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Biomedical subjects

M G Giovannini

Publications and source records attributed to M G Giovannini.

At least 19 recordsLinked to original sources
PubMed1991

Release of endogenous glutamate and aspartate from the frog spinal cord in vitro.

The release of the endogenous excitatory amino acids aspartate and glutamate from an in vitro slice preparation of the frog spinal cord was investigated by using high performance liquid chromatography (HPLC). Spinal slices were incubated at 4 degrees C to minimize amino acid uptake; samples of the bathing solution were collected at 10 min intervals for amino acid assay. In each experiment electrophysiological responses were recorded continuously from ventral roots after stimulation of an adjacent dorsal root, so that changes in the profile of amino acid release could be correlated with electrophysiological responses. At rest the release of glutamate and aspartate was 52.9 +/- 5.8 and 66.9 +/- 5.6 pmol/10 min, respectively, and was unaffected by low Ca2+ media or tetrodotoxin. After trains of high frequency stimulation applied to a dorsal root the release of glutamate and aspartate was significantly and reproducibly enhanced by 33 +/- 13 and 49 +/- 18%, respectively. The stimulus-dependent release was blocked by low Ca2+ media, tetrodotoxin or topical application of the neuropeptide thyrotropin releasing hormone. The present study provides direct evidence to support the role of glutamate and aspartate as neurotransmitters in the spinal cord.

Animals↗
PubMed1991

A decrease in brain catecholamines prevents oxiracetam antagonism of the effects of scopolamine on memory and brain acetylcholine.

The effect of oxiracetam on passive avoidance conditioned response and acetylcholine (ACh) levels in rats with selective lesions of the central monoaminergic pathways was investigated. The lesions were followed by a marked decrease in cortical serotonin (-88%), noradrenaline (-54%) and striatal dopamine (-57%) levels, while neither the performance of a passive avoidance conditioned response nor brain ACh levels were affected. Scopolamine (hyoscine) administration (0.63 mg/kg, s.c.) to lesioned rats exerted the expected amnesic effect, associated with a decrease in hippocampal, cortical and striatal ACh levels. In the rats with degeneration of dopaminergic and noradrenergic but not serotoninergic pathways, oxiracetam (50 and 100 mg/kg, s.c.) was unable to prevent both amnesia and the decrease in brain ACh levels caused by scopolamine. The effect of oxiracetam was prevented by haloperidol (0.2 mg/kg, s.c.). Our findings support the hypothesis that an interaction between monoaminergic and cholinergic neurotransmitter systems may be involved in the actions of nootropic drugs on cognitive functions.

Acetylcholine↗
PubMed1991

Effect of thyrotropin releasing hormone (TRH) on acetylcholine release from different brain areas investigated by microdialysis.

1. The effect of thyrotropin releasing hormone (TRH) administration upon acetylcholine (ACh) release in freely moving rats was investigated by means of transversal microdialysis coupled to h.p.l.c. TRH administered either s.c. or via local perfusion increased the ACh release from cortex and hippocampus but not from the striatum. The increase in ACh release was maintained after 7 days of s.c. administration of TRH. 2. After s.c. injection of the neuropeptide, the increase in ACh release was dose-dependent and reached a maximum at 40 min after administration. The maximal percentage increases were 18, 52, 66 and 89% at doses of 1, 2.5, 5 and 10 mg kg-1 and 35, 48 and 54% at doses of 2.5, 5 and 10 mg kg-1 in the cortex and hippocampus, respectively. The effect of TRH was dependent on neuronal activity since it was completely inhibited by perfusion with tetrodotoxin (TTX), 5 X 10(-7) M. 3. Perfusion with TRH, 2.5 micrograms microliters-1, caused 198% and 150% increase in ACh release 60 and 80 min after the beginning of the perfusion in the cortex and hippocampus, respectively. After this initial peak, a 100% increase in ACh release persisted throughout the perfusion. 4. Systemic TRH administration was followed by marked hyperactivity and stereotyped behaviour that showed a time course shorter than that of the increase in ACh release. 5. These findings demonstrate that TRH exerts a strong stimulant action on cortical and hippocampal cholinergic pathways.

Acetylcholine↗
PubMed1991

Isotope dilution mass spectrometry: definitive methods and reference materials in clinical chemistry.

The application of gas chromatography-mass spectrometry (GC-MS) for the quantitative analysis of endogenous compounds as well as drugs in clinical biochemistry is taken into account. For this purpose, it is of the utmost importance the choice of a good internal standard (IS). It is demonstrated that the best IS in GC-MS is a stable isotope-labelled analogue of the analyte. In this case only we can speak of isotope dilution mass spectrometry. All the advantages and disadvantages inherent to the use of such a molecule as IS are discussed. The use of a calibration curve to perform the quantitative analysis is described, together with the criteria applied for the definition of the reliability of a method (precision, accuracy and limit of quantification). Examples taken from the literature are described.

Chemistry, Clinical↗
PubMed1989

Cholinergic and noradrenergic denervations decrease labelled purine release from electrically stimulated rat cortical slices.

The origin of cortical purine release was investigated by measuring [3H]purine release from electrically stimulated cortical slices of rats after neurotoxic lesions of cholinergic, noradrenergic and serotoninergic pathways innervating the cortex. Purines were labelled by incubating the cortical slices with [3H]adenine. The 3H efflux at rest and during stimulation, analysed by high performance liquid chromatography, consisted of adenosine, inosine, hypoxanthine and a small amount of nucleotides. Twenty days after unilateral or bilateral lesion of the nucleus basalis a marked decrease in choline acetyltransferase activity was associated with a decrease in [3H]purine release. A linear relationship was found between the decrease in choline acetyltransferase activity and [3H]purine release. A partial recovery in both choline acetyltransferase activity and [3H]purine release was observed eight months after the lesion. Twenty days after intra-cerebroventricular injection of 6-hydroxydopamine a 59% decrease in cortical noradrenaline content was associated with a 44% decrease in [3H]purine release. Conversely, no change in [3H]purine release was found in rats in which a 89% decrease in cortical serotonin content was induced by intra-cerebroventricular injection of 5,7-dihydroxytryptamine. The decrease in [3H]purine release after the lesion of the cholinergic and noradrenergic pathways may depend on metabolic changes, a loss of a stimulating influence of acetylcholine and noradrenaline or may indicate a release of [3H]purine from cholinergic and noradrenergic fibres.

Adenosine↗
PubMed1989

The relationship between the behavioral effects of cognition-enhancing drugs and brain acetylcholine. Nootropic drugs and brain acetylcholine.

The effect of different doses of the cognition-enhancing agent oxiracetam on the disruption of acquisition of a passive avoidance conditioned response (step-through) by scopolamine 0.6 mg/kg s.c., and on the impairment of performance by scopolamine 0.2 mg/kg s.c. in an eight-arm maze was investigated in adult male Wistar rats. At the end of the experiments, ACh levels were measured in the cerebral cortex and hippocampus by an HPLC method, in order to ascertain whether a relationship exists between the effects of the two drugs on behavior and cholinergic function. In the step-through test, oxiracetam at doses of 50 and 100 mg/kg s. c., given 30 min. before scopolamine, antagonized both scopolamine-induced behavioral disruption and decrease in ACh levels but was inactive at the dose of 300 mg/kg s. c. In the eight-arm maze test, oxiracetam prevented scopolamine-induced disruption of the performance and decrease in brain ACh at the dose of 30 mg/kg s. c. but was inactive at the dose of 100 mg/kg s. c. In conclusion, when scopolamininduced disruption of behavior is prevented or reduced, the scopolamininduced decrease in ACh level, particularly in the hippocampus, is also reduced. The present results therefore offer further confirmation of a relationship between the cognition-enhancing effects of oxiracetam and its effects on hippocampal and cortical cholinergic mechanisms.

Acetylcholine↗
PubMed1989

S-aryl (tetramethyl) isothiouronium salts as possible antimicrobial agents, IV.

The 1-octanol/water partition coefficients of a series of antimicrobial S-aryl(tetramethyl)isothiouronium salts, were determined. The concentration of the solute in the two phases was measured by means of HPLC. The values of the hydrophobic parameters thus obtained were correlated to the corresponding capacity factors (log Kw) and compared with those expected on the basis of additive-constitutive principles. The deviations were interpreted quantitatively on the basis of electronic interactions of the substituents in the benzene ring. Equations correlating structural features and antimicrobial activity of some studied compounds by means of molecular parameters were formulated.

Anti-Bacterial Agents↗
PubMed1988

The cDNA sequence coding for prepro-PGS (prepro-magainins) and aspects of the processing of this prepro-polypeptide.

Amphibian skin is well known as a source of peptides homologous to bioactive peptides found in mammalian gut and brain. A systematic investigation of the skin secretions from Xenopus laevis revealed several peptides not derivable from known precursors. The sequence elucidation, utilizing fast atom bombardment/mass spectrometry, of two peptides, PGS and PGS Gly-10;Lys-22, is reported. These have been independently characterized and named magainins and found to have antimicrobial activity. A mixed sequence oligonucleotide probe complementary to the mRNA sequence coding for PGS was synthesized and used to screen a Xenopus skin cDNA library. A full length cDNA species encoding prepro-PGS was isolated and characterized, and its sequence is reported. The deduced precursor sequence was found to contain one copy of PGS Gly-10;Lys-22 and five copies of PGS. The proteolytic processing of this prepro-polypeptide is discussed.

Amino Acid Sequence↗
PubMed1988

Levitide, a neurohormone-like peptide from the skin of Xenopus laevis. Peptide and peptide precursor cDNA sequences.

A novel peptide, levitide, less than Glu-Gly-Met-Ile-Gly-Thr-Leu-Thr-Ser-Lys-Arg-Ile-Lys-Gln-NH2 has been isolated from skin secretions of the South African frog Xenopus laevis and sequenced by fast atom bombardment mass spectrometry. Synthetic oligonucleotides were used as probes to screen a X. laevis skin cDNA library for species coding for preprolevitide. Two such clones were detected and their sequences are reported here. Preprolevitide is 88 residues long, exhibits a putative signal sequence at the amino terminus, and contains the levitide peptide at the carboxyl terminus. The levitide precursor shows a striking nucleotide and amino acid (86%) sequence homology with the precursor of xenopsin, a biologically active octapeptide from Xenopus skin, and also encodes a 25-residue amphipathic peptide that is released by processing at a single arginine residue.

Amino Acid Sequence↗
PubMed1988

Purinergic modulation of cortical acetylcholine release is decreased in aging rats.

The effect of adenosine, N-ethylcarboxamide adenosine (NECA), and caffeine on acetylcholine (ACh) release was investigated in cortical slices prepared from 3 and 22-24-month-old rats. The slices were perfused with Krebs solution and electrically stimulated at 0.2, 1, and 5 Hz stimulation frequency. In old rats, ACh released by stimulation at 1 and 5 Hz was about half as large as in adult rats. In 22-24-month-old rats, the potency of adenosine was strongly reduced, and a similar significant inhibition of ACh release was obtained with concentrations of 1 microM adenosine in adult and 300 microM in old rats. Conversely, NECA, which has no effect on ACh release in adult rats, brought about a 40% decrease in old rats. Caffeine at 50 microM concentration enhanced, and at 500 microM inhibited, the evoked ACh release in adult rats, but was inactive in old rats. The possibility is envisaged that aging may modify purinergic modulation of ACh release by inducing conformational changes in purinergic receptors or changing adenosine metabolism.

Acetylcholine↗
PubMed1988

S-Aryl(tetramethyl)isothiouronium salts as possible antimicrobial agents--III.

A series of S-Aryl(tetramethyl)isothiouronium salts were prepared and evaluated in vitro for antimicrobial activity. Some compounds revealed interesting inhibiting action on Gram-positive bacteria which is noteworthy in view of the large number of strains antagonized and of the low MIC values. The possible influence of decomposition kinetics to the corresponding mercaptoderivatives was intestigated and compared with those of salts previously studied.

Anti-Bacterial Agents↗
PubMed1987

Biosynthesis and degradation of peptides derived from Xenopus laevis prohormones.

Peptides present in the skin secretion of the South African frog, Xenopus laevis, have been analysed by fast atom bombardment mass spectrometry and h.p.l.c. in the mass range 500-3200 Da. We have investigated the effects of successive glandular secretions induced by noradrenaline injections on these peptide levels and have found that the replenishment of the whole range of peptides is complete within 2-6 days. Intact secretory vesicles free of cellular contaminants contain a relatively large number of peptides with molecular masses in the range 2400-2700 Da. We have termed these peptides primary products or spacer peptides, since they originate from spacer regions of the precursors to xenopsin and caerulein. However, if the secretory vesicles are disrupted during the collection procedure and the solution containing the secretion is kept at room temperature for up to 2 h, relatively little of the larger peptides remain. By comparing the relative levels of the various peptides present in these secretions we have found that the larger peptides are proteolytically cleaved into smaller fragments by a novel cleavage at the N-terminal side of a lysine residue (at Xaa-Lys bonds where Xaa is Leu, Gly, Ala or Lys). Preliminary evidence has been obtained suggesting that the larger intact peptides possess lytic activity whereas the smaller proteolytic fragments appear relatively inactive. This may represent a mechanism by which the secretions are rendered harmless to the frog itself, since prolonged exposure would be expected to result in toxic effects. The dorsal glands of X. laevis thus appear similar to endocrine glands, since they are involved in peptide biosynthesis, secretion and subsequent proteolytic degradation.

Amino Acid Sequence↗
PubMed1987

Determination of testosterone and its tissue metabolites (DHT and 3 alpha-diol) in human plasma and prostatic tissue by isotopic dilution mass spectrometry.

5 alpha-Dihydrotestosterone has been widely measured in human prostatic tissue using RIA since it is involved in the pathogenesis of human prostatic hyperplasia and seems to be the best index for the follow-up of patients affected by prostatic cancer under endocrine treatment. A GC-MS method for the simultaneous determination of testosterone (T), 5 alpha-dihydrotestosterone (DHT) and 5 alpha-androstan-3 alpha, 17 beta-diol (3 alpha-diol) in prostatic tissue based on the isotopic dilution technique was developed. Tri-deuterated internal standards of each compound were previously synthetized in our laboratory. After extraction and purification on Sep-Pak C18 and Sephadex LH-20, T and its metabolites were measured as heptafluorobutyric ester (HFB) derivatives. Quantitative analysis was performed on a VG 7070 EQ mass spectromer equipped with a fused silica capillary column using the Selected Ion Monitoring technique. Steroid values (mean +/- SD; ng/g tissue) found in nine human hypertrophic prostates were: T: 0.71 +/- 0.43; DHT: 4.46 +/- 1.41; 3 alpha-diol: 0.34 +/- 0.23. Preliminary results obtained from the detection of the three androgens in human prostatic hyperplasia treated for 3 months with GnRH before surgery seem to indicate that DHT concentration decreases more than 10 times. Values obtained (n = 1; ng/g tissue) were: T: 0.194; DHT: 0.255; 3 alpha-diol: 0.015.

Androstenediol↗
PubMed1987

Effect of pyroglutamic acid stereoisomers on ECS and scopolamine-induced memory disruption and brain acetylcholine levels in the rat.

The acquisition of a passive avoidance conditioned response was disrupted in the rat by electroconvulsive shock (ECS) and scopolamine administration. D,L-pyroglutamic acid (D,L-PCA) 500 and 1000 mg/kg, administered as arginine salt 120 min before the retest, prevented both the ECS and scopolamine-induced amnesia. Arginine alone was ineffective. Scopolamine brought about a 52 and 39% decrease, respectively, in cortical and hippocampal acetylcholine (ACh) levels, measured by means of a gas-chromatographic method. D,L-PCA 500 and 1000 mg/kg also prevented the decrease in brain ACh level. When the two isomers were studied separately, D-PCA was more effective than L-PCA and antagonized scopolamine-induced amnesia at the doses of 250 and 500 mg/kg. In conclusion, D,L-PCA is active on cortical and hippocampal cholinergic mechanisms and, like other 2-oxopyrrolidone derivatives, shows cognition-enhancing properties.

Acetylcholine↗
PubMed1986

Isothermal gas chromatographic determination of nanogram amounts of chlorimipramine, chlorpromazine and their N-desmethyl metabolites in plasma using nitrogen-selective detection.

A gas chromatographic method using nitrogen-selective detection for the quantitative determination of nanogram amounts of chlorimipramine, chlorpromazine and their nor1 and nor2 derivatives in plasma is described. Derivatization with trifluoroacetic anhydride of nor1 and nor2 metabolites allowed the chromatographic separation of these compounds. A three-step solvent extraction procedure was performed using n-heptane containing 1% isoamyl alcohol and n-hexane and compared with a plasma clean-up procedure using C18 Sep-Pak cartridges. The two procedures were characterized by similar degrees of precision. The use of C18 Sep-Pak cartridges, however, produced a significant time and material saving over the conventional extraction method.

Chlorpromazine↗
PubMed1985

Phosphatidylserine increases acetylcholine release from cortical slices in aged rats.

Acetylcholine release was investigated in cortical slices superfused with choline-enriched Krebs solution containing physostigmine. Slices were prepared from 3 and 24 month old rats treated with either Tris buffer or sonicated suspensions of phosphatidylserine and phosphatidylcholine in Tris buffer. Slices were electrically stimulated at frequencies of 1, 2 and 5 Hz for 5 min periods preceded and followed by rest periods. ACh content of the superfusate was quantified by bioassay. In the 24 month old rats treated with Tris buffer, acetylcholine release, at all frequencies tested, was approximately 50% lower than that in the 3 month old rats. On the contrary, no significant decrease in ACh release was found in the 24 month old rats treated for 30 days with phosphatidylserine (15 mg/kg IP). The same treatment did not increase acetylcholine release in 3 month old rats. Acetylcholine release in 24 month old rats receiving a single administration of phosphatidylserine (15 mg/kg IP) or phosphatidylcholine (15 mg/kg IP) for 30 days was as low as in the 24 month old rats receiving the Tris buffer only. It is proposed that the chronic phosphatidylserine treatment may reduce the age-induced decrease in acetylcholine release by acting on the stimulus-secretion coupling mechanism.

Acetylcholine↗
PubMed1985

Pregnanediol-3 alpha-glucuronide measured in diluted urine by mass spectrometry with fast atom bombardment/negative-ion ionization.

We describe a mass-spectrometric method based on the fast atom bombardment ionization technique in the negative-ion mode for measuring pregnanediol-3 alpha-glucuronide in diluted urine from women. The procedure requires addition of testosterone-17 beta-D-glucuronide (2.5 micrograms/25 microL) to the urine sample as internal standard, and the sample is added directly to the fast atom bombardment target with no further manipulation. We have assessed and evaluated the method by the traditional criteria of reliability.

Adult↗