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E Cherubini

Publications and source records attributed to E Cherubini.

At least 19 recordsLinked to original sources
PubMed1992

Single-channel currents of NMDA type activated by L- and D-homocysteic acid in cerebellar granule cells in culture.

The whole-cell and the outside-out configurations of the patch-clamp technique were used to study macroscopic and single-channel currents evoked by the enantiomers of homocysteic acid, the sulphur containing analogue of glutamate, in cerebellar granule cells in culture. L-Homocysteic acid (L-HC, 15 microM) and D-homocysteic acid (D-HC, 50 microM) induced whole-cell currents of comparable amplitude, that were abolished by the N-methyl-D-aspartate (NMDA) receptor antagonist 2-amino-5-phosphonovalerate (AP-5) (20 microM). AMPA (100 microM) induced whole-cell currents which were not modified by AP-5 (20 microM) but were blocked by CNQX. In the outside-out configuration, both L-HC and D-HC (15 microM) elicited single-channel currents of the same conductance, mean open time and reversal potential as the NMDA-induced single-channel events. In the presence of Mg2+ (2 mM), D- and L-HC-induced single-channel currents were voltage-dependent. These data suggest that in cerebellar granule cells in culture, both L-HC and D-HC activate the same NMDA receptor channel complex.

Animals↗
PubMed1992

Inactivation characteristics of a sustained, Ca(2+)-independent K+ current of rat hippocampal neurones in vitro.

1. Current or voltage clamp recordings from CA3 neurones of the adult rat hippocampal slice were performed to study the inactivation properties of a slow outward K+ current identified as the delayed rectifier (IK). 2. In current clamp experiments, burst firing evoked from resting membrane potential by intracellular current injection was reduced or blocked by conditioning hyperpolarizing pre-pulses of 20-40 mV amplitude. This effect was inhibited by tetraethylammonium (TEA; 20 mM) but was unaffected by Cs+ (3 mM), 4-aminopyridine (4-AP; 2 mM), carbachol (30-50 microM), mast cell degranulating peptide (MCDP; 300 nM), thyrotrophin releasing hormone (TRH; 1 microM) or by a Ca(2+)-free solution containing Mn2+ or Co2+ (2 mM). 3. Single-electrode voltage clamp experiments were carried out on neurones superfused with Ca(2+)-free solution, containing tetrodotoxin (TTX; 1 microM), Mn2+ or Co2+ (2 mM), 4-AP (2 mM), Cs+ (3 mM) and carbachol (30 microM). Step depolarizations from a holding potential of -55 mV activated an outward current which reached a plateau after 200 ms, followed by an outward tail current. Such an outward current had the characteristics of IK. 4. The outward currents were significantly potentiated by conditioning hyperpolarizing pre-pulses suggesting the IK was reduced by a voltage-dependent inactivation process. Removal of inactivation was a function of the amplitude of the conditioning hyperpolarizing pre-pulse. At a holding potential of -55 mV removal of inactivation was time dependent with a time constant of 211 ms. High K+ (12.5 or 21.5 mM) solutions did not affect the inactivation characteristics of IK. 5. Tetraethylammonium (20 mM) or low concentrations of Ba2+ (0.1 mM) readily depressed the outward current without significantly affecting the inactivation process. Dendrotoxin (200 nM) also depressed such a slow current but, in addition, increased the inactivation process of IK. 6. It is suggested that removal of inactivation of IK by hyperpolarization can modulate cell excitability by fully restoring the ability of IK to inhibit burst firing of CA3 hippocampal neurones.

Animals↗
PubMed1991

N-methyl-D-aspartate receptor-mediated spontaneous activity in cerebellar granule cells in culture.

An outside-out configuration of the patch-clamp method was used to study the properties of spontaneous and glutamate-evoked single channel activity in cerebellar granule cells in culture. Both spontaneous occurring events and glutamate-evoked single channel currents had similar conductances (17.7, 36.5; 17.3, 35.8 pS) and mean open times (0.7, 7.1; 0.6, 7.8 ms). In magnesium free solution, the spontaneous channels were blocked by the NMDA-receptor antagonist APV (0.1 mM) and by the NMDA-channel blocker MK-80 (0.1 mM). The spontaneous activity disappeared at negative holding potentials when the bathing solution contained magnesium (5 mM). Maximum conductance of spontaneous channels increased with days in culture (DIC) from 9 pS at 1 DIC to 59 pS at 10 DIC. These experiments suggest that endogenous transmitter activates NMDA-receptors in cerebellar granule cells in culture.

2-Amino-5-phosphonovalerate↗
PubMed1991

L-homocysteic acid mediates synaptic excitation at NMDA receptors in the hippocampus.

beta-p-Chlorophenylglutamate (Chlorpheg), a specific L-homocysteate (L-HC) uptake blocker, was tested on the L-HC- and L-glutamate-induced currents and on the excitatory postsynaptic potentials (EPSPs) evoked in CA1 rat hippocampal neurons by Schaffer collaterals stimulation. In the presence of tetrodotoxin (TTX; 1 microM), Chlorpheg (0.5-2 mM) potentiated L-HC- but not L-glutamate-induced currents. In normal magnesium containing medium and at resting membrane potential, Chlorpheg (1.5-1 mM) increased the amplitude and duration of the EPSPs evoked by Schaffer collaterals stimulation. This effect was prevented by bath application of the N-methyl-D-aspartate (NMDA) receptor antagonist CPP (20 microM). Chlorpheg enhanced also the NMDA component of the EPSP, evoked in the presence of 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 10 microM), bicuculline (20 microM) and glycine (100 microM). This effect was blocked by CPP (20 microM). It is concluded that L-HC is an endogenous NMDA agonist at the Schaffer collateral-CA1 synapse.

6-Cyano-7-nitroquinoxaline-2,3-dione↗
PubMed1991

GABA: an excitatory transmitter in early postnatal life.

In the adult mammalian CNS, GABA is the main inhibitory transmitter. It inhibits neuronal firing by increasing a Cl- conductance. Bicuculline blocks this effect and induces interictal discharges. A different picture is present in neonatal hippocampal neurones, where synaptically released or exogenously applied GABA depolarizes and excites neuronal membranes--an effect that is due to a different Cl- gradient. In fact, during the early neonatal period, GABA acting on GABAA receptors provides most of the excitatory drive, whereas excitatory glutamatergic synapses are quiescent. It is suggested that during development GABA exerts mainly a trophic action through membrane depolarization and a rise in intracellular Ca2+.

Animals↗
PubMed1991

Depression of a sustained calcium current by kainate in rat hippocampal neurones in vitro.

1. High-threshold, slow inactivating inward Ca2+ currents were studied in CA1 pyramidal neurones from rat hippocampal slices using the single-electrode voltage clamp technique. 2. Kainate (50-400 nM) induced a dose-dependent depression of the amplitude of the slow Ca2+ current. At a dose of 200 nM the current amplitude was reduced from -0.63 +/- -0.06 to -0.32 +/- 0.06 nA. Such an effect of kainate was associated with the development of a small inward current (-0.11 +/- 0.03 nA). Kynurenic acid (1 mM) or 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 20 microM) fully prevented these actions of kainate. 3. The structurally related kainate analogue alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA; 200 nM) depressed the slow Ca2+ current by 30 +/- 7%, an effect also blocked by CNQX. 4. In low-Na+ medium slow Ca2+ currents were followed by sustained inward tail currents. Kainate reduced both the steady-state Ca2+ current (from -0.98 +/- 0.14 to -0.63 +/- 0.15 nA) and the tail current (from -0.40 +/- 0.04 to -0.14 +/- 0.03 nA). 5. The inactivation process of the slow Ca2+ current was tested by a double-pulse protocol and was found to be enhanced by kainate. 6. Equimolar replacement of Ca2+ by Ba2+ produced larger inward currents followed by prolonged tails. Kainate reduced the Ba2+ steady-state current from -1.77 +/- 0.18 to -1.44 +/- 0.24 nA and the tail current from -0.47 +/- 0.15 to -0.17 +/- 0.05 nA. 7. In current clamp experiments Ca2+ action potentials were recorded from cells loaded with the Ca2+ chelator BAPTA. In these conditions kainate failed to reduce the Ca2+ action potential, while in the absence of BAPTA kainate shortened the Ca2+ action potentials by 30%. 8. It is suggested that low concentrations of kainate reduced the slow Ca2+ current by promoting its inactivation perhaps through a rise in free intracellular Ca2+.

Animals↗
PubMed1991

Persistent pulsatile release of glutamate induced by N-methyl-D-aspartate in neonatal rat hippocampal neurones.

1. Intracellular recordings were made from CA3 hippocampal neurones in vitro, during the first ten days of postnatal life and in adulthood. 2. Repeated (three to six) applications of N-methyl-D-aspartate (NMDA), in the presence of tetrodotoxin (TTX, 1-3 microM) and K+ channel blockers (tetraethylammonium chloride or bromide (TEA), 10 mM, and Cs+, 2 mM; or 4-aminopyridine (4-AP), 30-50 microM, and Cs+, 2 mM) induced in neonatal but not in adult neurones, periodic inward currents (PICs) which persisted for several hours after the last application of NMDA. 3. PICs which were due to non-specific cation currents had a frequency of 0.10 +/- 0.04 Hz, and an amplitude of 1.1 +/- 0.28 nA at holding potentials between -40 and -50 mV. The amplitude was a linear function of the membrane potential over the range -70 to +20 mV. They reversed polarity at 4.1 +/- 9.8 mV. 4. K+ channel blockers alone failed to induce PICs. Repeated (three to six) brief applications of high (12 mM) K+ medium also induced PICs. The frequency and amplitude of K(+)-induced PICs were however considerably reduced by concomitant applications of the NMDA receptor antagonist D,L-3-[( +/- )-2-carboxypiperazin-4-yl-]propyl-1-phosphonic acid (CPP, 20 microM). PICs could be induced also by caffeine (1 mM) in the presence of the phosphodiesterase inhibitor 3-isobutyl-1-methyl-xanthine (IBMX, 200 microM), TTX, TEA and Cs+. 5. Intracellular injection of the calcium chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA) did not prevent the induction of PICs by NMDA. However PICs were blocked by removal of the external calcium and by the calcium antagonists cobalt (2 mM) and cadmium (50 microM). 6. In spite of blockade of propagated synaptic activity by TTX, PICs were synchronous in a pair of intracellularly recorded cells. They were also synchronous with extracellular spikes recorded by electrodes located into stratum pyramidal or stratum radiatum. 7. Once established, PICs were unaffected by NMDA receptor antagonists D(-)2-amino-5-phosphonovaleric acid (AP-5, 50 microM), CPP (20 microM) and the NMDA channel blocker ketamine (10 microM). They were reversibly blocked by the broad spectrum excitatory amino acid antagonist kynurenic acid (1 mM) and by the selective non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10 microM). 8. It is concluded that PICs are generated in neonatal neurones by a synchronous, pulsatile release of glutamate from presynaptic nerve terminals, secondary to oscillations in intracellular calcium.

Action Potentials↗
PubMed1991

Strychnine-sensitive glycine responses of neonatal rat hippocampal neurones.

1. Intracellular recordings employing current and voltage clamp techniques were used to study the effects of glycine on rat CA3 hippocampal neurones during the first 3 weeks of postnatal (P) life. 2. Glycine (0.3-1 mM) depolarized neurones from rats less than 4 days old (P4). Neurones from older neonates (P5-P7) were hyperpolarized by glycine, whereas adult neurones were unaffected. 3. Both depolarizing and hyperpolarizing responses were associated with large conductance increases; they reversed polarity at a potential which changed with the extracellular chloride concentration. The responses persisted in tetrodotoxin (1 microM) or in a solution with a much reduced calcium concentration. 4. Strychnine (1 microM) but not bicuculline (10-50 microM) antagonized the effects of glycine. The action of strychnine was apparently competitive with a dissociation constant of 350 nM. 5. In voltage clamp experiments, glycine elicited a non-desensitizing outward current at -60 mV. When a maximal concentration of glycine was applied at the same time as gamma-aminobutyric acid (GABA), the conductance increase induced by the two agonists was additive, suggesting the activation of different populations of channels. 6. Concentrations of glycine lower than 100 microM did not affect membrane potential. However, at 30-50 microM glycine increased the frequency of spontaneous GABA-mediated synaptic responses; this action was not blocked by strychnine. 7. It is concluded that during the first 2 weeks of life glycine acts at strychnine-sensitive receptors to open chloride channels.

Animals↗
PubMed1990

Inactivation of a slow Ca2+ current in CA1 neurones of the adult rat hippocampal slice.

CA1 neurones of the adult rat hippocampal slice preparation were voltage clamped at or near -40 mV membrane potential using a single electrode clamp method. Depolarizing voltage commands from a holding potential of -40 mV elicited voltage-dependent inward Ca2+ currents comprising a fast and a slow component. The latter one was investigated for its susceptibility to inactivation, which was maximally expressed at around 0 mV membrane potential. When extracellular Ca2+ was replaced by Ba2+, inward currents became much larger and were followed by long tail currents. Similar data were observed in neurones injected with the Ca2+ chelator BAPTA. It is suggested that inactivation of the slow Ca2+ current depends at least partly on the levels of intracellular free Ca2+ in hippocampal neurones.

Animals↗
PubMed1990

Submicromolar concentrations of zinc irreversibly reduce a calcium-dependent potassium current in rat hippocampal neurons in vitro.

The action of the endogenous divalent cation zinc on Ca2+ and Ca2(+)-dependent currents was studied in rat hippocampal CA1 and CA3 neurons in vitro, by means of a single electrode voltage clamp technique. Bath application of zinc (0.5-1 microM) produced a small membrane depolarization associated with an increase in synaptic noise and cell excitability and a depression of the afterhyperpolarization following a train of action potentials. The effects on the afterhyperpolarization, could not be reversed on washout. In voltage-clamped neurons, zinc induced a steady inward current and reduced, at resting membrane potential, the peak amplitude of the outward current underlying the afterhyperpolarization, IAHP. In caesium loaded neurons (in the presence of tetrodotoxin and tetraethylammonium), zinc reduced the slow inactivating Ca2+ current activated from a holding potential of -40 mV. Similar results were observed with nickel and cobalt at comparable concentrations, with Zn2+ greater than Ni2+ greater than Co2+, in their order of potency. In contrast to nickel and cobalt the effects of zinc did not reverse on washout. These results suggest that low concentrations of zinc enhance cell excitability by reducing IAHP. In addition, zinc reduces the slow inactivating voltage-dependent Ca2+ current. The irreversible effect of this metal ion is compatible with a toxic, intracellular site of action.

Animals↗
PubMed1990

GABA mediated excitation in immature rat CA3 hippocampal neurons.

Intracellular recordings from rat hippocampal neurons in vitro during the first postnatal week revealed the presence of spontaneous giant depolarizing potentials (GDPs). These were generated by the synchronous discharge of a population of neurons. GDPs reversed polarity at -27 and -51 mV when recorded with KCl or K-methylsulphate filled electrodes, respectively. GDPs were blocked by the GABAA receptor antagonist bicuculline (10 microM). Iontophoretic or bath applications of GABA (10-300 microM) in the presence of tetrodotoxin (1 microM), induced a membrane depolarization or in voltage clamp experiments an inward current which reversed polarity at the same potential as GDPs. The response to GABA was blocked in a non-competitive manner by bicuculline (10 microM) and did not desensitize. GABA mediated GDPs were presynaptically modulated by N-methyl-D-aspartate (NMDA) and non-NMDA receptors. Their frequency was reduced or blocked by NMDA receptor antagonists and by the rather specific non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). The frequency of GDPs was enhanced by glycine and D-serine (10-30 microM) in a strychnine insensitive manner. This effect was blocked by AP-5, suggesting that it was mediated by the allosteric modulatory site of the NMDA receptor. These observations suggest that most of the 'excitatory' drive in immature neurons is mediated by GABA acting on GABAA receptors; furthermore excitatory amino acids modulate the release of GABA by a presynaptic action on GABAergic interneurons.

Animals↗
PubMed1990

GABAergic mechanisms in the CA3 hippocampal region during early postnatal life.

The developmental pattern of GABAergic neurons in the rat hippocampus during the first week of postnatal life shows several particularities both from a morphological and physiological point of view: (1) GABA immunoreactive neurons which are initially localized in a deep and superficial layer, progressively disappear from these two layers. From the end of the first postnatal week, GABAergic neuronal somata appear throughout the whole hippocampus, but GABA immunoreactive terminal structures are not frequent until the second postnatal week. (2) Intracellular observations in slices reveal the presence in CA3 pyramidal neurons between P0 and P6 (postnatal days) of spontaneous giant depolarizing potentials (GDPs); these are mediated by GABA acting on GABAA receptors and modulated presynaptically by NMDA receptors. During this period of development, GABA and GABAA analogues have a depolarizing action at resting membrane potential. Bicuculline at this developmental stage blocks completely spontaneous and evoked synaptic potentials. During the second postnatal week, when GABA responses shift from depolarizing to hyperpolarizing, bicuculline induces spontaneous interictal discharges. It is suggested that the positive feedback of the GABAergic interneuron on the pyramidal neuron during the first week of life may account for the generation of GDPS which may play an important role in synaptogenesis.

Action Potentials↗
PubMed1990

The allosteric glycine site of the N-methyl-D-aspartate receptor modulates GABAergic-mediated synaptic events in neonatal rat CA3 hippocampal neurons.

We report in this study that, in the presence of magnesium, bath application of micromolar concentrations of glycine have prominent effects on synaptic events and N-methyl-D-aspartate (NMDA) responses in neonatal but not in adult hippocampal slices. Intracellular recordings were made from 71 rat CA3 hippocampal neurons in neonatal slices. In keeping with our earlier study, during the first postnatal week, CA3 neurons exhibited giant depolarizing potentials (GDPs). These GDPs are mediated by gamma-aminobutyric acid (GABA) acting on type A GABA (GABAA) receptors and modulated presynaptically by NMDA receptors. In the majority of cells (18 out of 31), glycine (10-30 microM) increased the frequency of GDPs (from 0.14 to 0.29 Hz). This effect was mimicked by D-serine (10-20 microM) and blocked by the NMDA receptor antagonists D-(-)-2-amino-5-phosphonovalerate (50 microM) and DL-2-amino-7-phosphonoheptanoate (50 microM) and by the GABAA antagonist bicuculline (10 microM) but not by strychnine (1 microM). Subthreshold concentrations of glycine (or D-serine) and NMDA, when given together, enhanced synaptic noise and the frequency of GDPs. In the presence of tetrodotoxin (1 microM), glycine and D-serine (up to 50 microM) did not modify the NMDA-induced inward currents in CA3 pyramidal cells. However the reduction of NMDA-mediated currents by 7-chlorokynurenate (10-20 microM) was reversed by glycine and D-serine (100-200 microM). In contrast, glycine (up to 100 microM) had no effect on membrane potential, input resistance, or NMDA responses after postnatal day 10. It is concluded that GABA-mediated events are facilitated by glycine acting on presynaptically located NMDA receptors.

Aging↗