Currents In Hippocampal Neurons Research Articles

The K+ channel opener diazoxide enhances glutamatergic currents and reduces GABAergic currents in hippocampal neurons

1. The effect of diazoxide, an opener of ATP-sensitive K+ channels (KATP channels) has been investigated in the rat hippocampal slices by the use of extracellular and intracellular recording techniques. 2. In control solution, diazoxide enhanced the CA1 and CA3 field excitatory postsynaptic potentials (EPSPs) and produced interictal activities in CA3. These effects were neither prevented by KATP blockers, including glibenclamide (3-30 microM) or tolbutamide (500 microM), nor mimicked by another KATP opener such as galanine (1 microM); thus these effects are probably not mediated by KATP channels. 3. Using intracellular recording, we then studied, in CA3 pyramidal neurons, the effect of diazoxide on the EPSPs and the fast and slow inhibitory postsynaptic potentials (IPSPs). 4. In presence of bicuculline (10 microM) and phaclofen (50 microM), to block, respectively, fast and slow IPSPs, diazoxide reversibly enhanced the EPSPs. 5. In presence of 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 10 microM), to block EPSPs, diazoxide reversibly decreased both fast and slow IPSPs. 6. These effects of diazoxide on the EPSPs and fast and slow IPSPs were associated neither with a change of the reversal potential of the EPSPs or the fast and slow IPSPs nor with a change of the input resistance and membrane potential. 7. Using single electrode voltage-clamp technique, we then tested the effects of diazoxide on the currents generated by applications of glutamate or gamma-aminobutyric acid (GABA) -A and -B analogues. 8. In presence of tetrodotoxin (TTX; 1 microM), diazoxide reversibly enhanced the peak currents evoked by alpha-amino-3-hydroxy-5-methyl-4- isoxazolepropionate (AMPA; 3-5 microM), quisqualate (5-10 microM) and N-methyl-D-aspartate (NMDA; 10 microM), but not those evoked by kainate (1-3 microM). 9. In presence of TTX (1 microM), diazoxide reversibly decreased the GABA- (1-5 mM), isoguvacine- (30-60 microM), and baclofen- (10-30 microM) mediated peak currents. 10. It is concluded that, in the hippocampus, diazoxide enhances the excitatory glutamatergic currents and reduces the GABAergic inhibition, thus generating paroxystic activities. We suggest that these effects are mediated by second messenger cascades.

Valproate and sodium currents in cultured hippocampal neurons.

Cultured rat hippocampal neurons with short processes were investigated using the whole cell voltage clamp under conditions appropriate for isolating Na+ currents. After incubation of the neuron culture for a period of 15-30 min in 1 mM sodium valproate, several parameters of the Na+ current were changed. The peak Na+ conductance gp, measured using hyperpolarizing prepulses, was reduced by valproate in a voltage-dependent manner. In the membrane voltage range from -30 to +20 mV, this reduction showed a linear dependence on voltage, increasing from about zero to approximately 30% of gp, the maximum peak Na+ conductance of the neuron. At the holding voltage of -70 mV, the inactivation parameter h infinity decreased from 0.88 in the control to 0.64 in the valproate solution. This reduction originated mainly from a 10 mV shift in the sigmoid relation between h infinity and membrane voltage along the voltage axis to hyperpolarizing potentials. The decay of the maximum peak Na+ current (inactivation) could be fitted by a biexponential function. Time constants of the fast and slow component at -20 mV decreased in valproate by about 50%. Valproate also retarded the recovery from inactivation, as determined at the holding voltage. The sigmoid recovery from inactivation could reasonably be described by an exponential function with time constant tau r and delay time delta t. Both tau r and delta t increased more than 200% in valproate. Our results indicate that valproate affected the Na+ current in hippocampal neurons in a way that contributed to a considerable depression of Na+ reactivation.(ABSTRACT TRUNCATED AT 250 WORDS)

Modulation of decay kinetics and frequency of GABA A receptor-mediated spontaneous inhibitory postsynaptic currents in hippocampal neurons

Inhibitory postsynaptic currents mediated by spontaneous activation of GABA A receptors were studied using whole-cell voltage-clamp recordings in granule cells of the adult rat (postnatal day 60+) dentate gyrus in 400-μm-thick coronal half-brain slices maintained at 34–35°C. The average amplitude of spontaneous inhibitory postsynaptic currents remained constant during a given recording period (i.e. no rundown was noted). The spontaneous currents had an average conductance between 200–400 pS, were mediated by Cl − flux through GABA A receptor/channels since they reversed at the Cl − equilibrium potential and were blocked by bicuculline or picrotoxin. Their mono-exponential decay time-constants (range: 4.2–7.2 ms) were prolonged by midazolam and pentobarbital in a dose-dependent manner. The effect of midazolam was reversed by the benzodiazepine receptor antagonist flumazenil (RO 15–1788) which, by itself, had no effect on the decay time-constant. The decay time-constant was also dependent on membrane voltage and on temperature. A 132-mV change in membrane potential produced an e-fold prolongation of the decay while the Q 10 (between 22–37°C) of the decay rate was 2.1. Within a given neuron, the frequency of spontaneous GABAergic events was remarkably constant over long time-periods, though the mean frequency among different cells showed large variability. Spontaneous miniature inhibitory postsynaptic currents also persisted under experimental conditions such as the presence of extracellular tetrodotoxin (1 μM), Cd 2+ (200 μM) or lowered extracellular Ca 2+/elevated Mg 2+, which effectively abolished all stimulus-evoked GABAergic neurotransmission. The frequency of tetrodotoxin-resistant miniature events was increased by elevating extracellular K + concentration and was diminished by the GABA B receptor agonist (−)baclofen only at a dose (50 μM) which was an order of magnitude larger than that required to depress stimulus-evoked responses. These findings are consistent with different mechanisms being responsible for the spontaneous and stimulus-evoked release of GABA from interneuron terminals and also identify pre- and postsynaptic modulatory factors of the endogenous, action-potential-independent, GABAergic neurotransmission as being important determinants of the excitability level of mammalian CNS neurons.

The actions of intermediate and long-chain n-alkanols on unitary NMDA currents in hippocampal neurons

The actions of the n-alkanols butanol, pentanol, and octanol on unitary currents passing through N-methyl-D-aspartate (NMDA) ion channels have been studied in cultured CA1 hippocampal neurons. The cell-attached patch clamp method, with L-homocysteic acid included in the patch pipette, was used to record single channel NMDA currents at the cell resting potential or for hyperpolarizing patch potentials. With the n-alkanols added to the bath solution, the mean open times for the NMDA channel were diminished and the channel conductance was unchanged. A decrease in mean open time to about 70% of control value was found with butanol (3 mM), pentanol (1 mM), and octanol (0.02 mM). In addition the n-alkanols had small effects to decrease the frequency of channel openings and to increase the amplitude of the unitary currents. The effects of the alcohols on intracellular calcium levels, during NMDA applications, were also measured using the fluorescent dye FURA II.

Initial Characterization of the Nicotinic Acetylcholine Receptors in Rat Hippocampal Neurons

The properties of the neuronal nicotinic acetylcholine receptor in primary cultures of hippocampal cells from fetal rats (17-18 days gestation) were studied using the whole-cell patch-clamp technique in Na(+)-external, Cs(+)-internal and nominally Mg(2+)-free solutions. The nicotinic agonists acetylcholine, (+)anatoxin-a, and (-) and (+)nicotine all evoked inward whole-cell currents in hippocampal neurons that were voltage clamped near their resting potentials. Sensitivity to (+)anatoxin-a was first detected at around day 6, and thereafter the magnitude of the response increased as a function of number of days in culture up to about 40 days. The whole-cell current waveforms consisted of more than one peak whose relative amplitude depended on the agonist concentration. These currents were reversibly blocked by micromolar concentrations of d-tubocurarine, mecamylamine, and dihydro-beta-erythroidine. At nanomolar concentrations, neuronal bungarotoxin, alpha-bungarotoxin and alpha-cobratoxin caused an irreversible blockade of the currents but they were unaffected by tetrodotoxin, atropine, DL-2-amino-5-phosphonovaleric acid, Mg2+, and 6,7-dinitroquinoxaline-2,3-dione. In addition, the currents were also blocked in a reversible manner by methyllycaconitine at picomolar concentration. The current-voltage plots elicited by both (+)anatoxin-a and acetylcholine revealed larger inward currents and smaller or no outward currents. The present results demonstrate the existence of an inwardly rectifying, snake neurotoxin-sensitive functional nicotinic acetylcholine receptor ion channel in rat hippocampal neurons.

Quisqualate receptor-mediated depression of calcium currents in hippocampal neurons

The modulation of Ca2+ currents by the excitatory neurotransmitter glutamate and its analogs was investigated in hippocampal neurons in culture. In the presence of glutamate receptor-gated ion channel antagonists, all of the analogs tested caused either a small reversible depression or had no effect on the Ca2+ current. However, in neurons dialyzed with GTP gamma S, quisqualate and glutamate but not NMDA, kainate, AMPA, or L-APB caused marked and irreversible depressions of the Ca2+ current. This inhibition was only observed if Ca2+ was present in either the internal or external medium. Intracellular H-7, staurosporine, IP3, cAMP, cGMP, or calmodulin inhibitors failed to prevent the quisqualate-induced Ca2+ current inhibition. These observations are consistent with an interaction between a G protein-coupled glutamate receptor and Ca2+ channels.

Nonstationary fluctuation analysis of postsynaptic currents in hippocampal neurons reveals single synaptic channel properties

Nonstationary fluctuation analysis of postsynaptic currents in hippocampal neurons reveals single synaptic channel properties

Nonstationary fluctuation analysis of postsynaptic currents in hippocampal neurons reveals single synaptic channel properties

Nonstationary fluctuation analysis of postsynaptic currents in hippocampal neurons reveals single synaptic channel properties

Mild acidosis protects hippocampal neurons from injury induced by oxygen and glucose deprivation

Hippocampal neurons are extremely sensitive to ischemic injury; two plausible mechanisms have been implicated in mediating such damage. The first involves overexposure of neurons to excitatoryN-methyl- d-aspartate (NMDA) receptor agonists, which mobilize damaging concentrations of intracellular calcium; the second involves the generation of damaging tissue acidosis. A recent report shows that exposure to pH 6.6 can block NMDA-induced calcium currents in hippocampal neurons. This suggests that moderate acidity might protect against NMDA-mediated neurotoxicity and ischemic injury in vivo. We have observed such projection in vitro using primary hippocampal cultures. At an extracellular pH of 7.4, 6 h of glucose-free anoxia caused delayed and profound damage to neurons which was partially attenuated by the NMDA receptor antagonist, 2-amino-5-phosphonovaleric acid (APV). Dropping the pH to 6.5 provided virtually complete protection against this insult. Thus, acidosis need not be viewed exclusively as a damaging component of ischemic insults.

Glutamine currents in hippocampal neurons are attributable to contaminating glutamate

Physiological responses of cultured rat hippocampal neurons to exogenously applied glutamine were studied using tight-seal, whole-cell voltage clamp techniques in nominal extracellular magnesium. Four mM glutamine induced peak inward currents which were4.6 ± 1.7% (± 1S.D.) of current responses to 100 μM glutamate. Ten μM 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), 1 mM magnesium, 20 μM MK-801, and 200 μM d-aminophosphonovalerate (APV) blocked these responses in a pattern similar to N-methyl- d-aspartate antagonism. Current-voltage plots produced by 4 mM glutamine and 10 μM glutamate were identical. Even in the absence of extracellular magnesium, 4 mM glutamine was not toxic to our cultured neurons. The data suggest that physiological responses to glutamine are attributable to low concentrations of glutamate in commercial preparations and that glutamine alone has insignificant physiological and toxic effects.

Temporal integration by a slowly inactivating K+ current in hippocampal neurons

Nature 336, 379–381 (1988). IN this letter, the last two sentences on page 379 as printed are misleading, and should read: Both the outward current and the underlying conductance were blocked by very low concentrations (30 jxM or so) of 4-aminopyridine (4-AP). In this property ID resembles some other fast-activating, slowly inactivating K+ currents, for example in axons8 and sensory ganglion cells9.

Temporal integration by a slowly inactivating K+ current in hippocampal neurons.

A central aspect of neuronal function is how each nerve cell translated synaptic input into a sequence of action potentials that carry information along the axon, coded as spike frequency. When transduction from a graded depolarizing input to spikes is studied by injecting a depolarizing current, there is often a remarkably long delay to the first action potential, both in mammalian and molluscan neurons. Here, I report that the delayed excitation in rat hippocampal neurons is due to a slowly inactivating potassium current, ID. ID co-exists with other voltage-gated K+ currents, including a fast A current and a slow delayed rectifier current. As ID activates in the subthreshold range, and takes tens of seconds to recover from inactivation, it enables the cell to integrate separate depolarizing inputs over long times. ID also makes the encoding properties of the cell exceedingly sensitive to the prevailing membrane potential.