Different responses of CA1 and CA3 regions to hypoxia in rat hippocampal slice

1. To study the effects of brief periods of hypoxia on cellular functions in the rat hippocampal slice, extracellular and intracellular recordings were made from pyramidal neurons, and interstitial potassium activity ([K+]o) was measured in the pyramidal cell layers. Slices were perfused in an interface chamber at 36-37 degrees C with medium containing 8.5 mM [K+]o. Hypoxia was induced by switching the overflow gas from O2-CO2 to N2-CO2. 2. Brief periods of hypoxia (5-60 s) produced electrographic seizures with typical tonic and clonic components in 53% of 293 slices that generated spontaneous interictal bursts. Hypoxia-induced seizures were usually initiated in and restricted to the Ca1 region; only 2.5% of these slices generated seizures in CA3. In contrast to the CA1 region, the CA3 region could undergo spreading depression during hypoxia. The probability of seizure generation in CA1 was increased with increasing duration of hypoxia and was greatly reduced by lowering the bath temperature a few degrees. 3. [K+]o gradually increased in the CA1 and CA3 cell layers during the 20 s leading up to an hypoxia-induced seizure. [K+]o rose to approximately 9.8 mM (from a base line of 8.5 mM) in CA1 just before a seizure and to 11.4 mM during the seizure. After hypoxia, [K+]o reached a higher level in CA1 than in CA3, regardless of whether 1 microM tetrodotoxin was present to eliminate differences in cell firing in the two regions. CA1 pyramidal cells and glia gradually depolarized by several millivolts during and after hypoxia; no initial hyperpolarizing phase was detected. 4. Burst input from CA3 was necessary for hypoxia-induced seizures. The frequency and intensity of spontaneous burst-firing in CA3 remained steady in the period leading up to a CA1 seizure episode. In contrast, the intensity of synaptically driven bursts in CA1 grew markedly just before seizure onset. N-methyl-D-aspartate (NMDA) receptors participated in the crescendo of increasingly synchronous activity in CA1, because the competitive NMDA receptor antagonist, D-2-amino-5-phosphonovaleric acid (D-APV, 30 microM), stereoselectively reduced seizure intensity. 5. Hypoxia-induced seizures were followed by a depressant phase, which was manifested most prominently by a prolonged (up to several minutes) reduction in the frequency and intensity of burst-firing in the CA3 region, hyperpolarization of CA1 neurons, and undershoot of [K+]o. In normal (3.5 mM) [K+]o, synaptically driven population spikes in CA1 were only reduced in amplitude by hypoxia; hypoxia did not induce seizures in 3.5 mM [K+]o.(ABSTRACT TRUNCATED AT 400 WORDS)     

Propofol-mediated impairment of CA1 long-term potentiation in mouse hippocampal slices

Propofol (2,6-diisopropylphenol) is a short-acting intravenous anesthetic. Propofol is known to impair maintenance of long-term potentiation (LTP) in synaptic responses from Schaffer collateral-commissural (SC) pathway to CA1 pyramidal cells in the hippocampus, but the threshold concentration of propofol needed to elicit this action is unknown. The actions of propofol in vivo (e.g., amnesia, sedation, hypnosis and immobility) depend on its concentration, and thus it is necessary to determine the concentration required to impair CA1 LTP in order to assess the impact of impairment in vivo. In the present study, we investigated the effects of various concentrations of propofol on synaptic plasticity, primarily by measuring LTP at SC pathway to CA1 pyramidal cell synapses in mouse hippocampal slices. Continuous application of 50 microM propofol from 20 min before tetanus stimulation suppressed potentiation of the synaptic responses by tetanus stimulation. The suppression was pronounced from 10 min post-tetanus and about 55% suppression of the potentiation was observed at 60 min after tetanus. Propofol at 5 or 20 microM did not have this effect. The presence of gamma-aminobutyric acid type A (GABA(A)) receptors antagonist, picrotoxin, abrogated the suppression of LTP by 50 microM propofol. Propofol 50 microM did not affect long-term depression (LTD). These results suggest that the suppression of hippocampal CA1 LTP via GABA(A) receptors requires a much higher propofol concentration compared with that needed to induce amnesia.     

Endogenous adenosine regulates the effects of low-frequency stimulation on the induction of long-term potentiation in CA1 neurons of guinea pig hippocampal slices

A train of low-frequency afferent stimuli (LFS, 1 Hz, 1000 pulses), given 60 min prior to a tetanus (100 Hz, 100 pulses), suppresses the induction of long-term potentiation (LTP) in which a short-term potentiation decreases gradually back to the pre-tetanic level within 40-50 min (LTP suppression). We investigated the effects of adenosine A1 or A2 receptor antagonists (8-cyclopentyltheophylline (8-CPT) and CP-66713, respectively) on LTP suppression in CA1 neurons of guinea pig hippocampal slices. When the LFS was delivered in the presence of 8-CPT (1 microM), LTP suppression was not significantly affected. However, when LFS was delivered in the presence of CP-66713 (10 microM), LTP suppression was inhibited, leading to successful LTP induction. These results indicate that endogenous adenosine, acting via A2 receptors, is involved in the mechanism of LTP suppression.     

The long-term suppressive effect of prior activation of synaptic inputs by low-frequency stimulation on induction of long-term potentiation in CA1 neurons of guinea pig hippocampal slices

We have investigated the effects of prior activation of afferent inputs by a train of low-frequency stimulation (LFS) on the induction of long term potentiation (LTP) induced by highfrequency stimulation (tetanus, 100 Hz, 100 pulses) in CA1 neurons of guinea pig hippocampal slices. The parameters of the LFS were altered systematically: the frequency (1 or 5 Hz); the number of pulses (80, 200 or 1000); and the time lag from the LFS to the tetanus (20, 60 or 100 min). Conditioning effects of the LFS on the induction of LTP were evaluated in terms of the slope of the field excitatory postsynaptic potential (S-EPSP) and the amplitude of the population spike (A-PS). LTP could reliably be induced by 100 Hz tetanic stimulation delivered to a naive slice. In contrast, the attempt to induce LTP 60 min after LFS of 200 or 1000 pulses at 1 Hz resulted only in short-term potentiation while the LFS itself produced no significant change in the responses. The suppressive effect on LTP was significantly reduced for 1 Hz LFS with a smaller number of pulses (80 pulses), or a shorter (20 min) or longer (100 min) time lag from the LFS to the tetanus, or with LFS at a higher frequency (5 Hz). When the LFS of 1000 pulses at 1 Hz was delivered in the presence of the n-methyl-d-aspartate (NMDA) receptor antagonist AP5 (d,l-4-amino-5-phosphonovalerate, 50 M), which was washed out after the end of the LFS, the tetanus given 60 min after application of the LFS produced stable LTP, indicating the involvement of NMDA receptor/channels in the mechanisms of this particular form of synaptic plasticity-long-term suppression of LTP.     

K-dependent inhibition in the dentate-CA3 network of guinea pig hippocampal slices.

1. The occurrence of potassium-dependent inhibitory postsynaptic potentials (K-IPSPs) in relation to burst discharges induced by 4-aminopyridine (4-AP; 30 microM) was studied in CA3, granule and hilar neurons in guinea pig hippocampal slices with the use of paired extra- and/or intracellular recording. 2. Slow small (2-5 mV) and large (up to 30 mV) K-IPSPs were observed in CA3, granule and in some hilar neurons during 4-AP applications in the presence of blockers for fast synaptic transmission, picrotoxin (50 microM), and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 5-10 microM). Amplitudes of K-IPSPs were linearly related to voltage, and they reversed in sign close to -100 mV, as expected for synaptic potentials generated by an increase in K-conductance. 3. In CA3 neurons, 4-AP applied in the presence of picrotoxin elicited burst discharges and K-IPSPs. CNQX blocked the burst discharge activity and increased the amplitude of K-IPSPs. 4. In granule cells, 4-AP applied in the presence of picrotoxin elicited K-IPSPs and only inconsistently small excitatory postsynaptic potentials (EPSPs). The EPSPs were blocked by CNQX, but CNQX application did not affect the K-IPSPs. However, in granule cells it could be observed that blockade of Cl-inhibition by picrotoxin in the presence of CNQX increased the amplitude of K-IPSPs. 5. In hilar neurons, 4-AP applied in the presence of picrotoxin elicited mainly burst discharges. CNQX blocked the burst discharges only in a few cells. In most hilar neurons K-IPSPs were observed at the beginning of the 4-AP effect, but subsequently K-IPSPs were replaced by burst discharges. 6. To determine the type of cells that burst in picrotoxin and 4-AP, neurons were stained intracellularly with horseradish peroxidase. Neurons stained in the granule cell layer did not burst and were morphologically identified as granule cells. Neurons stained in the hilar region burst and were nonpyramidal, nongranule cells. Bursting cells stained in the CA3 area were all pyramidal cells. 7. The hilar neurons varied considerably in size and dendritic organization. They could be classified as aspiny and spiny cells, the latter including mossy cells. 8. We conclude that K-dependent inhibition may explain the long-lasting IPSPs observed in in vivo recordings from hippocampal cells. In a hippocampal lamella, burst discharge activity of hilar neurons including presumed excitatory mossy cells is associated with inhibition of granule cells.(ABSTRACT TRUNCATED AT 400 WORDS)     

PO2-profiles in hippocampal slices of the guinea pig

Summary For a detailed analysis of the oxygen supply of hippocampal slices, tissue PO₂ (P_{t,o 2}) was recorded polarographically in the neural layers of thick and thin slice preparations from the guinea pig. The experiments showed that the P_t,o₂-gradients were extremely steep in the outer zones of vital slices. In an air equilibrated salt solution the surface PO₂ was reduced to less than 50% within ca. 25 m. Minimum values were measured at a depth of ca. 150 m. A rise of temperature lowered the oxygen supply in the deeper layers of the excised tissue. An elevation of the surface PO₂ hardly improved P_{t,o 2} in the deep structures, because the O₂-consumption of the hippocampal slices increased with rising PO₂.Supported by DFG (Sp 108/6)Dedicated to Professor Dr. H. Caspers on the occasion of his 60th birthday     

Histamine decreases calcium-mediated potassium current in guinea pig hippocampal CA1 pyramidal cells

The action of histamine on CA1 pyramidal cells was studied in a hippocampal slice preparation. In the presence of tetrodotoxin (TTX) and tetraethylammonium (TEA), histamine had little effect on the calcium spikes. Using the single-electrode voltage-clamp technique, the actions of histamine on membrane currents were tested. In TTX, histamine (1 microM) decreased outward current only at potentials more depolarized than approximately -50 mV, where calcium-mediated potassium current is predominant. In the presence of manganese, histamine was without effect. Histamine (10 microM) did not affect the transient outward potassium current (A-current), the inward M-current resulting from small hyperpolarizing steps, or the inward Q-current elicited by larger hyperpolarizing steps. Blocking potassium currents with TEA or replacing calcium with barium revealed a slow inward current normally carried by calcium. With TTX present to block sodium currents, histamine (10 microM) did not reduce the inward current. The outward current reduced by a maximally effective concentration of histamine (10 microM) can be further decreased by manganese. The results support the conclusion that histamine selectively decreases the calcium-mediated potassium conductance in CA1 pyramidal cells of hippocampus. The possibility is raised that there is a component of calcium-mediated potassium current that is insensitive to histamine.     

Requirement of extracellular Ca2+ after tetanus for induction of long-term potentiation in guinea pig hippocampal slices

Long-term potentiation (LTP) is the persistent facilitation of synaptic responses induced by a brief tetanic stimulation (tetanus). Replacement of a standard solution containing 2.0 mM Ca2+ with one containing 1.0 mM Ca2+ for 5 min, either immediately after or during tetanus, significantly inhibited the induction of LTP. Furthermore, similar results were obtained when Verapamil or Loperamide, presumable Ca2+ antagonists, was applied immediately after tetanus. From these results, it was concluded that the induction of LTP requires extracellular Ca2+ after as well as during tetanus.     

Actions of pentylenetetrazol (PTZ) on CA3 neurons in hippocampal slices of guinea pigs

Summary The actions of the convulsant drug pentylenetetrazol (PTZ) were studied on CA3 neurons of hippocampal slices (300–400 m thick). In 9 out of 109 neurons, epileptic reactions were elicited during a single application of PTZ. After repeated applications of PTZ, 53 neurons showed periodic paroxysmal activity. They developed according to the following sequence: (i) Paroxysmal hyperpolarizations, (ii) burst activity, and (iii) typical paroxysmal depolarization shifts (PDS). The rate of occurrence was about 8/min. Paroxysmal hyper- and depolarizations appeared synchronously in pairs of neurons. The developmental sequence occurred in reverse during washing. After the onset of paroxysmal activity, bursts and PDS persisted if PTZ concentration in tissue ranged between about 2 and 10 mmol/l. When this range was exceeded in either direction, epileptic activity was abolished at a calcium concentration of 2.75 mmol/l. Decreasing and increasing the calcium concentration shifted the epileptogenic concentration range to lower and higher levels, respectively. It is concluded that repetition of PTZ application alters membrane properties of neurons and thus leads to paroxysmal events triggered by synaptic processes.Supported by Deutsche Forschungsgemeinschaft Sp 108/3 and Bi 278/1-3     

Glycine antagonists block the induction of long-term potentiation in CA1 of rat hippocampal slices

The effects of the strychnine-insensitive glycine receptor antagonists, cycloleucine and 7-chlorokynurenic acid, on the induction of long-term potentiation (LTP) in CA1 of rat hippocampal slices were examined. A 5 min administration of cycloleucine (20-100 microM) or 7-chlorokynurenic acid (1-5 microM) during the delivery of high-frequency stimulation blocked the induction of LTP without affecting baseline synaptic transmission. Coapplication of 100 microM glycine with cycloleucine or 7-chlorokynurenic acid masked the inhibitory effect on the induction of LTP, supporting the hypothesis that these compounds act as glycine antagonists. These results indicate that glycine is a necessary factor for the induction of LTP in CA1 of the rat hippocampus.     

Post-inhibitory excitation of adenosine on neurotransmission in guinea pig hippocampal slices.

The postsynaptic field potential (population spike potential; PS) was recorded from the granule cell layer of guinea pig hippocampal slices. Adenosine at low concentrations ranging from 10 nM to 1 microM enhanced the amplitude of PS, whereas at concentrations over 10 microM it inhibited the neurotransmission. There appeared to be a rebound phenomenon after the removal of adenosine at inhibitory concentrations and the amplitude of the PS overshot the initial amplitude (we called this post-inhibitory excitation; PIE). Neither depressants such as gamma-aminobutyric acid (GABA; 1 mM) nor sodium pentobarbital (100 microM) by itself induced PIE. After application of GABA or sodium pentobarbital together with adenosine (0.1 microM), however, removal of all agents could induce the PIE. PIE as well as the excitatory effect of adenosine at low concentrations was counteracted by application of H-7 (100 microM), melittin or polymyxin B, potent protein kinase C (PKC) inhibitors, suggesting that the excitatory effect of adenosine is mediated by a metabolic process involving PKC. These results indicate that PIE induced by adenosine at high concentrations is due to a mechanism similar to the excitatory effect induced by adenosine at low concentrations, and that during application of adenosine at high concentrations the excitation is masked by its potent inhibitory effect.     

Statistical analysis of long-term potentiation of large excitatory postsynaptic potentials recorded in guinea pig hippocampal slices: binomial model

Summary Excitatory postsynaptic potentials (EPSPs) were recorded in guinea pig hippocampal slices (area CA1) from 15 neurones after stimulation of stratum radiatum (str. rad.) and stratum oriens. EPSP amplitudes increased in 8 neurones (10 post-tetanic regions) recorded 15 to 45 min after tetanic stimulation of str. rad. The increase was considered to represent long-term potentiation (LTP). Quantal analysis was performed by two methods assuming binomial statistics: the histogram method using deconvolution of noise and the variance method. According to both methods, LTP was associated with an increase in mean quantal content (m) which correlated with LTP magnitude. A statistically significant increase in quantal size (v) was found only by the histogram method and the increase was not correlated with LTP magnitude. A separate analysis of EPSPs with small LTP magnitude demonstrated that with the histogram method only v was increased but not m. A smaller increase in m for the pooled data of both methods did not correlate with LTP magnitude for this EPSP subset. The increase in m for the whole EPSP set corresponds to previous results on the quantal analysis of LTP in in vivo preparations and favours a presynaptic location of major mechanisms underlying LTP maintenance. The increase in v indicates the existence of another mechanism responsible for the maintenance of a small part of LTP. This mechanism might involve either pre- or postsynaptic changes or both.     

Properties of the visually driven neurons of cat's hippocampal regions CA 1 and CA 3

Neurons in areas CA 1 and 3 of cat's dorsal hippocampus were studied. Fifteen percent of the investigated cells were influenced by visual stimuli. Eighty five such neurons were investigated. The organization of their receptive fields was tested with stationary and moving visual stimuli. Twenty eight percent of neurons had small receptive fields (10-20 deg square). Forty one neurons responded to stationary flashing spots. They were ON-OFF, ON and OFF types with phasic (66%) and tonic (34%) characteristics. Seventy five responded to dark and bright stimuli moving across their receptive fields. Twenty five neurons were direction-sensitive and 21 responded better to the dark moving stimuli than to the bright ones. No significant differences in the response properties of neurons in the CA 1 and CA 3 fields were observed.     

Different effects of dynorphin A on in vitro guinea pig hippocampal CA3 pyramidal cells with various degrees of paired-pulse facilitation

We investigated the effects of dynorphin A (Dyn A), a heptadecapeptide, on the population spikes of the guinea pig hippocampal CA3 pyramidal neurons, in vitro, using paired-pulse stimulation of the mossy fibers. Dyn A produced facilitatory and inhibitory effects on the population spikes in the preparations with lower and higher degrees of paired-pulse facilitation, respectively. Morphine and D-Ala2, D-Leu5-en-kephalin, mu- and delta-agonist, respectively, predominantly potentiated the population spikes, while kappa-agonists such as U-50, 488H and bremazocine mainly caused an inhibition. These results suggest that Dyn A has two separate (excitatory and inhibitory) effects on the guinea pig hippocampal CA3 neurons through mu-(delta) and kappa-opioid receptors, respectively.     

Desynchronization of carbachol-induced theta-like activities by alpha-adrenergic agents in guinea pig hippocampal slices

The effect of alpha-adrenergic agents on theta-like activity (TLA) in guinea-pig hippocampal slices was studied. TLA was induced by a cholinergic agent, carbachol. TLA had a frequency of 4.66+/-0.08 Hz (mean+/-S.E.M.) and an amplitude of 96.3+/-8.3 microV in the dentate gyrus (DG). The alpha-adrenergic agents epinephrine and clonidine increased the frequency and decreased the amplitude of TLA in a concentration-dependent manner. The agents also increased antidromic and orthodromic population spike (PS) amplitudes of the granule cells, but did not have any effect on population EPSP. Another alpha-adrenergic agent, guanabentz, had the same effect as clonidine. The adenylate cyclase inhibitor SQ22536 suppressed the increase in PS amplitude by clonidine. The results suggest that these alpha2-adrenergic agents facilitate the activity of granule cells through the activation of the alpha2-adrenoreceptor and cAMP pathway, and the facilitation causes the desynchronization of TLA.     

Effects of methohexital on bioelectrical reactions in guinea pig hippocampal slices during hypoxia

The barbiturate methohexital (42 and 140 micromol/l) was tested for an acute effect on anoxic depolarization (AD) and evoked potentials (EP) in hippocampal slices of guinea pigs exposed to repeated hypoxic conditions (n = 78). The dosages of methohexital resemble the range of plasma levels measured in patients with an intraoperative burst suppression electroencephalogram. Direct current potential and EP were recorded in the CA1 region. Hypoxia was terminated either when AD had reached its peak, or 2 min after maximum AD. Excluding the actions of methohexital on cerebral blood flow, reperfusion phase and the delayed mechanisms of cellular protection, a dose-dependent direct effect on EP even after repeated hypoxic conditions could be observed.     

Effect of the mono- and tetra-sialogangliosides, GM1 and GQ1b, on long-term potentiation in the CA1 hippocampal neurons of the guinea pig

 Effects of the mono- and tetra-sialogangliosides, GM1 and GQ1b, on long-term potentiation (LTP) were investigated in the CA1 neurons of guinea-pig hippocampal slices. The magnitude of LTP induced by a strong tetanus (100 Hz, 100 pulses) was not significantly affected by application of either ganglioside. In contrast, when LTP was induced by a weak tetanus (100 Hz, 4 pulses), a significantly greater LTP was induced in the presence of either ganglioside. Similarly, when slices were incubated in low-Ca²⁺ (1.0–1.1 mM) medium for more than 2 h, the LTP was usually small or absent, but showed a significant increase in amplitude of population spike (A-PS) when the slices were incubated with either GM1 or GQ1b (4–5 μg/ml). In addition, the application of GQ1b (4 μg/ml) reversed the blocking effect of an NMDA-receptor antagonist, APP-5 (10 μM), on the induction of LTP and resulted in forming LTP. Based on these findings, we conclude that GM1 and GQ1b exert positive modulatory effects on the induction of LTP in hippocampal CA1 neurons and suggest that GM1 and GQ1b may participate in the induction of LTP as donors of Ca²⁺ ions. Received: 21 April 1998 / Accepted: 5 May 1998     

Intracellular correlates of spatial memory acquisition in hippocampal slices: long-term disinhibition of CA1 pyramidal cells

Despite many advances in our understanding of synaptic models of memory such as long-term potentiation and depression, cellular mechanisms that correlate with and may underlie behavioral learning and memory have not yet been conclusively determined. We used multiple intracellular recordings to study learning-specific modifications of intrinsic membrane and synaptic responses of the CA1 pyramidal cells (PCs) in slices of the rat dorsal hippocampus prepared at different stages of the Morris water maze (WM) task acquisition. Schaffer collateral stimulation evoked complex postsynaptic potentials (PSP) consisting of the excitatory and inhibitory postsynaptic potentials (EPSP and IPSP, respectively). After rats had learned the WM task, our major learning-specific findings included reduction of the mean peak amplitude of the IPSPs, delays in the mean peak latencies of the EPSPs and IPSPs, and correlation of the depolarizing-shifted IPSP reversal potentials and reduced IPSP-evoked membrane conductance. In addition, detailed isochronal analyses revealed that amplitudes of both early and late IPSP phases were reduced in a subset of the CA1 PCs after WM training was completed. These reduced IPSPs were significantly correlated with decreased IPSP conductance and with depolarizing-shifted IPSP reversal potentials. Input-output relations and initial rising slopes of the EPSP phase did not indicate learning-related facilitation as compared with the swim and na?ve controls. Another subset of WM-trained CA1 PCs had enhanced amplitudes of action potentials but no learning-specific synaptic changes. There were no WM training-specific modifications of other intrinsic membrane properties. These data suggest that long-term disinhibition in a subset of CA1 PCs may facilitate cell discharges that represent and record the spatial location of a hidden platform in a Morris WM.     

低氧大鼠海马脑片CA1区长时程增强的变化及ACTH的作用

目的: 研究ACTH对低氧海马脑片CA1区长时程增强的影响。方法: 细胞外记录海马脑片CA1区诱发的群峰电位(PS)和强直刺激诱发的长时程增强(LTP)。结果: 低氧(5% O₂+90% N₂+5% CO₂或11.2% O₂+83.8% N₂+5% CO₂)灌流海马脑片后, LTP的诱出时间显著延长、诱出率明显降低, PS迅速减少并逐渐消失。预先灌流ACTH可改善上述效应。结论: 低氧可损害海马脑片CA1区LTP的诱发过程, ACTH可使之改善。     

Effect of scopolamine and HP 029, a cholinesterase inhibitor, on long-term potentiation in hippocampal slices of the guinea pig

The effect of scopolamine and a cholinesterase inhibitor on long-term potentiation (LTP) of population spikes was studied in a guinea pig hippocampal slice preparation. After brief application of each drug (10 min), LTP in CA1 and CA3 was induced by tetanus stimulation delivered to commissural/associational fibers and mossy fibers, respectively. Scopolamine at concentration of 10 microM had no effect on LTP in CA1 but significantly suppressed LTP in CA3. The cholinesterase inhibitor, 9-amino-1,2,3,4-tetrahydroacridine-1-ol maleate (HP 029) at concentration of 10 microM significantly enhanced LTP both in CA1 and CA3. These results suggest that the cholinergic system is involved in producing LTP in CA3. Another mechanism of the effect of HP 029 on LTP in CA1 is discussed.