Biomedical subjects

Y Kovalchuk

Publications and source records attributed to Y Kovalchuk.

9 recordsLinked to original sources

PubMed2004

Distinct roles of Galpha(q) and Galpha11 for Purkinje cell signaling and motor behavior.

G-protein-coupled metabotropic glutamate group I receptors (mGluR1s) mediate synaptic transmission and plasticity in Purkinje cells and, therefore, critically determine cerebellar motor control and learning. Purkinje cells express two members of the G-protein G(q) family, namely G(q) and G11. Although in vitro coexpression of mGluR1 with either Galpha11 or Galpha(q) produces equally well functioning signaling cascades, Galpha(q)- and Galpha11-deficient mice exhibit distinct alterations in motor coordination. By using whole-cell recordings and Ca2+ imaging in Purkinje cells, we show that Galpha(q) is required for mGluR-dependent synaptic transmission and for long-term depression (LTD). Galpha11 has no detectable contribution for synaptic transmission but also contributes to LTD. Quantitative single-cell RT-PCR analyses in Purkinje cells demonstrate a more than 10-fold stronger expression of Galpha(q) versus Galpha11. Our findings suggest an expression level-dependent action of Galpha(q) and Galpha11 for Purkinje cell signaling and assign specific roles of these two G(q) isoforms for motor coordination.

Animals↗

PubMed2003

"In vivo" monitoring of neuronal network activity in zebrafish by two-photon Ca(2+) imaging.

The zebrafish larva is a powerful model for the analysis of behaviour and the underlying neuronal network activity during early stages of development. Here we employ a new approach of "in vivo" Ca(2+) imaging in this preparation. We demonstrate that bolus injection of membrane-permeable Ca(2+) indicator dyes into the spinal cord of zebrafish larvae results in rapid staining of essentially the entire spinal cord. Using two-photon imaging, we could monitor Ca(2+) signals simultaneously from a large population of spinal neurons with single-cell resolution. To test the method, Ca(2+) transients were produced by iontophoretic application of glutamate and, as observed for the first time in a living preparation, of GABA or glycine. Glycine-evoked Ca(2+) transients were blocked by the application of strychnine. Sensory stimuli that trigger escape reflexes in mobile zebrafish evoked Ca(2+) transients in distinct neurons of the spinal network. Moreover, long-term recordings revealed spontaneous Ca(2+) transients in individual spinal neurons. Frequently, this activity occurred synchronously among many neurons in the network. In conclusion, the new approach permits a reliable analysis with single-cell resolution of the functional organisation of developing neuronal networks.

Animals↗

PubMed2001

Impairment of mossy fiber long-term potentiation and associative learning in pituitary adenylate cyclase activating polypeptide type I receptor-deficient mice.

The pituitary adenylate cyclase activating polypeptide (PACAP) type I receptor (PAC1) is a G-protein-coupled receptor binding the strongly conserved neuropeptide PACAP with 1000-fold higher affinity than the related peptide vasoactive intestinal peptide. PAC1-mediated signaling has been implicated in neuronal differentiation and synaptic plasticity. To gain further insight into the biological significance of PAC1-mediated signaling in vivo, we generated two different mutant mouse strains, harboring either a complete or a forebrain-specific inactivation of PAC1. Mutants from both strains show a deficit in contextual fear conditioning, a hippocampus-dependent associative learning paradigm. In sharp contrast, amygdala-dependent cued fear conditioning remains intact. Interestingly, no deficits in other hippocampus-dependent tasks modeling declarative learning such as the Morris water maze or the social transmission of food preference are observed. At the cellular level, the deficit in hippocampus-dependent associative learning is accompanied by an impairment of mossy fiber long-term potentiation (LTP). Because the hippocampal expression of PAC1 is restricted to mossy fiber terminals, we conclude that presynaptic PAC1-mediated signaling at the mossy fiber synapse is involved in both LTP and hippocampus-dependent associative learning.

Animals↗

PubMed2000

NMDA receptor-mediated subthreshold Ca(2+) signals in spines of hippocampal neurons.

We have used rapid confocal microscopy to investigate the mechanism of Ca(2+) signals in individual dendritic spines of hippocampal CA1 pyramidal cells. The experiments focused on the signals that occur during single weak synaptic responses that were subthreshold for triggering postsynaptic action potentials. These Ca(2+) signals were not strongly affected by blocking the EPSPs with the AMPA receptor antagonist CNQX. The signals were also not strongly reduced by blocking T-type voltage-gated Ca(2+) channels (VGCCs) with Ni(2+) or by blocking a broad range of VGCCs with intracellular D890. The spine Ca(2+) signals were blocked by NMDA receptor channel (NMDAR) antagonist and had the voltage dependence characteristic of these channels. Neither ryanodine nor cyclopiazonic acid (CPA), substances known to deplete intracellular Ca(2+) stores, substantially reduced the amplitude of synaptically evoked Ca(2+) signals. CPA slowed the recovery phase of Ca(2+) signals in spines produced by synaptic stimulation or by backpropagating action potentials, suggesting a role of intracellular stores in Ca(2+) reuptake. Thus, we find that Ca(2+) release from intracellular stores is not required to produce spine Ca(2+) signals. We conclude that synaptic Ca(2+) signals in spines are primarily caused by Ca(2+) entry through NMDARs. Although these channels are largely blocked by Mg(2+) at voltages near the resting potential, they can nevertheless produce significant Ca(2+) elevation. The resulting Ca(2+) signals are an integral component of individual evoked or spontaneous synaptic events and may be important in the maintenance of synaptic function.

2-Amino-5-phosphonovalerate↗

PubMed1999

Two-photon Na+ imaging in spines and fine dendrites of central neurons.

Dendritic spines are assumed to be the smallest units of neuronal integration. Because of their miniature size, however, many of their functional properties are still unclear. New insights in spine physiology have been provided by two-photon laser-scanning microscopy which allows fluorescence imaging with high spatial resolution and minimal photodamage. For example, two-photon imaging has been employed successfully for the measurement of activity-induced calcium transients in individual spines. Here, we describe the first application of two-photon imaging to measure Na+ transients in spines and dendrites of CA1 pyramidal neurons in hippocampal slices. Whole-cell patch-clamped neurons were loaded with the Na(+)-indicator dye SBFI (sodium-binding benzofuran-isophthalate). In situ calibration of SBFI fluorescence with ionophores enabled the determination of the actual magnitude of the [Na+]i changes. We found that back-propagating action potentials (APs) evoked Na+ transients throughout the proximal part of the dendritic tree and adjacent spines. The action-potential-induced [Na+]i transients reached values of 4 mM for a train of 20 APs and monotonically decayed with a time constant of several seconds. These results represent the first demonstration of activity-induced Na+ accumulation in spines. Our results demonstrate that two-photon Na+ imaging represents a powerful tool for extending our knowledge on Na+ signaling in fine cellular subcompartments.

Action Potentials↗

PubMed1996

Long-term potentiation and functional synapse induction in developing hippocampus.

Long-term potentiation (LTP) is a cellular mechanism that potentially underlies learning and memory. To test the hypothesis that LTP is involved in activity-dependent synapse formation, we combined whole-cell recordings and confocal microscopy to investigate hippocampal glutamatergic synapses at their earliest stages of development. Here we report that, during the first postnatal week, the hippocampal glutamatergic network becomes gradually functional owing to the transformation of precursor, pure NMDA (N-methyl-D-aspartate)-receptor-based synaptic contacts into conducting AMPA (alpha-amino-3-hydroxy-5-methylisoxazole-4-proprionate)/NMDA-re cep tor-type synapses. This functional synapse induction is caused by an associative form of LTP, so it is input-specific and easily triggered experimentally by pairing presynaptic stimulation with postsynaptic depolarization. Our results challenge previous views that LTP occurs in the hippocampus only at later stages of development and that its induction requires dendritic spines. They also provide direct evidence that LTP is important for the activity-dependent formation of conducting glutamatergic synapses in the developing mammalian brain.

Animals↗

PubMed1995

Pre- and postsynaptic determinants of EPSC waveform at cerebellar climbing fiber and parallel fiber to Purkinje cell synapses.

Excitatory postsynaptic currents (EPSCs) at the parallel fiber and climbing fiber to Purkinje cell synapses were studied by whole-cell clamping Purkinje cells in cerebellar slices. Reducing glutamate release with adenosine or GABA decreased the amplitude of the EPSCs, with a larger suppression being produced at the parallel fiber synapse. Reducing glutamate release also speeded the decay of the EPSCs, and this effect was not a series resistance artefact since postsynaptic reduction of the current with CNQX did not speed the EPSC decay. Blocking glutamate uptake slowed the decay of the EPSCs. At the climbing fiber synapse, adenosine had little suppressive effect on the smaller EPSC evoked by the second of a pair of stimuli. Blocking desensitization of postsynaptic AMPA receptors prolonged the EPSC decay, preferentially increased the size of the second EPSC, and resulted in adenosine having a similar suppressive effect on the first and second EPSC. These data suggest that, at these synapses, the fall of glutamate concentration in the synaptic cleft overlaps with the decay of the EPSC, and that the EPSC size and duration are controlled by the amount of glutamate released, the rate of glutamate uptake, and desensitization.

Adenosine↗

PubMed1994

Arachidonic acid depresses non-NMDA receptor currents.

Arachidonic acid has been proposed as an intercellular messenger in the nervous system. It is released when glutamate acts on postsynaptic receptors, potentiates NMDA receptor currents and depresses glutamate uptake. Here we report the effects of arachidonic acid on non-NMDA receptor currents, studied by whole-cell clamping isolated neurons and neurons in tissue slices. In cultured cerebellar granule cells and in freshly isolated hippocampal pyramidal cells arachidonic acid decreased the current produced by iontophoresed AMPA. This depression was not due to increased desensitization of the AMPA receptor. In cerebellar slices, arachidonic acid depressed the non-NMDA component of the synaptic current at the mossy fibre to granule cell and the parallel fibre to Purkinje cell synapses. However, this depression was not always seen, possibly because the lipophilic arachidonic acid is absorbed by superficial cells in the slice and does not reach the synapse being studied. Depression of non-NMDA receptor currents by arachidonic acid may reflect the presence of an arachidonic acid binding site on the non-NMDA receptor, but non-NMDA receptor subunits show much less sequence homology with fatty acid binding proteins than does the NMDA receptor.

Amino Acid Sequence↗

PubMed1993

Glutamate induces long-term increase in the frequency of single N-methyl-D-aspartate channel openings in hippocampal CA1 neurons examined in situ.

Long-term potentiation is currently a leading candidate for a physiological memory mechanism in CNS. The interpretation of this phenomenon is contradictory in many respects. However, there is clear evidence that long-term potentiation is critically dependent on activation of N-methyl-D-aspartate receptors. Recently it has been shown that extracellularly applied glutamate also induces long-lasting changes in the properties of synaptic transmission in the hippocampus that can be attributed to long-term potentiation. The involvement of presynaptic mechanisms has been reported. Here we demonstrate a definite increase in both the open time and open-state probability of N-methyl-D-aspartate-operated channels, induced by prolonged application of glutamate to hippocampal slices.

Animals↗