title: Characterization of Ca2+ signals induced in hippocampal CA1 neurones by the synaptic activation of NMDA receptors.
abstract: 1. A combination of confocal microscopy, whole-cell patch-clamp recording, intracellular dialysis and pharmacological techniques have been employed to study Ca2+ signalling in CA1 pyramidal neurones, within rat hippocampal slices. 2. In the soma of CA1 neurones, depolarizing steps applied through the patch-pipette resulted in transient increases in the fluorescence emitted by the Ca2+ indicator fluo-3. The intensity of the fluorescence transients was proportional to the magnitude of the Ca2+ currents recorded through the pipette. Both the somatic fluorescence transients and the voltage-activated Ca2+ currents ran down in parallel over a period of between approximately 15-45 min. The fluorescence transients were considered, therefore, to be caused by increases in cytosolic free Ca2+. 3. Under current-clamp conditions, high-frequency (tetanic) stimulation (100 Hz, 1 s) of the Schaffer collateral-commissural pathway led to compound excitatory postsynaptic potentials (EPSPs) and somatic Ca2+ transients. The somatic Ca2+ transients were sensitive to the N-methyl-D-aspartate (NMDA) receptor antagonist D-2-amino-5-phosphonopentanoate (AP5; 100 microM). These transients, but not the EPSPs, disappeared with a time course similar to that of the run-down of voltage-gated Ca2+ currents. Tetanus-induced somatic Ca2+ transients could not be elicited under voltage-clamp conditions. 4. Fluorescence images were obtained from the dendrites of CA1 pyramidal neurones starting at least 30 min after obtaining whole-cell access to the neurone. Measurements were obtained only after voltage-gated Ca2+ channel activity had run down completely. 5. Tetanic stimulation of the Schaffer collateral-commissural pathway resulted in compound EPSPs and excitatory postsynaptic currents (EPSCs), under current- and voltage-clamp, respectively. In both cases, these were invariably associated with dendritic Ca2+ transients. In cells voltage-clamped at -35 mV, the fluorescent signal increased on average 2-fold during the tetanus and decayed to baseline values with a half-time (t1/2) of approximately 5 s. 6. The alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptor antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 10 microM) partially reduced the tetanus-induced EPSC without affecting the Ca2+ transients. In contrast, AP5, which also depressed the EPSC, substantially reduced or eliminated the Ca2+ transients. 7. In normal (i.e. 1 mM Mg(2+)-containing) medium, NMDA receptor-mediated synaptic currents displayed the typical region of negative slope conductance in the peak I-V relationship (between -90 and -35 mV). The dendritic tetanus-induced Ca2+ transients also displayed a similar anomalous voltage dependence, decreasing in size from -35 to -90 mV.(ABSTRACT TRUNCATED AT 400 WORDS)
authors.affiliation: Department of Pharmacology, School of Medical Sciences, University of Bristol.
authors.forename: S
authors.initials: S
authors.lastname: Alford
authors.forename: B G
authors.initials: BG
authors.lastname: Frenguelli
authors.forename: J G
authors.initials: JG
authors.lastname: Schofield
authors.forename: G L
authors.initials: GL
authors.lastname: Collingridge
mesh_terms: D000818:Animals; D002118:Calcium / Q000502:physiology*; D015220:Calcium Channels / Q000502:physiology; D003712:Dendrites / Q000502:physiology; D004558:Electric Stimulation; D005071:Evoked Potentials / Q000187:drug effects / Q000502:physiology; D006624:Hippocampus / Q000166:cytology / Q000502:physiology*; D066298:In Vitro Techniques; D015640:Ion Channel Gating / Q000502:physiology; D007834:Lasers; D008853:Microscopy; D009434:Neural Pathways / Q000502:physiology; D009474:Neurons / Q000502:physiology*; D017966:Pyramidal Cells / Q000378:metabolism; D051381:Rats; D017459:Receptors, Amino Acid / Q000378:metabolism; D016194:Receptors, N-Methyl-D-Aspartate / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology*; D013050:Spectrometry, Fluorescence; D013569:Synapses / Q000502:physiology*
chemical_list: D015220:Calcium Channels; D017459:Receptors, Amino Acid; D016194:Receptors, N-Methyl-D-Aspartate; D002118:Calcium