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P Calabresi

Publications and source records attributed to P Calabresi.

At least 91 records · Page 5Linked to original sources
PubMed1998

Endogenous ACh enhances striatal NMDA-responses via M1-like muscarinic receptors and PKC activation.

Cortical glutamatergic fibres and cholinergic inputs arising from large aspiny interneurons converge on striatal spiny neurons and play a major role in the control of motor activity. We have investigated the interaction between excitatory amino acids and acetylcholine (ACh) on striatal spiny neurons by utilizing intracellular recordings, both in current- and in voltage-clamp mode in rat brain slices. Muscarine (0.3-10 microM) produced a reversible and dose-dependent increase in the membrane depolarizations/inward currents induced by brief applications of N-methyl-D-aspartate (NMDA), while it did not affect the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)-induced responses. These concentrations of muscarine did not alter the membrane potential and the current-voltage relationship of the recorded cells. Neostigmine (0.3-10 microM), an ACh-esterase inhibitor, mimicked this facilitatory effect. The facilitatory effects of muscarine and neostigmine were antagonized either by scopolamine (3 microM) or by pirenzepine (10-100 nM), an antagonist of M1-like muscarinic receptors, but not by methoctramine (300 nM), an antagonist of M2-like muscarinic receptor. Accordingly, these facilitatory effects were mimicked by McN-A-343 (1-10 microM), an agonist of M1-like muscarinic receptors, but not by oxotremorine (300 nM), an agonist of M2-like receptors. Tetrodotoxin (TTX) did not block the facilitatory effect produced by the activation of muscarinic receptors suggesting that this effect is postsynaptically mediated. The action of neostigmine was prevented either by the intracellular calcium (Ca2+) chelator BAPTA (200 mM) or by preincubating the slices with inhibitors of protein kinase C (PKC) (staurosporine 100 nM or calphostin C 1 microM). McN-A-343 did not alter the excitatory post synaptic potentials (EPSPs) evoked by corticostriatal stimulation in the presence of physiological concentration of magnesium (Mg2+ 1.2 mM), while it enhanced the duration of these EPSPs recorded in the absence of external magnesium. Our data show that endogenous striatal ACh exerts a positive modulatory action on NMDA responses via M1-like muscarinic receptors and PKC activation.

(4-(m-Chlorophenylcarbamoyloxy)-2-butynyl)trimethy↗
PubMed1998

Blockade of M2-like muscarinic receptors enhances long-term potentiation at corticostriatal synapses.

Acetylcholine (ACh) exerts a crucial role in learning and memory. The striatum contains the highest concentration of this transmitter in the brain. This structure expresses two different forms of synaptic plasticity, long-term depression (LTD) and long-term potentiation (LTP), which might contribute to the storage of motor skills and some cognitive processes. We have investigated the role of M2-like muscarinic receptors in striatal LTP by utilizing intracellular recordings in vitro from a rat corticostriatal slice preparation. Methoctramine (250 nM), an antagonist of M2-like muscarinic receptors, enhanced striatal LTP induced in the absence of external magnesium (Mg2+) by high-frequency stimulation (HFS) of corticostriatal fibres. Methoctramine did not affect the amplitude of excitatory postsynaptic potentials (EPSPs) when bath applied either before or after the conditioning tetanus suggesting that a critical increase of ACh concentrations is produced only during HFS. Methoctramine per se failed to enhance the NMDA-mediated EPSPs recorded in the absence of external Mg2+ and in the presence of 10 microM CNQX. Methoctramine antagonized the presynaptic inhibitory action of neostigmine, an inhibitor of ACh-esterase, and oxotremorine, an agonist of M2-like muscarinic receptors. These data indicate that the activation of M2-like muscarinic receptors exerts a negative influence on striatal LTP, probably by reducing the release of glutamate from corticostriatal fibres and they suggest a complex modulatory effect of ACh in striatal synaptic plasticity.

Acetylcholine↗
PubMed1998

Benzylacyclouridine enhances 5-fluorouracil cytotoxicity against human prostate cancer cell lines.

At a nontoxic growth inhibitory concentration benzyloxyacyclouridine (BAU), a potent and specific inhibitor of uridine phosphorylase (UrdPase), enhanced 5-fluorouracil (5-FU) cytotoxic activity against human prostate cancer PC-3 and DU-145 cell lines. The BAU/5-FU combination exhibited greater antitumor activity in vivo using PC-3 human xenografts compared to 5-FU alone, with no associated increase in animal host toxicity. The mechanism(s) responsible for the enhanced in vitro and in vivo activity of this combination may involve enhanced formation of the 5-FU nucleotide metabolites FdUMP, FdUTP, and FUTP resulting in enhanced inhibition of thymidylate synthase (TS) and increased incorporation of fluoropyrimidine metabolites into tumoral RNA and DNA.

Animals↗
PubMed1998

Predictive sensitivity of human cancer cells in vivo using semipermeable polysulfone fibers.

An in vivo experimental model was developed to predict efficiently and accurately chemosensitivity of human tumors. Human cancer cells either from cultured cell lines or from patients' tumors were injected directly into semipermeable polysulfone fibers subsequently implanted into immunocompetent rats. Results suggest utility of this novel model system for predicting tumor sensitivity to a wide range of anticancer agents and for potentially guiding the treatment of cancer patients in the clinical setting.

Animals↗
PubMed1998

L-type Ca2+ channel blockers attenuate electrical changes and Ca2+ rise induced by oxygen/glucose deprivation in cortical neurons.

BACKGROUND AND PURPOSE: Experimental evidence supports a major role of increased intracellular calcium [Ca2+]i levels in the induction of neuronal damage during cerebral ischemia. However, the source of Ca2+ rise has not been fully elucidated. To clarify further the role and the origin of Ca2+ in cerebral ischemia, we have studied the effects of various pharmacological agents in an in vitro model of oxygen (O2)/glucose deprivation. METHODS: Pyramidal cortical neurons were intracellularly recorded from a slice preparation. Electrophysiological recordings and microfluorometric measurements of [Ca2+]i were performed simultaneously in slices perfused with a glucose-free physiological medium equilibrated with a 95% N2/5% CO2 gas mixture. RESULTS: Eight to twelve minutes of O2/glucose deprivation induced an initial membrane hyperpolarization, followed by a delayed, large but reversible membrane depolarization. The depolarization phase was accompanied by a transient increase in [Ca2+]i levels. When O2/glucose deprivation exceeded 13 to 15 minutes, both membrane depolarization and [Ca2+]i rise became irreversible. The dihydropyridines nifedipine and nimodipine significantly reduced either the membrane depolarization or the [Ca2+]i elevation. In contrast, tetrodotoxin had no effect on either of these parameters. Likewise, antagonists of ionotropic and group I and II metabotropic glutamate receptors failed to reduce the depolarization of the cell membrane and the [Ca2+]i accumulation. Finally, dantrolene, blocker of intracellular Ca2+ release, did not reduce both electrical and [Ca2+]i changes caused by O2/glucose depletion. CONCLUSIONS: This work supports a role of L-type Ca2+ channels both in the electrical and ionic changes occurring during the early phases of O2/glucose deprivation.

Animals↗
PubMed1998

High-dose intravenous methotrexate for patients with nonleukemic leptomeningeal cancer: is intrathecal chemotherapy necessary?

PURPOSE: Standard treatments for neoplastic meningitis are only modestly effective and are associated with significant morbidity. Isolated reports suggest that concurrent systemic and intrathecal (i.t.) therapy may be more effective than i.t. therapy alone. We present our experience, which includes CSF and serum pharmacokinetic data, on the use of high-dose (HD) intravenous (i.v.) methotrexate (MTX) as the sole treatment for neoplastic meningitis. PATIENTS AND METHODS: Sixteen patients with solid-tumor neoplastic meningitis received one to four courses (mean, 2.3 courses) of HD (8 g/m2 over 4 hours) i.v. MTX and leucovorin rescue. Serum and CSF MTX concentrations were measured daily. Toxicity, response, and survival were retrospectively compared with a reference group of 15 patients treated with standard i.t. MTX during the same time interval. RESULTS: Peak methotrexate concentrations ranged from 3.7 to 55 micromol/L (mean, 17.1 micromol/L) in CSF and 178 to 1,700 micromol/L (mean, 779 micromol/L) in serum. Cytotoxic CSF and serum MTX concentrations were maintained much longer than with i.t. dosing. Toxicity was minimal. Cytologic clearing was seen in 81% of patients compared with 60% of patients treated intrathecally (P = .3). Median survival in the HD i.v. MTX group was 13.8 months versus 2.3 months in the i.t. MTX group (P = .003). CONCLUSION: HD i.v. MTX is easily administered and well tolerated. This regimen achieves prolonged cytotoxic serum MTX concentrations and CSF concentrations at least comparable to those achieved with standard i.t. therapy. Cytologic clearing and survival may be superior in patients treated with HD i.v. MTX. Prospective studies and a reconsideration of the use of i.t. chemotherapy for patients with neoplastic meningitis are warranted.

Adult↗
PubMed1998

Phase I clinical and pharmacological studies of benzylacyclouridine, a uridine phosphorylase inhibitor.

Benzylacyclouridine (BAU, IND 039655) is a potent and specific inhibitor of uridine phosphorylase (UrdPase; EC 2.4.2.3). This enzyme plays a major role in regulating uridine homeostasis and also catalyzes the conversion of fluoropyrimidine nucleosides to their respective bases. Inhibition of UrdPase enzyme activity 18-24 h after 5-fluorouracil (5-FU) administration increased plasma levels of uridine and enhanced the therapeutic index of 5-FU by rescuing normal tissues. Moreover, in vitro preclinical studies have also shown that inhibiting UrdPase enzyme activity by BAU prior to administration of 5-FU increased cytotoxicity in a number of human cancer cell lines. A series of preclinical studies was performed in dogs and pigs to evaluate the pharmacological and pharmacodynamic properties of BAU. These data showed a sustained elevation in plasma uridine concentration in both animal models. The rapid degradation of a tracer dose of uridine into uracil was virtually arrested by BAU administered both p.o. or i.v. The t1/2 of BAU was 1.8-3.6 h in dogs, with bioavailability levels of 85% (30 mg/kg) and 42.5% (120 mg/kg). In pigs, the half-life varied from 1.6 to 2.3 h, with a bioavailability of 40% at 120 mg/kg. The drug was distributed into most tissues with a tissue: plasma ratio of approximately 0.7. On the basis of these preclinical studies, we performed a Phase I clinical trial of BAU in patients with advanced cancer. Patients received 200, 400, 800, and 1600 mg/m2 BAU as a single oral dose. Toxicities included grade 2 anemia, grade 1 fever, grade 1 fatigue, grade 1 constipation, and grade 1 elevation in alkaline phosphatase; none of these toxicities were observed to be dose dependent. The maximum tolerated dose and dose-limiting toxicity were not reached at the doses given. BAU plasma concentrations and area under the curve correlated linearly with the oral dose level. The pharmacokinetics of BAU were consistent with a first-order clearance, with average peak concentrations ranging from 19 microM (200 mg/m2) to 99 microM (1600 mg/m2) and tbeta1/2 ranging from 3.0 to 3.9 h at the four dose levels. Compared with baseline plasma uridine, treatment of patients with 200, 400, 800, and 1600 mg/m2 BAU increased peak uridine concentrations by 120, 150, 250, and 175%, respectively. On the basis of this clinical study, the suggested Phase II starting dose of BAU in combination with 5-FU is 800 mg/m2. Studies combining BAU with 5-FU and incorporating appropriate molecular and biochemical end points to assess the effects of this drug combination on tumor and/or surrogate tumor tissue are under way.

Aged↗
PubMed1997

A sensitive ELISPOT assay to detect low-frequency human T lymphocytes.

We extended the sensitivity of the ELISPOT assay by including an antigen-driven proliferation step prior to a final restimulation with antigen and irradiated antigen presenting cells (APCs). This improved sensitivity made the modified ELISPOT assay better suited to the detection of rare or low frequency T lymphocytes than the standard ELISPOT assay or alternatives such as limiting dilution analysis or in situ hybridization. Use of ELISA-grade plastic or polyvinylidene difluoride (PVDF) plates for the detection of different cytokines improved the signal-to-noise ratio for counting cytokine spots, and use of video computer imaging software improved objective quantitation. Analysis of antigen-reactive peripheral blood mononuclear cells (PBMC) from multiple sclerosis (MS) patients using both the traditional and our modified ELISPOT assay demonstrate a > 10-fold increase in numbers of myelin basic protein (MBP)-responsive T cells detected (an average of less than 1 spot forming cell (SFC) per 2 x 10(5) PBMC with the standard assay compared to 19 SFC per 2 x 10(5) PBMC with the modified assay). In addition, the modified ELISPOT assay could be performed with frozen PBMC, which permitted greater flexibility in sample processing, multiple use of a single sample as an internal standard, and simultaneous analysis of samples collected at different time points. This modified ELISPOT assay has many applications, including analysis of cytokine profiles in rare T cell populations, identification of antigen-responsive individuals as PBMC donors for T lymphocyte cloning or for therapeutic intervention, and assessment of vaccine or therapeutic efficacy as a surrogate clinical marker.

Amino Acid Sequence↗
PubMed1997

Endogenous adenosine mediates the presynaptic inhibition induced by aglycemia at corticostriatal synapses.

Energy deprivation, as a result of aglycemia, leads to depression of the central synaptic transmission. Endogenous adenosine has been implicated in this depressant effect. We have studied the possible involvement of endogenous adenosine in the depression of corticostriatal excitatory transmission induced by glucose deprivation by using intracellular recordings in brain slices. After stimulation of corticostriatal fibers, EPSPs were recorded from striatal spiny neurons. Adenosine (3-300 microM) or brief periods (5-10 min) of aglycemia reduced the EPSP amplitude but did not alter the membrane potential and the resistance of the recorded cells. These inhibitory effects were not associated with an alteration of the postsynaptic sensitivity to exogenous glutamate but were coupled with an increased paired-pulse facilitation, suggesting the involvement of presynaptic mechanisms. A delayed postsynaptic membrane depolarization/inward current was detected after 15-20 min of glucose deprivation. The presynaptic inhibitory effects induced by adenosine and aglycemia were both antagonized either by the nonselective adenosine receptor antagonist caffeine (2.5 mM) or by the A1 receptor antagonists 8-cyclopentyl-1,3-dimethylxanthine (CPT, 1 microM) and 1,3-dipropyl-8-cyclopentylxanthine (CPX, 300 nM). Conversely, these antagonists affected neither the delayed membrane depolarization/inward current nor the underlying conductance increase produced by glucose deprivation. The ATP-sensitive potassium channel blockers tolbutamide (1 mM) and glipizide (100 nM) had no effect on the aglycemia-induced decrease of EPSP amplitude. Our data demonstrate that endogenous adenosine acting on A1 receptors mediates the presynaptic inhibition induced by aglycemia at corticostriatal synapses, whereas ATP-dependent potassium channels do not play a significant role in this presynaptic inhibition.

Adenosine↗
PubMed1997

Abnormal synaptic plasticity in the striatum of mice lacking dopamine D2 receptors.

Dopamine D2 receptors (D2Rs) are of crucial importance in the striatal processing of motor information received from the cortex. Disruption of the D2R gene function in mice results in a severe locomotor impairment. This phenotype has analogies with Parkinson's disease symptoms. D2R-null mice were used to investigate the role of this receptor in the generation of striatal synaptic plasticity. Tetanic stimulation of corticostriatal fibers produced long-term depression (LTD) of EPSPs in slices from wild-type (WT) mice. Strikingly, recordings from D2R-null mice showed the converse: long-term potentiation (LTP). This LTP, unlike LTD, was blocked by an NMDA receptor antagonist. In magnesium-free medium, LTP was also revealed in WT mice and found to be enhanced by L-sulpiride, a D2R antagonist, whereas it was reversed into LTD by LY 17555, a D2R agonist. In D2R-null mice this modulation was lost. Thus, our study indicates that D2Rs play a key role in mechanisms underlying the direction of long-term changes in synaptic efficacy in the striatum. It also shows that an imbalance between D2R and NMDA receptor activity induces altered synaptic plasticity at corticostriatal synapses. This abnormal synaptic plasticity might cause the movement disorders observed in Parkinson's disease.

2-Amino-5-phosphonovalerate↗
PubMed1997

Hypoxia in striatal and cortical neurones: membrane potential and Ca2+ measurements.

Simultaneous measurements of membrane potential and intracellular Ca2+ were used to study the effects of hypoxia on striatal and cortical neurones. Striatal neurones responded to hypoxia with a reversible membrane depolarization coupled with a transient increase in intracellular Ca2+. Thirty minutes of hypoxia caused an irreversible membrane depolarization associated with a massive raise in Ca2+ levels, leading to cell death. Conversely, cortical neurones were more resistant to O2 deprivation. Hypoxia (4-10 min) induced minimal changes in both membrane potential and Ca2+ signals. Longer periods (20-30 min) caused an initial membrane hyperpolarization followed by a large but reversible depolarization coupled with a transient increase in Ca2+ signals. These results support the hypothesis of a differential sensitivity of central neurones to hypoxia, suggesting that striatal neurones are more vulnerable than cortical cells.

Animals↗
PubMed1997

Opposite membrane potential changes induced by glucose deprivation in striatal spiny neurons and in large aspiny interneurons.

We have studied the electrophysiological effects of glucose deprivation on morphologically identified striatal neurons recorded from a corticostriatal slice preparation. The large majority of the recorded cells were spiny neurons and responded to aglycemia with a slow membrane depolarization coupled with a reduction of the input resistance. In voltage-clamp experiments aglycemia caused an inward current. This current was associated with a conductance increase and reversed at -40 mV. The aglycemia-induced membrane depolarization was not affected by tetrodotoxin (TTX) or 6-cyano-7-nitroquinoxaline-2,3-dione plus aminophosphonovalerate, antagonists acting respectively on AMPA and NMDA glutamate receptors. Also, the intracellular injection of bis(2-aminophenoxy)ethane-N,N, N',N'-tetra-acetic acid, a calcium (Ca2+) chelator, and low Ca2+/high Mg2+-containing solutions failed to reduce this phenomenon. Conversely, it was reduced by lowering external sodium (Na+) concentration. A minority of the recorded cells had the morphological characteristics of large aspiny interneurons and the electrophysiological properties of "long-lasting afterhyperpolarization (LA) cells." These cells responded to aglycemia with a membrane hyperpolarization/outward current that was coupled with an increased conductance. This current was not altered by TTX, blockers of ATP-dependent potassium (K+) channels, and adenosine A1 receptor antagonists, whereas it was reduced by solutions containing low Ca2+/high Mg2+. This current reversed at -105 mV and was blocked by barium, suggesting the involvement of a K+ conductance. We suggest that the opposite membrane responses of striatal neuronal subtypes to glucose deprivation might account for their differential neuronal vulnerability to aglycemia and ischemia.

Animals↗
PubMed1997

Activation of group III metabotropic glutamate receptors depresses glutamatergic transmission at corticostriatal synapse.

Intracellular recordings were performed from a rat corticostriatal slice preparation in order to characterize the effects of group III metabotropic glutamate receptor (mGluR) agonists on excitatory transmission at corticostriatal synapses. The amplitude of excitatory postsynaptic potentials (EPSPs), evoked by cortical stimulation, was significantly decreased by agonists acting at group III metabotropic glutamate receptors. Both L-2-amino-4-phosphonobutanoate (L-AP4) and L-serine-O-phosphate (L-SOP) were effective in reducing the amplitude of cortically evoked EPSPs, in a dose-dependent manner. The EC50 value for the effect of L-SOP and L-AP4 was 0.89 microM and 9.95 microM, respectively. Both L-AP4 and L-SOP had negligible effects on the intrinsic membrane properties of the recorded neurons and did not alter the postsynaptic response to focal application of glutamate, suggesting a presynaptic site of action. The presynaptic inhibition of both L-SOP and L-AP4 was fully antagonized by 250 microM (s)-2-methyl-2-amino-4-phosphonobutanoate (MAP4), whilst it was unaffected by 500 microM RS-methyl-4-carboxyphenylglycine (MCPG). Conversely, the presynaptic inhibitory effect on the EPSP amplitude exerted by 10 microM 1S,3R-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-ACPD) was antagonized by 500 microM MCPG, whilst it was not blocked by 250 microM MAP4. Finally, the reduction of the EPSP amplitude produced by a saturating dose of L-SOP was further increased by 10 microM 1S,3R-ACPD, suggesting an additive effect of these compounds. The present results are consistent with the idea that group III mGluRs exert a presynaptic inhibitory modulation of the excitatory glutamatergic transmission at corticostriatal synapses.

Aminobutyrates↗
PubMed1997

Synaptic plasticity and physiological interactions between dopamine and glutamate in the striatum.

Several electrophysiological studies have addressed the interaction between glutamate and dopamine within the striatum. Although the results obtained from these studies were often conflicting, more recently the characterization of new forms of synaptic plasticity in the basal ganglia provided a possible integrative explanation of the different electrophysiological data regarding the interaction between these transmitters. In this review we will try to summarize and discuss the available data concerning the possible impact of the functional role of D1 and D2 receptor activation on the modulation of the glutamatergic corticostriatal pathway. Moreover, we will also describe the function of the striatum in the integration of glutamatergic and dopaminergic inputs to produce long-term changes of synaptic efficacy (long-term depression, long-term potentiation). Finally, we will consider the implication of the interaction between dopamine and glutamate in the regulation of energetic metabolism whose failure is responsible for neuronal death.

Animals↗