Ionic basis of the caesium-induced depolarisation in rat supraoptic nucleus neurones.

1. The effects of external Cs(+) on magnocellular neurosecretory cells were studied during intracellular recordings from 93 supraoptic nucleus neurones in superfused explants of rat hypothalamus. 2. Bath application of 3-5 mM Cs(+) provoked reversible membrane depolarisation and increased firing rate in all of the neurones tested. Voltage-current analysis revealed an increase in membrane resistance between -120 and -55 mV. The increase in resistance was greater below -85 mV than at more positive potentials. 3. Voltage-clamp analysis showed that external Cs(+) blocked the hyperpolarisation-activated inward current, I(H). Under current clamp, application of ZD 7288, a selective blocker of I(H), caused an increase in membrane resistance at voltages < or = -65 mV. Voltage-current analysis further revealed that blockade of I(H) caused hyperpolarisation when the initial voltage was < -60 mV but had no effect at more positive values. 4. Current- and voltage-clamp analysis of the effects of Cs(+) in the presence of ZD 7288, or ZD 7288 and tetraethyl ammonium (TEA), revealed an increase in membrane resistance throughout the range of voltages tested (-120 to -45 mV). The current blocked by Cs(+) in the absence of I(H) was essentially voltage independent and reversed at -100 mV. The reversal potential shifted by +22.7 mV when external [K(+)] was increased from 3 to 9 mM. We conclude that, in addition to blocking I(H), external Cs(+) blocks a leakage K(+) current that contributes significantly to the resting potential of rat magnocellular neurosecretory cells.

Excitatory role of the hyperpolarization-activated inward current in phasic and tonic firing of rat supraoptic neurons.

The properties and functional roles of the hyperpolarization-activated inward current (I(H)) in magnocellular neurosecretory cells (MNCs) were investigated during sharp microelectrode recordings from supraoptic neurons in superfused explants of rat hypothalamus. Under current clamp, voltage responses to hyperpolarizing current pulses featured depolarizing sags that were abolished by the I(H) blocker ZD 7288. Under voltage clamp, subtraction of current responses to hyperpolarizing steps recorded in the absence and presence of ZD 7288 was used to investigate the properties of I(H). Current-voltage analysis revealed that steady-state I(H) amplitude increases with hyperpolarization, with half-maximal activation of the underlying conductance occurring at -78 mV. The time course of activation of I(H) during hyperpolarizing steps was monoexponential with time constants (100-800 msec) decreasing with hyperpolarization. The effects of ZD 7288 on I(H) were slow (tau, approximately 15 min), irreversible, and half-maximal at 1.8 micrometer. When tested on continuously active MNCs, application of 30-60 micrometer ZD 7288 caused a significant reduction in firing rate. In phasically active MNCs, the drug decreased burst duration and intraburst firing frequency and caused an increase in the duration of interburst intervals. These effects were accompanied with a small hyperpolarization of the membrane potential. In contrast, ZD 7288 had no effect on spike duration, on the amplitude of calcium-dependent afterpotentials, or on the frequencies and amplitudes of spontaneous synaptic potentials. These results confirm the presence of I(H) in MNCs of the rat supraoptic nucleus and suggest that the presence of this conductance provides an excitatory drive that contributes to phasic and tonic firing.

Kappa-opioid receptor activation inhibits post-spike depolarizing after-potentials in rat supraoptic nucleus neurones in vitro.

Endogenous agonists acting at kappa-opioid receptors modulate the discharge activity of hypothalamic supraoptic nucleus vasopressin cells in vivo. Phasic activity in vasopressin cells is known to depend critically on intrinsic mechanisms involving post-spike depolarizing after-potentials and we hypothesized that inhibition of phasic bursting by an endogenous kappa-agonist may result from reducing the magnitude of depolarizing after-potentials. To investigate this possibility, intracellular sharp electrode recordings were obtained from supraoptic nucleus cells impaled in superfused explants of rat hypothalamus. Bath application of the selective kappa-agonist, U50,488H (0.1-1 microM), decreased the spontaneous firing rate of magnocellular neurosecretory cells (by 94. 0+/-4.5% at 1 microM, mean+/-SEM; P = 0.02, n = 4). U50,488H did not alter membrane potential (0.9+/-0.8 mV hyperpolarization at 1 microM, P = 0.17, n = 8) or input resistance (11.0+/-4.5% increase at 1 microM, P = 0.09, n = 5). U50,488H (0.1 and 1 microM, both n = 5) reduced depolarizing after-potential amplitude (by 29.9+/-9.3 and 78.0+/-10. 6%, respectively, P<0.001) in eight cells in which the baseline membrane potential was kept constant by dc-current injection and in which a depolarizing after-potential was evoked every 25-40 s by a brief (40-80 ms) train of 3-6 action potentials (the number of spikes in the trains was kept constant for each cell). Thus, kappa-opioid receptor activation reduces depolarizing after-potential amplitude in supraoptic nucleus cells and this may underlie the reduction in burst duration of vasopressin cells caused by an endogenous kappa-agonist in vivo.

Caesium blocks depolarizing after-potentials and phasic firing in rat supraoptic neurones.

1. The effects of Cs+ on the action potential, post-train after-hyperpolarization (AHP), Ca2+-dependent post-spike depolarizing after-potential (DAP) and phasic firing were examined during intracellular recordings from magnocellular neurosecretory cells (MNCs) in superfused rat hypothalamic explants. 2. Extracellular Cs+ reversibly inhibited (IC50, approximately 1 mM) DAPs, and associated after-discharges, that followed brief spike trains in each of sixteen cells tested. Although bath application of Cs+ also provoked a small reversible depolarization, inhibition of the DAP was retained when membrane voltage was kept constant by current injection. 3. Application of Cs+ had no significant effects on spike duration (n = 8), frequency-dependent spike broadening (n = 8), spike hyperpolarizing after-potentials (n = 14), or the amplitude of the isolated AHP (n = 7). Caesium-evoked inhibition of the DAP, therefore, does not result from diminished spike-evoked Ca2+ influx, and may reflect direct blockade of the conductance underlying the DAP. 4. Inhibition of the DAP was associated with an enhancement of the amplitude and duration of the AHP, indicating that the currents underlying the AHP and the DAP overlap in time following a train of action potentials, and that the relative magnitude of these currents is an important factor in determining the shape and time course of post-train after-potentials. 5. Bath application of Cs+ reversibly abolished phasic firing in each of seven cells tested. This effect was reversible and persisted at all subthreshold voltages tested. These results indicate that the current underlying the DAP is necessary for the genesis of plateau potentials and phasic firing in MNCs.

Changes of spontaneous miniature excitatory postsynaptic currents in rat hippocampal pyramidal cells induced by aniracetam.

Spontaneous miniature excitatory postsynaptic currents (mEPSCs) in rat hippocampal pyramidal neurones in slices (CA1 region) were recorded at 35-37 degrees C using the whole-cell patch-clamp technique before and after addition of aniracetam (1 mM) to determine how a partial blockade of desensitization alters the relationship between the amplitude (A) and kinetics of mEPSCs, and to evaluate the factors that determine their variability. The rise time (taur) and the time constant of decay of mEPSCs (taud) are essentially amplitude independent in control conditions, but become clearly amplitude dependent in the presence of aniracetam. The slopes of the best fitting lines to taud:A and taur:A data pairs were (+/- SD; ms/pA; n = 5): (1) (control) 0.07 +/- 0.02 and 0.008 +/- 0.003; (2) (aniracetam) 0.40 +/- 0.19 and 0.22 +/- 0.22. The amplitude-dependent prolongation of taud is explained by the concentration dependence of two related processes, the buffering of glutamate molecules by AMPA receptor channels, and the occupancy of the double-bound activatable states. A slower deactivation makes an amplitude-independent contribution. Desensitization reduces the amplitude dependence of taud by minimizing repeated openings of alpha-amino-3-hydroxy-methyl-isoxazole (AMPA) receptor channels. A greater amplitude dependence of taur probably involves both pre- and postsynaptic factors. The variability of A and taud values did not change significantly, but the factors underlying the variability of taud values were much affected. The greater amplitude dependence and the greater scatter about the best fitting lines to taud:A data pairs are approximately balanced by the greater mean values. The greater scatter of taud about the best fitting lines probably occurs because the saturation of AMPA receptors is not the same at different synapses with different numbers of AMPA receptors.