Characterization of Ca2+ signals induced in hippocampal CA1 neurones by the synaptic activation of NMDA receptors.

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)