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Biomedical subjects

M A Bosch

Publications and source records attributed to M A Bosch.

At least 19 recordsLinked to original sources
PubMed2002

Role of platelet-activating factor in phosphatidylcholine secretion in primary cultures of rat type II pneumocytes.

This study was designed to investigate the effect of platelet-activating factor (PAF) in the secretory response of type II pneumocytes, that are involved in the synthesis and secretion of the pulmonary surfactant. PAF increased phosphatidylcholine secretion in a concentration-dependent manner in the 10(-5) - 10(-10) M range, with a maximum phosphatidylcholine secretion of up to 3.3 fold the basal values (3.4 +/- 0.3% phosphatidylcholine secreted). This effect was prevented by the synthetic PAF-receptor antagonist WEB 2086. A study of the mechanism through which PAF exerts its stimulatory effect was carried out adding different agents that are well known stimulants of phosphatidylcholine secretion. Thus, PAF increased the TPA- and terbutaline-stimulated phosphatidylcholine secretion, that are PKC and PKA activators respectively, suggesting the involvement of both protein kinases in the process. This involvement was further supported by the use of inhibitors of protein kinases and by the stimulation of cAMP production in type II pneumocytes incubated with PAF.

Animals↗
PubMed2001

Estrogen biphasically modifies hypothalamic GABAergic function concomitantly with negative and positive control of luteinizing hormone release.

The principal role of estrogen is its control of the female ovulatory cycle via negative and positive feedback on gonadotropin secretion. However, a detailed, cohesive picture of how the steroid specifically regulates the excitability of hypothalamic neurons involved in the central control of gonadotropin secretion is still emerging. Here, we used an ovariectomized female guinea pig model to test the hypothesis that estrogen acts on GABAergic neurons in the preoptic area (POA) to elicit a biphasic profile of luteinizing hormone (LH) secretion. Intracellular electrophysiological recordings revealed that estradiol benzoate (EB; 25 microgram, s.c.) decreased the hyperpolarizing response of GABAergic neurons to the GABA(B) receptor agonist baclofen 24 hr after treatment. This effect of GABA(B) receptor stimulation in unidentified POA neurons was still depressed 42 hr after EB administration. By the use of a ribonuclease protection assay, however, EB reduced glutamic acid decarboxylase mRNA expression 42 hr but not 24 hr after its administration. Thus, estrogen attenuated the autoinhibition of GABAergic POA neurons during the initial LH suppressive (i.e., negative feedback) phase and subsequently reduced GABAergic function during the LH surge (i.e., positive feedback). These studies demonstrate that the effects of estrogen on hypothalamic GABAergic neurons coincide with the inhibitory and stimulatory actions, respectively, of the steroid on LH secretion. Furthermore, the data provide novel insights into the mechanism by which estrogen regulates hypothalamic GABAergic neurons, which are critical for the biphasic modulation of LH release observed over the course of the female ovulatory cycle.

Action Potentials↗
PubMed2000

Involvement of calcium in the stimulation of phosphatidylcholine secretion in primary cultures of rat type II pneumocytes by Escherichia coli lipopolysaccharide.

The purpose of this study was to evaluate the mechanism by which Escherichia coli lipopolysaccharide stimulates the secretion of phosphatidylcholine in primary cultures of rat type II pneumocytes. The stimulatory effect of lipopolysaccharide on phosphatidylcholine secretion was additive to those of terbutaline and TPA (protein kinase A and C activators respectively) and this effect was not suppressed by inhibitors of both protein kinases. On the other hand, lipopolysaccharide did not modify the increase on phosphatidylcholine secretion induced by the endoplasmic reticulum Ca2+-ATPase inhibitor thapsigargin, and enhanced slightly the calcium-ionophore A23187 stimulated phosphatidylcholine secretion. In addition, the stimulatory effect of lipopolysaccharide was suppressed by BAPTA, an intracellular Ca2+ chelator, and KN-62, a specific inhibitor of Ca2+-calmodulin-dependent protein kinase. These results, together with the lipopolysaccharide-mediated increase in the cytosolic [Ca2+], suggest that stimulation of phosphatidylcholine secretion by lipopolysaccharide in type II pneumocytes occurs by a calcium-dependent transduction mechanism via Ca2+-calmodulin-dependent protein kinase activation.

Adrenergic beta-Agonists↗
PubMed1999

A powerful GABA(B) receptor-mediated inhibition of GABAergic neurons in arcuate nucleus.

We combined histofluorescence with in situ hybridization to identify GABAergic neurons in the arcuate nucleus (ARC) following electrophysiological recording, using GAD65 as a marker. Intracellular recordings 91 were made in hypothalamic slices prepared from ovariectomized guinea pigs. Over 90% of ARC neurons tested with the GABA(B) receptor agonist baclofen responded with a membrane hyperpolarization or an outward current. The hyperpolarization was dose dependent, and the GABA(B) receptor antagonist CGP 35,348 produced a rightward shift in the agonist dose-response curve. Agonist potency was lower, and the efficacy greater, in GAD-positive neurons. The use of this novel technique for identifying GABAergic neurons thus reveals differences in the pharmacodynamics of GABA(B) receptor activation between GABAergic and non-GABAergic ARC neurons.

Animals↗
PubMed1999

Chronic prenatal cocaine treatment down-regulates mu-opioid receptor mRNA expression in the brain of fetal Rhesus Macaque.

Ribonuclease protection assays (RPA) were performed to quantify mu-opioid receptor mRNA expression in specific brain regions of day 70 Rhesus Macaque fetuses that were exposed to cocaine (3 mg/kg) or saline from days 22-70 of gestation. The content of mu-receptor mRNA was high in the diencephalon and moderate in the mesencephalon. In contrast, mu-receptor mRNA was lightly expressed in areas such as the frontal cortex, striatum and the temporal lobe. The content of mu-opioid receptor mRNA was significantly higher in the diencephalon than in other brain regions (P < 0.001; n = 4). Cocaine exposure significantly decreased the expression of mu-receptor mRNA in the fetal diencephalon (P < 0.05; n = 4 in each group). Our data would indicate that prolonged gestational cocaine exposure causes mu-opioid receptor mRNA down-regulation in specific brain regions of the fetus.

Animals↗
PubMed1998

A powerful GABA(B) receptor-mediated inhibition of GABAergic neurons in the arcuate nucleus.

We combined histofluorescence with in situ hybridization to identify GABAergic neurons in the arcuate nucleus (ARC) following electrophysiological recordings, using GAD65 as a marker. Intracellular recordings were made in hypothalamic slices prepared from ovariectomized guinea pigs. Over 90% of ARC neurons tested with the GABA(B) receptor agonist baclofen responded with a membrane hyperpolarization or an outward current. The hyperpolarization was dose-dependent, and the GABA(B) receptor antagonist CGP 35,348 produced a rightward shift in the agonist dose-response curve. Agonist potency was lower, and the efficacy greater, in GAD-positive neurons. The use of this novel technique for identifying GABAergic neurons thus reveals differences in the pharmacodynamics of GABA(B) receptor activation GABAergic and non-GABAergic ARC neurons.

Animals↗
PubMed1998

The inflammatory cytokines tumor necrosis factor alpha and interleukin-1beta stimulate phosphatidylcholine secretion in primary cultures of rat type II pneumocytes.

Tumor necrosis factor alpha and interleukin-1beta increase surfactant secretion in type II pneumocytes in a time- and dose-dependent manner. This stimulatory effect was additive to that of lipopolysaccharide, suggesting that cytokines and lipopolysaccharide may exert their actions through different signal transduction pathways. Tumor necrosis factor alpha and interleukin-1beta did not modify the increase on phosphatidylcholine secretion induced by the direct protein kinase C activator tetradecanoylphorbol 13-acetate, whereas this effect was inhibited by the protein kinase C inhibitors bisindolylmaleimide (2 x 10(-6) M) and 1-(5-isoquinolinylsulphonyl)-2-methyl piperazone (10(-4) M). In addition, the stimulatory effect of tumor necrosis factor alpha and interleukin-1beta was not suppressed by the intracellular Ca2+ chelator BAPTA (5 x 10(-6) M) or by KN-62 (3 x 10(-5) M), a specific inhibitor of Ca2+-calmodulin-dependent protein kinase. These results suggest that tumor necrosis factor alpha or interleukin-1beta stimulate phosphatidylcholine secretion via protein kinase C activation in a Ca2+-independent manner.

Animals↗
PubMed1998

Glucagon-like peptide-1-(7-36)amide increases pulmonary surfactant secretion through a cyclic adenosine 3',5'-monophosphate-dependent protein kinase mechanism in rat type II pneumocytes.

Glucagon-like peptide-1 (GLP-1) receptor messenger RNA has been identified in cells considered type II pneumocytes that are involved in the synthesis and secretion of the pulmonary surfactant. In an attempt to open new insights into the control of surfactant secretion, we studied the effects of glucagon-related peptides in this process. Accordingly, type II pneumocytes were isolated from Wistar rat lungs and cultured overnight with [methyl-14C]choline, and then the basal and stimulated secretions of [14C]phosphatidylcholine were measured. GLP-1(7-36)amide stimulated phosphatidylcholine secretion in a concentration-dependent manner in the 1-100 nM range; the concentration of the peptide that produced a half-maximal response was 10 nM. Exendin-4 induced similar effects. No changes were observed when GLP-1-(1-37), GLP-2, or exendin-(9-39) was added to the medium. However, the latter reversed the stimulatory effects of GLP-1-(7-36)amide and exendin-4. A study of the mechanism through which GLP-1-(7-36)amide exerts its stimulatory effect was carried out using different agents that are well known stimulants of phosphatidylcholine secretion. GLP-1-(7-36)amide did not produce any change in the stimulatory effect observed with terbutaline or 8-bromo-cAMP, suggesting the involvement of a cAMP-dependent protein kinase in the stimulatory effect of this peptide on phosphatidylcholine secretion. It was further supported by the use of inhibitors of protein kinases and by the stimulation of cAMP production in type II pneumocytes incubated with either GLP-1-(7-36)amide or exendin-4.

Adrenergic beta-Agonists↗
PubMed1997

Impaired phosphatidylcholine biosynthesis and ascorbic acid depletion in lung during lipopolysaccharide-induced endotoxaemia in guinea pigs.

Injection of guinea pigs with a single dose of Escherichia coli lipopolysaccharide (3.2 mg/100 g) induces a reversible endotoxic shock that was evaluated by measuring plasma glucose levels and aspartate aminotransferase activity at 24 h after lipopolysaccharide injection. The hypoglycaemia and the increase in plasma aminotransferase activity observed, correlated with the alterations found during the recovery phase of endotoxic shock. When lipid peroxidation and some antioxidant systems were measured in lungs from treated animals, we only found differences in ascorbic acid content, that was decreased by 50%. Lipopolysaccharide treatment results in a depression of pulmonary phosphatidylcholine synthesis, that correlates with the surfactant deficiencies associated with respiratory illnesses in septic shock. Guinea pigs fed on a diet with a low content in ascorbic acid were more sensitive to endotoxin. In these animals we found no detectable levels of ascorbic acid in lung, whereas both vitamin E lung levels and pulmonary phosphatidylcholine synthesis were significantly decreased. Our results point out the significance of ascorbic acid in the protection against oxidative lung injury associated to endotoxaemia, and validate our shock model for further studies on the mechanisms of this pathological condition.

Animals↗
PubMed1996

Downregulation of mu-opioid receptor mRNA in the mediobasal hypothalamus of the female guinea pig following morphine treatment.

Ribonuclease protection assays were performed on the basal hypothalamus (BH), the preoptic area (POA) and the thalamus (THAL) from female guinea pigs that were ovariectomized and implanted with morphine (n = 8) or placebo (n = 8) pellets for 1 week. An antisense [32P]rUTP labeled riboprobe, representing a 280 bp fragment of the guinea pig mu-opioid receptor gene (spanning putative TM II through eight residues of TM IV), protected a single RNA band of 280 bp. In contrast to the rat, the guinea pig THAL expressed less mu-opioid receptor mRNA than both POA and BH. Morphine treatment caused a significant decrease (15.6 +/- 5.8%) in mu-opioid receptor mRNA expression in the BH, while POA and THAL were not different from placebo controls. Therefore, in conjunction with our previous findings of a downregulation of mu-opioid receptors, mu-opioid receptor mRNA is downregulated in the mediobasal hypothalamus of female guinea pigs following chronic morphine treatment.

Analgesics, Opioid↗
PubMed1995

Effect of Escherichia coli lipopolysaccharide on surfactant secretion in primary cultures of rat type II pneumocytes.

The purpose of this study is to evaluate the effect of the Escherichia coli lipopolysaccharide on the secretion of phosphatidylcholine, the principal component of pulmonary surfactant, in primary cultures of rat alveolar type II pneumocytes. Lipopolysaccharide stimulated phosphatidylcholine secretion in a time- and dose-dependent manner. At a concentration of 200 micrograms/ml, lipopolysaccharide stimulated the release of phosphatidylcholine 4-fold over the basal secretory rate, and the concentration producing half the maximal response was 20 micrograms/ml. The stimulatory effect of lipopolysaccharide on phosphatidylcholine secretion was additive to that of the protein kinase C activator TPA, which is a potent stimulator of surfactant secretion. Lipopolysaccharide did not activate protein kinase C, which suggests that stimulation of phosphatidylcholine secretion by the endotoxin was through a mechanism independent of protein kinase C activation.

Animals↗
PubMed1994

Type II pneumocytes revisited: intracellular membranous systems, surface characteristics, and lamellar body secretion.

BACKGROUND: Type II pneumocytes, the producers of pulmonary surfactant, have been extensively studied during the last 20 years because of the importance of their metabolism in lung function and integrity. The ultrastructural studies of the 1970s and 1980s have shown that these cells present unique elements. EXPERIMENTAL DESIGN: In this work, we used thin-section, freeze-fracture, and fracture-flip electron microscopy techniques to obtain new information on the ultrastructural peculiarities of isolated rat type II pneumocytes, focusing our study on the intracellular membranous systems and their interrelationships and the microanatomy of their plasma membrane during secretory process. RESULTS: In thin-sections of pneumocytes postfixed with osmium tetroxide and potassium ferricyanide, we observed that lamellar bodies (LBs) are usually connected to membranes of the endoplasmic reticulum, and seem to emerge and grow from them. Unusual connections between the endoplasmic reticulum and mitochondria were detected, as well as numerous "bar-like structures" (BLSs), most of them in the early stages of development and often generating from the nuclear membrane. Membranes of the smooth endoplasmic reticulum that closely follow the outlines of mitochondria also appear to be the origin of some BLSs. Possible transition forms, BLS--LB, were also detected, although they were rare. New images of the surface of the pneumocytes and its changes during LB secretion showed a segregation and clearing of membrane particles at the areas of LB extrusion. CONCLUSIONS: We propose that LBs can originate directly from membranes of the endoplasmic reticulum or from BLSs. An indirect participation of mitochondria appears possible. The plasma membrane of pneumocytes displays structural changes associated with the secretion of LBs as visualized by a redistribution of intramembrane and surface particles.

Animals↗
PubMed1993

Effect of dimethylthiourea in phosphatidylcholine biosynthesis by rat lung during reversible endotoxic shock.

A rat model of endotoxic shock was used to evaluate the effects of dimethylthiourea, a putative hydroxyl radical scavenger, in the alterations of lung phosphatidylcholine biosynthesis found during endotoxemia. Treatment of rats with dimethylthiourea, just before lipopolysaccharide injection, resulted in a decreased lipid peroxidation and an increase in phosphatidylcholine biosynthesis, although it did not prevent the body weight loss or the increase in lung weight and lung protein content associated with the lung injury induced by lipopolysaccharide. Our results suggest that phosphatidylcholine biosynthesis is impaired by processes in which hydroxyl radicals are implicated, although other oxygen free radical species, not removed by dimethylthiourea, can be also involved in lipopolysaccharide mediated lung injury.

Animals↗
PubMed1993

Biochemical and electron microscopy analysis of the endotoxin binding to microtubules in vitro.

The mechanisms involved in cellular activation and damage by bacterial endotoxins are not completely defined. In particular, there is little information about possible intracellular targets of endotoxins. Recently, the participation of a microtubule associated protein in endotoxin actions on macrophages has been suggested. In the present work, we have studied the effect of E. coli lipopolysaccharide on the polymerization of microtubular protein in vitro. Electrophoretic analysis of the polymerization mixtures showed that the endotoxin inhibited the polymerization when present at high concentrations. At lower concentrations, LPS selectively displaced the microtubule associated protein MAP-2 from the polymerized microtubules. Electron microscopy showed that LPS binds to microtubules of tubulin + MAPs and to microtubules of purified tubulin (without MAPs) polymerized with taxol. Gel filtration experiments confirmed the binding of LPS to tubulin, and by ligand blot assays an interaction LPS-MAP-2 was detected. The ability of LPS to interact with microtubular proteins suggests a possible participation of microtubules on the cellular effects of endotoxins.

Animals↗
PubMed1993

Visualization of lipopolysaccharide aggregates by freeze-fracture and negative staining.

The morphology of Escherichia coli 0111:B4 lipopolysaccharide (LPS) in aqueous medium was studied by freeze-fracture and negative staining. Samples processed by freeze-fracture showed LPS aggregates that were mainly spherical or elliptical and of rather homogeneous size. Negative staining, however, showed a more heterogeneous population, although globular structures revealed by both procedures had a similar size. Considering the mechanisms involved in the processing of the samples, we suggest that the genuine shape of LPS aggregates is more likely to be globular, thus being artefactual those forms visualized by negative staining, traditionally associated with the organization of LPS in aqueous suspension.

Escherichia coli↗