparticles, nanocapsules and microparticles made of poly-epsilon-caprolactone (PECL) and a submicron emulsion. Results indicated that the three submicron systems, nanoparticles, nanocapsules and emulsion, increased more than 3-fold the indomethacin concentration in the cornea, aqueous humour and iris-ciliary body at 0.5 and 1 h post-instillation. Furthermore, an increased indomethacin ocular bioavailability of 300% was observed after instillation of the submicron systems in comparison with the value obtained for a commercial solution. In contrast, the microparticles hardly increased the ocular bioavailability of indomethacin. The mechanism of interaction of the colloidal carriers with the corneal epithelium was investigated by confocal laser scanning microscopy. Confocal images indicated that submicron particles penetrate into the corneal epithelium cells by an endocytic mechanism. The similar behaviour of the three colloidal carriers suggests that any of their specific ingredients (PCEL, lecithin and oil) acts as a penetration enhancer or an endocytotic stimulator. On the other hand, the favourable ocular penetration of indomethacin when encapsulated in the colloidal carriers, but not in the microparticles, led us to assume that the colloidal nature of these carriers is the main factor responsible for the increased ocular bioavailability of indomethacin. PECL nanoparticles and nanocapsules as well as submicron emulsions are shown to be novel corneal drug carriers, thus representing a useful approach for increasing the ocular bioavailability of drugs.

Analysis of Variance↗
PubMed1995

Immunodetection of serotonin transporter from mouse brain.

A DNA fragment encoding amino acid sequences from the amino terminal region of the mouse serotonin transporter was isolated and sequenced. This transporter is widely distributed throughout the mouse brain, as deduced by heminested reverse transcription-polymerase chain reaction (RT-PCR) assays. To identify the serotonin transporter protein, we have developed specific antibodies against a fusion protein containing its amino terminal region, a domain which shows a low degree of homology between the different neurotransmitter transporters. Western blot analysis of mouse brain membranes detected the serotonin transporter as a 71 kDa polypeptide.

Amino Acid Sequence↗
PubMed1995

An autoradiographical saturation kinetic study of the different benzodiazepine binding sites in rat brain by using [3H] flunitrazepam as a radioligand.

The present study reports on the equilibrium association constant (KD) and receptor density (Bmax) values of a number of brain areas from the mesencephalon, cerebral cortex, hippocampus, and cerebellum for the overall benzodiazepine (BZ) binding sites as well as for benzodiazepine binding site subtype 1 (BZ1) and subtype 2 (BZ2), determined by autoradiographical procedures using [3H] flunitrazepam. The differences between BZ1 and BZ2 binding sites were analyzed using the specific BZ1 agonist zolpidem as inhibitor of the radioligand. Statistically significant differences in the affinities of BZ2 with respect to BZ and BZ1 binding site were mainly found in cortical layers when pKD (negative logarithms of KD values) values were compared (ANOVA-SNK test). The distribution of Bmax, as well as the percentages of BZ1 and BZ2 and Hill coefficients which, surprisingly, are always close to 1 (> 0.9) for all the saturation kinetics analyzed, are also described. The possibility of heterogeneity related to anatomical distribution in the different subtypes is discussed.

Analysis of Variance↗
PubMed1995

Immunodetection of the large form of the gamma 2 subunit of mammalian GABAA receptor.

Two forms of the GABAA receptor gamma 2-subunit, named short (gamma 2S) and long (gamma 2L), and generated by alternative RNA splicing, have been identified in mammalian brain by molecular cloning techniques. We have produced antibodies against a synthetic peptide containing the 8-amino acid insertion present in the long form but not in the short one. Using the antipeptide serum, we have identified the gamma 2L subunit in membrane preparations of GABAA receptors from rat and mouse cerebellum and its cellular location in cerebellum.

Alternative Splicing↗
PubMed1995

Identification of a cyclic adenosine 3',5'-monophosphate-dependent protein kinase phosphorylation site in the carboxy terminal tail of human D1 dopamine receptor.

Each of the dopamine receptor subtypes contains several consensus sites for phosphorylation in their intracellular domains. We have used fusion proteins of the carboxy terminal tail of D1 and D5 dopamine receptors to study the phosphorylation of these proteins by cyclic adenosine 3',5' monophosphate (cAMP)-dependent protein kinase (PKA) and protein kinase C (PKC). The fusion protein of D1 dopamine receptor was efficiently phosphorylated by PKA, but not by PKC. Site-directed mutagenesis of serine 380 to an alanine residue precluded the phosphorylation by the kinase. No phosphorylation of the D5 dopamine receptor fusion protein was observed with either PKA or PKC, which indicates that these receptor subtypes might differ in their mechanisms of regulation.

Amino Acid Sequence↗
PubMed1995

Autoradiographical study of types 1 and 2 of benzodiazepine receptors in rat brain after chronic ethanol treatment and its withdrawal.

The effect of chronic ethanol treatment, and its withdrawal on benzodiazepine binding sites in rat brain is described in this autoradiographical study using the benzodiazepine agonist [3H]flunitrazepam. Several areas of the rat cerebral cortex, hippocampus, mesencephalon, cerebellum and lateral geniculate nucleus were studied in animals, chronically treated with ethanol, and 24 or 48 hr after ethanol withdrawal. The [3H]flunitrazepam binding and the relative percentages of binding to BZ1 and BZ2 sites, using zolpidem as a BZ1 site inhibitor, are described. The cerebellum and red nucleus, which only express BZ1 binding sites, appear to be areas significantly modified by ethanol and its withdrawal. In no other structure did ethanol modify [3H]flunitrazepam binding nor change the relative percentage of BZ1 and BZ2 sites.

Animals↗
PubMed1995

Effect of ethanol treatment on rate and equilibrium constants for [3H] muscimol binding to rat brain membranes: alteration of two affinity states of the GABAA receptor.

Equilibrium binding curves were biphasic in control and ethanol-treated rats. [3H]Muscimol binds to sites of high (KDA of approximately 10 nM) and low (KDB of approximately 0.3-0.4 microM) affinity. Chronic ethanol treatment produced a decrease in BmaxA value, and the hyperbolic binding profiles were progressively affected by the chronic and in vitro ethanol treatments, with most of this effect corresponding to the high-affinity site. IC50 and Ki values were calculated for several competing ligands, using membranes from both control and ethanol-treated animals. The association and dissociation curves were also biphasic, using a radioligand concentration precluding a significant occupancy of the low-affinity sites, which suggests the existence of two forms or affinity states of the monoliganded receptor. Chronic ethanol treatment did not produce changes in the values of the dissociation rate constants (fast and slow phases). By contrast, we report for the first time a decrease in the values of the association rate constants, with this decrease being higher for the slow phase. Consequently, the dissociation equilibrium constants are two times higher in chronically ethanol-treated animals for both phases.

Animals↗
PubMed1995

Molecular cloning of alpha 1d-adrenergic receptor and tissue distribution of three alpha 1-adrenergic receptor subtypes in mouse.

A partial cDNA encoding most of the third intracellular loop of the mouse alpha 1d-adrenergic receptor subtype was amplified from hippocampus by reverse transcription-polymerase chain reaction (RT-PCR) using degenerate oligodeoxynucleotide primers. This DNA fragment was used as a probe to isolate an alpha 1d-adrenergic receptor cDNA from a mouse brain cDNA library. The deduced amino acid sequence encodes a potential protein of 562 amino acids, and northern hybridization of poly(A)+ RNA isolated from mouse brain detected a single 3.0-kb transcript. Partial cDNA fragments of the alpha 1b- and alpha 1a-adrenergic receptor subtypes were also amplified from mouse brain and sequenced. Analysis of the mRNA expression by RT-PCR indicated that the alpha 1-adrenergic receptors are widely distributed in mouse tissues. The alpha 1d subtype is expressed in brain areas such as hippocampus, striatum, and brainstem and also in many extracerebral tissues, such as lung, liver, heart, kidney, and spleen. The alpha 1a subtype is also expressed in many tissues, whereas the alpha 1b subtype has a more restricted expression, with high levels in striatum, brainstem, and diencephalus.

Amino Acid Sequence↗
PubMed1995

Pneumonia caused by Listeria monocytogenes.

Pneumonia due to Listeria monocytogenes is extremely uncommon. We report the case of an 87-year-old woman with no underlying immunosuppressive disease who presented with listerial pneumonia. Cutaneous anergy and a decrease in total lymphocyte count in this elderly woman could predispose her to listerial infection.

Adult↗
PubMed1994

Effect of morphine and abstinence syndrome on [3H]bromoxidine binding to alpha 2-adrenoceptors in rat brain.

At 4 days after the implantation of two subcutaneous 75 mg morphine pellets in the back skin, rats were morphine-dependent. In the three layers studied in the occipital cortex we found that the values of the alpha 2-adrenergic agonist [3H]bromoxidine binding increased with respect to animals implanted with placebo pellets. Typical behavioral and physiological symptoms of the abstinence syndrome appeared 30 minutes after administration of naloxone, [3H]bromoxidine binding values being similar to those obtained in animals implanted with placebo pellets. The pattern of response of the [3H]bromoxidine binding was similar in the hippocampus and the superficial gray layer of the superior colliculus of the mesencephalon, but the differences were not statistically significant in these areas. This paper concludes that exist brain regional differences in the alpha 2-adrenoceptors response under morphine-treatment and possibly under naloxone-induced morphine abstinence syndrome.

Adrenergic alpha-Agonists↗
PubMed1994

Effect of chronic treatment with ethanol and withdrawal of ethanol on binding of [3H]SCH23390 to D1 dopamine receptor in rat visual cortex and hippocampus. An autoradiographic study.

Male Wistar rats, treated with ethanol for 8 weeks and pair-control animals, were used to study the effects of chronic treatment with ethanol, and withdrawal of ethanol for 24 and 48 hr on [3H]SCH23390 binding. The visual cortex (Laminae III-IV and Lamina VI), the superficial grey layer of the superior colliculus, and the molecular layer of the dentate gyrus of the hippocampus were the cerebral areas analysed. Non significant changes were observed in hippocampus and Laminae III-IV of the visual cortex after treatments with alcohol. More interesting results were obtained from Lamina VI, where the chronic treatment with ethanol did not modify the binding of [3H]SCH23390, whereas the withdrawal of ethanol produced a statistically significant increase in binding values. In addition, superficial grey layer of the superior colliculus showed a significant increase in binding values between 48 hr withdrawal and ethanol treated groups. The results herein reported suggest that some structures involved in visual functions are related to responses of adaptation to ethanol.

Animals↗