The electrophysiology of prefrontal serotonin systems: therapeutic implications for mood and psychosis

A newly described synaptic action of serotonin (5-HT) in the cerebral cortex is reviewed, and implications for mood and psychosis are discussed. Recordings in brain slices show that 5-HT induces a rapid increase in excitatory postsynaptic potentials/currents (EPSPs/EPSCs) in virtually all layer V pyramidal cells of neocortex. This effect is mediated by the 5-HT2A receptor, which has been linked to the action of hallucinogenic and atypical antipsychotic drugs. The increase in EPSCs is seen most prominently in medial prefrontal cortex and other frontal regions where 5-HT2A receptors are enriched. The induction of EPSCs by 5-HT appears to occur through a novel mechanism that does not depend on the activation of afferent impulse flow. Instead, 5-HT appears to act presynaptically, directly or indirectly, to induce a focal release of glutamate from a subpopulation of glutamatergic terminals impinging upon the apical (but not basilar) dendrites of layer V pyramidal cells; a working hypothesis of the transduction pathway (involving asynchronous transmitter release) for this process is presented. Consistent with a focal action upon glutamatergic nerve terminals, the 5-HT-induced EPSPs can be suppressed by presynaptic inhibitory modulators such as mu-opiate or group II/III metabotropic agonists. We suggest that the suppression of 5-HT-induced EPSCs by 5-HT2A antagonists and mu-opiate agonists may underlie certain shared clinical effects of 5-HT2A antagonists and mu-opiate agonists. We suggest further that since presynaptic group II/III metabotropic glutamate agonists suppress 5-HT-induced EPSCs, metabotropic glutamate agonists may also possess antidepressant and/or antipsychotic properties.     

Mechanisms underlying the enhancement of excitatory synaptic transmission in basolateral amygdala neurons of the kindling rat

To elucidate the mechanism underlying epileptiform discharges in kindled rats, synaptic responses in kindled basolateral amygdala neurons in vitro were compared with those from control rats by using intracellular and whole cell patch-clamp recordings. In kindled neurons, electrical stimulation of the stria terminalis induced epileptiform discharges. The resting potential, apparent input resistance, current-voltage relationship of the membrane, and the threshold, amplitude, and duration of action potentials in kindled neurons were not different from those in control neurons. The electrical stimulation of stria terminalis elicited excitatory postsynaptic potentials (EPSPs) and DL-2-amino-5-phosphonopentanoic acid (AP5)-sensitive and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX)-sensitive excitatory postsynaptic currents (EPSCs). The amplitude of evoked EPSPs and of evoked AP5-sensitive and CNQX-sensitive EPSCs were enhanced markedly, whereas fast and slow inhibitory postsynaptic potentials (IPSPs) induced by electrical stimulation of lateral amygdaloid nucleus were not significantly different. The rise time and the decay time constant of the evoked CNQX-sensitive EPSCs were shortened, whereas the rise time of the evoked AP5-sensitive EPSCs was shortened, but the decay time constants were not significantly different. In both tetrodotoxin (TTX)-containing medium and low Ca2+ and TTX-containing medium, the frequency and amplitude of spontaneous EPSCs were increased in kindled neurons. These increases are presumably due to nearly synchronous multiquantal events resulted from the increased probability of Glu release at the nerve terminals. The rise time of evoked CNQX- and AP5-sensitive EPSCs and the decay time constant of evoked CNQX-sensitive EPSCs were shortened, suggesting that excitatory synapses at the proximal dendrite and/or the soma in kindled neurons may contribute more effectively to generate evoked EPSCs than those at distal dendrites. In conclusion, the increases in the amplitudes of spontaneous and evoked EPSCs and in the frequency of spontaneous EPSCs may contribute to the epileptiform discharges in kindled neurons.     

Evidence from simultaneous intracellular recordings in rat hippocampal slices that area CA3 pyramidal cells innervate dentate hilar mossy cells

1. Simultaneous intracellular recordings of area CA3 pyramidal cells and dentate hilar "mossy" cells were made in rat hippocampal slices to test the hypothesis that area CA3 pyramidal cells excite mossy cells monosynaptically. Mossy cells and pyramidal cells were differentiated by location and electrophysiological characteristics. When cells were impaled near the border of area CA3 and the hilus, their identity was confirmed morphologically after injection of the marker Neurobiotin. 2. Evidence for monosynaptic excitation of a mossy cell by a pyramidal cell was obtained in 7 of 481 (1.4%) paired recordings. In these cases, a pyramidal cell action potential was followed immediately by a 0.40 to 6.75 (mean, 2.26) mV depolarization in the simultaneously recorded mossy cell (mossy cell membrane potentials, -60 to -70 mV). Given that pyramidal cells used an excitatory amino acid as a neurotransmitter (Cotman and Nadler 1987; Ottersen and Storm-Mathisen 1987) and recordings were made in the presence of the GABAA receptor antagonist bicuculline (25 microM), it is likely that the depolarizations were unitary excitatory postsynaptic potentials (EPSPs). 3. Unitary EPSPs of mossy cells were prone to apparent "failure." The probability of failure was extremely high (up to 0.72; mean = 0.48) if the effects of all presynaptic action potentials were examined, including action potentials triggered inadvertently during other spontaneous EPSPs of the mossy cell. Probability of failure was relatively low (as low as 0; mean = 0.24) if action potentials that occurred during spontaneous activity of the mossy cell were excluded. These data suggest that unitary EPSPs produced by pyramidal cells are strongly affected by concurrent synaptic inputs to the mossy cell. 4. Unitary EPSPs were not clearly affected by manipulation of the mossy cell's membrane potential. This is consistent with the recent report that area CA3 pyramidal cells innervate distal dendrites of mossy cells (Kunkel et al. 1993). Such a distal location also may contribute to the high incidence of apparent failures. 5. Characteristics of unitary EPSPs generated by pyramidal cells were compared with the properties of the unitary EPSPs produced by granule cells. In two slices, pyramidal cell and granule cell inputs to the same mossy cell were compared. In other slices, inputs to different mossy cells were compared. In all experiments, unitary EPSPs produced by granule cells were larger in amplitude but similar in time course to unitary EPSPs produced by pyramidal cells. Probability of failure was lower and paired-pulse facilitation more common among EPSPs triggered by granule cells.(ABSTRACT TRUNCATED AT 400 WORDS)     

Calcium dynamics in single spines during coincident pre- and postsynaptic activity depend on relative timing of back-propagating action potentials and subthreshold excitatory postsynaptic potentials

We compared the transient increase of Ca2+ in single spines on basal dendrites of rat neocortical layer 5 pyramidal neurons evoked by subthreshold excitatory postsynaptic potentials (EPSPs) and back-propagating action potentials (APs) by using calcium fluorescence imaging. AP-evoked Ca2+ transients were detected in both the spines and in the adjacent dendritic shaft, whereas Ca2+ transients evoked by single EPSPs were largely restricted to a single active spine head. Calcium transients elicited in the active spines by a single AP or EPSP, in spines up to 80 micro(m) for the soma, were of comparable amplitude. The Ca2+ transient in an active spine evoked by pairing an EPSP and a back-propagating AP separated by a time interval of 50 ms was larger if the AP followed the EPSP than if it preceded it. This difference reflected supra- and sublinear summation of Ca2+ transients, respectively. A comparable dependence of spinous Ca2+ transients on relative timing was observed also when short bursts of APs and EPSPs were paired. These results indicate that the amplitude of the spinous Ca2+ transients during coincident pre- and postsynaptic activity depended critically on the relative order of subthreshold EPSPs and back-propagating APs. Thus, in neocortical neurons the amplitude of spinous Ca2+ transients could encode small time differences between pre- and postsynaptic activity.     

Modulation by substance P of synaptic transmission in the mouse hippocampal slice

The modulatory action of substance P on synaptic transmission of CA1 neurons was studied using intra- or extracellular recording from the mouse hippocampal slice preparation. Bath-applied substance P (2-4 microM) or the selective NK1 receptor agonist substance P methylester (SPME, 10 nM-5 microM) depressed field potentials (recorded from stratum pyramidale) evoked by focal stimulation of Schaffer collaterals. This effect was apparently mediated via NK1 receptors since it was completely blocked by the selective NK1 antagonist SR 140333. The field potential depression by SPME was significantly reduced in the presence of bicuculline. Intracellular recording from CA1 pyramidal neurons showed that evoked excitatory postsynaptic potentials (EPSPs) and evoked inhibitory postsynaptic potentials (IPSPs) were similarly depressed by SPME, which at the same time increased the frequency of spontaneous GABAergic events and reduced that of spontaneous glutamatergic events. The effects of SPME on spontaneous and evoked IPSPs were prevented by the ionotropic glutamate receptor blocker kynurenic acid. In tetrodotoxin (TTX) solution, no change in either the frequency of spontaneous GABAergic and glutamatergic events or in the amplitude of responses of pyramidal neurons to 4 microM alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) or 10 microM N-methyl-D-aspartate (NMDA) was observed. On the same cells, SPME produced minimal changes in passive membrane properties unable to account for the main effects on synaptic transmission. The present data indicate that SPME exerted its action on CA1 pyramidal neurons via a complex network mechanism, which is hypothesized to involve facilitation of a subset of GABAergic neurons with widely distributed connections to excitatory and inhibitory cells in the CA1 area.     

Inhibition evoked from primary afferents in the electrosensory lateral line lobe of the weakly electric fish (Apteronotus leptorhynchus)

Distal versus proximal inhibitory shaping of feedback excitation in the electrosensory lateral line lobe: implications for sensory filtering. J. Neurophysiol. 80: 3214-3232, 1998. The inhibition controlling the indirect descending feedback (parallel fibers originating from cerebellar granule cells in the eminentia posterior pars granularis) to electrosensory lateral line lobe (ELL) pyramidal cells was studied using intracellular recording techniques in vitro. Parallel fibers (PF) contact stellate cells and dendrites of ventral molecular layer (VML) GABAergic interneurons. Stellate cells provide local input to pyramidal cell distal dendrites, whereas VML cells contact their somata and proximal dendrites. Single-pulse stimulation of PF evoked graded excitatory postsynaptic potentials (EPSPs) that were blocked by alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid and N-methyl--aspartate (NMDA) antagonists. The EPSPs peaked at 6.4 +/- 1.8 ms (mean +/- SE; n = 11) but took >50 ms to decay completely. Tetanic stimulation (100 ms, 100 Hz) produced a depolarizing wave with individual EPSPs superimposed. The absolute amplitude of the individual EPSPs decreased during the train. Spike rates, established by injected current, mostly were increased, but in some cells were decreased, by tetanic stimulation. Global application of a gamma-aminobutyric acid-A (GABAA) antagonist to the recorded cell's soma and apical dendritic region increased the EPSP peak and decay phase amplitudes. Tetanic stimulation always increased current-evoked spike rates after GABAA blockade during, and for several hundred milliseconds after, the stimulus. Application of a GABAB antagonist did not have any significant effects on the PF-evoked response. This, and the lack of any long hyperpolarizing inhibitory postsynaptic potentials, suggests that VML and stellate cell inhibition does not involve GABAB receptors. Focal GABAA antagonist applications to the dorsal molecular layer (DML) and pyramidal cell layer (PCL) had contrasting effects on PF-evoked EPSPs. DML GABAA blockade significantly increased the EPSP peak amplitude but not the decay phase of the EPSP, whereas PCL GABAA-blockade significantly increased the decay phase, but not the EPSP peak, amplitude. The order of antagonist application did not affect the outcome. On the basis of the known circuitry of the ELL, we conclude that the distal inhibition originated from GABAergic molecular layer stellate cells and the proximal inhibition originated from GABAergic cells of the ventral molecular layer (VML cells). Computer modeling of distal and proximal inhibition suggests that intrinsic differences in IPSP dynamics between the distal and proximal sites may be amplified by voltage-dependent NMDA receptor and persistent sodium currents. We propose that the different time courses of stellate cell and VML cell inhibition allows them to act as low- and high-pass filters respectively on indirect descending feedback to ELL pyramidal cells.     

Regulation of the NMDA component of EPSPs by different components of postsynaptic GABAergic inhibition: computer simulation analysis in piriform cortex

Regulation of the NMDA component of EPSPs by different components of postsynaptic GABAergic inhibition: computer simulation analysis in piriform cortex. J. Neurophysiol. 78: 2546-2559, 1997. Physiological analysis in the companion paper demonstrated that gamma-aminobutyric acid-A (GABAA)-mediated inhibition in piriform cortex is generated by circuits that are largely independent in apical dendritic and somatic regions of pyramidal cells and that GABAA-mediated inhibitory postsynaptic currents (IPSCs) in distal dendrites have a slower time course than those in the somatic region. This study used modeling methods to explore these characteristics of GABAA-mediated inhibition with respect to regulation of the N-methyl--aspartate (NMDA) component of excitatory postsynaptic potentials. Such regulation is relevant to understanding NMDA-dependent long-term potentiation (LTP) and the integration of repetitive synaptic inputs that can activate the NMDA component as well as pathological processes that can be activated by overexpression of the NMDA component. A working hypothesis was that the independence and differing properties of IPSCs in apical dendritic and somatic regions provide a means whereby the NMDA component and other dendritic processes can be controlled by way of GABAergic tone without substantially altering system excitability. The analysis was performed on a branched compartmental model of a pyramidal cell in piriform cortex constructed with physiological and anatomic data derived by whole cell patch recording. Simulations with the model revealed that NMDA expression is more effectively blocked by the slow GABAA component than the fast. Because the slow component is present in greater proportion in apical dendritic than somatic regions, this characteristic would increase the capacity of dendritic IPSCs to regulate NMDA-mediated processes. The simulations further revealed that somatic-region GABAergic inhibition can regulate the generation of action potentials with little effect on the NMDA component generated by afferent fibers in apical dendrites. As a result, if expression of the NMDA component or other dendritic processes were enabled by selective block of dendritic inhibition, for example, by centrifugal fiber systems that may regulate learning and memory, the somatic-region IPSC could preserve system stability through feedback regulation of firing without counteracting the effect of the dendritic-region block. Simulations with paired inputs revealed that the dendritic GABAA-mediated IPSC can regulate the extent to which a strong excitatory input facilitates the NMDA component of a concurrent weak input, providing a possible mechanism for control of "associative LTP" that has been demonstrated in this system. Postsynaptic GABAB-mediated inhibition had less effect on the NMDA component than either the fast or slow GABAA components. Depolarization from a concomitant alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) component also was found to have comparatively little effect on current through the NMDA channel because of its brief time course.     

Enhanced activation of NMDA receptor responses at the immature retinogeniculate synapse

The maturation of retinogeniculate excitatory transmission and intrathalamic inhibition was studied in slices of the dorsal LGN obtained from ferrets during the first 2 postnatal months. Response to optic tract stimulation at neonatal ages consisted of slow EPSPs lasting several hundred milliseconds. Application of the NMDA receptor antagonist D-(-)-2-amino-5-phosphonovaleric acid (D-APV) during the first 2 postnatal weeks resulted in EPSPs that were reduced in peak amplitude and dramatically curtailed in duration, indicating that NMDA receptors participate strongly in retinogeniculate transmission at the immature synapse. Gradually, EPSPs became shorter in duration such that after the second postnatal week, the retinogeniculate EPSPs were only a few milliseconds in duration. At this late stage of development responses were remarkably less affected by application of D-APV. These changes in contribution of NMDA receptors to retinogeniculate transmission were found to be due to the development of strong IPSPs, the result of gradual maturation of activation of GABAergic inhibition. Indeed, application of bicuculline methiodide to block GABAA receptor-mediated IPSPs strongly enhanced the NMDA component of the EPSPs in more mature cells. The voltage dependence and kinetics of NMDA-induced excitatory postsynaptic currents (NMDA EPSCs) were characterized by voltage-clamp recordings after blocking AMPA/kainate receptors with 6-cyano-7-nitroquinoxaline-2,3-dione and GABAA receptors wit' bicuculline methiodide. The voltage dependence of the NMDA EPSCs remained unaltered with age. During the first postnatal month the kinetic properties of the NMDA EPSCs also remained unaltered, but a reduction in EPSC duration was observed within the following weeks, well after the critical period of anatomical reorganization.(ABSTRACT TRUNCATED AT 250 WORDS)     

Amplification of tumor immunity by gene transfer of the co-stimulatory 4-1BB ligand: synergy with the CD28 co-stimulatory pathway

The superficial cells of the entorhinal cortex (EC), main input to the hippocampus, receive a serotonergic input from the raphe nuclei and express 5-hydroxytryptamine creatine sulfate complex (5-HT) receptors at high density. With the use of intracellular recordings, we investigated the effects of serotonin on synaptic inhibition of layer II and III neurons of the EC. Serotonin reduced both polysynaptic fast and slow inhibitory postsynaptic potentials (IPSPs) in projection neurons of the superficial EC. Polysynaptic fast and slow IPSPs were depressed by serotonin in a dose-dependent manner (0.1-100 microM). Serotonin in a concentration of 1 microM reduced the amplitudes of polysynaptic fast and slow IPSPs by approximately 40 and 50%, respectively. To identify the subtype of the 5-HT-receptor mediating the effects on polysynaptic IPSPs, we applied various 5-HT-receptor agonists and antagonists. Although the serotonin agonists for the 5-HT1B,2C,3 receptors were ineffective, the effects were mimicked by the 5-HT1A-receptor agonists (8-OH-DPAT, 5-CT) and prevented by the 5-HT1A-receptor antagonist NAN-190. To look at the direct effects of 5-HT on inhibitory interneurons, we elicited monosynaptic IPSPs in the absence of excitatory synaptic transmission. In contrast to the polysynaptic IPSPs, monosynaptic IPSPs were not significantly affected by serotonin. Recordings from putative inhibitory interneurons revealed that their excitatory postsynaptic potentials (EPSPs) were reversibly reduced by serotonin. We conclude that serotonin suppresses polysynaptic inhibition in projection neurons of layers II and III of the EC by depression of EPSPs on inhibitory interneurons via 5-HT1A receptors.     

Contributions of individual layer 2-5 spiny neurons to local circuits in macaque primary visual cortex

We studied excitatory local circuits in the macaque primary visual cortex (VI) to investigate their relationships to the magnocellular (M) and parvocellular (P) streams. Sixty-two intracellularly labeled spiny neurons in layers 2-5 were analyzed. We made detailed observations of the laminar and columnar specificity of axonal arbors and noted correlations with dendritic arbors. We find evidence for considerable mixing of M and P streams by the local circuitry in VI. Such mixing is provided by neurons in the primary geniculate recipient layer 4C, as well as by neurons in both the supragranular and infragranular layers. We were also interested in possible differences in the axonal projections of neurons with different dendritic morphologies. We found that layer 4B spiny stellate and pyramidal neurons have similar axonal arbors. However, we identified two types of layer 5 pyramidal neuron. The majority have a conventional pyramidal dendritic morphology, a dense axonal arbor in layers 2.4B, and do not project to the white matter. Layer 5 projection neurons have an unusual "backbranching" dendritic morphology (apical dendritic branches arc downward rather than upward) and weak or no axonal arborization in layers 2-4B, but have long horizontal axonal projections in layer 5B. We find no strong projection from layer 5 pyramidal neurons to layer 6. In macaque V1 there appears to be no single source of strong local input to layer 6; only a minority of cells in layers 2-5 have axonal branches in layer 6 and these are sparse. Our results suggest that local circuits in V1 mediate interactions between M and P input that are complex and not easily incorporated into a simple framework.     

Anoxic depression of excitatory and inhibitory postsynaptic potentials in rat neocortical slices

1. The effects of brief anoxia (4-6 min replacement of O2 by N2) on synaptic potentials evoked from layer IV and/or the white matter were studied in pyramidal neurons of layers II-III from rat neocortical slices. 2. The early and late components of excitatory postsynaptic potentials (EPSPs) showed differential sensitivity to anoxia: within 2 min the late EPSP (lEPSP) disappeared, whereas the amplitude of the early EPSP (eEPSP) decreased by 70% at 5 min of anoxia. Recovery was complete within 4-11 min. 3. Both fast and slow inhibitory postsynaptic potentials (IPSPs) were extremely sensitive to lack of O2 and were abolished earlier than the lEPSP evoked by the same stimulus. As well, recovery of the IPSPs was always more delayed than that of the EPSPs. 4. A transient increase in excitability during early anoxia and/or midrecovery, manifested as enhanced probability of spiking in 25% of neurons, is attributed to the higher sensitivity of IPSPs compared with EPSPs. 5. The anoxic-induced depression of the lEPSP and IPSPs, which are generated close to the soma, is not due to depolarization-induced occlusion; however, occlusion may cause an attenuation of the eEPSP at dendritic sites. 6. The depression of the EPSPs is not a result of a decreased transmembrane Na+ gradient after inactivation of Na-K-adenosine triphosphatase (Na-K-ATPase). Although ouabain induced a depolarization similar to that of anoxia, it did not affect EPSP amplitude.(ABSTRACT TRUNCATED AT 250 WORDS)     

Amplification of perforant-path EPSPs in CA3 pyramidal cells by LVA calcium and sodium channels

The perforant path forms a monosynaptic connection between the cells of layer II of the entorhinal cortex and the pyramidal cells in hippocampal area CA3. Although this projection is prominent anatomically, very little is known about the physiological properties of this input. The distal location of these synapses suggests that somatically recorded perforant-path excitatory postsynaptic potentials (EPSPs) may be influenced by the activation of voltage-dependent channels in CA3 cells. We observed that perforant-path EPSPs are reduced (by approximately 25%) by blockade of postsynaptic low-voltage-activated calcium and sodium channels, indicating that perforant-path EPSPs are amplified by the activation of these channels. These data suggest that the perforant path may represent an important and highly modifiable direct connection between the entorhinal cortex and area CA3.     

Serotonin depresses excitatory synaptic transmission and depolarization-evoked Ca2+ influx in rat basolateral amygdala via 5-HT1A receptors

The actions of serotonin on rat basolateral amygdala neurons were studied with conventional intracellular recording techniques and fura-2 fluorimetric recordings. Bath application of 5-hydroxytryptamine (5-HT or serotonin) reversibly suppressed the excitatory postsynaptic potential in a concentration-dependent manner without affecting the resting membrane potential and neuronal input resistance. Extracellular Ba2+ or pertussis toxin pretreatment did not affect the depressing effect of 5-HT suggesting that it is not mediated through activation of Gi/o protein-coupled K+ conductance. The sensitivity of postsynaptic neurons to glutamate receptor agonist was unaltered by the 5-HT pretreatment. In addition, the magnitude of paired-pulse facilitation was increased in the presence of 5-HT indicating a presynaptic mode of action. The effect of 5-HT was mimicked by the selective 5-HT1A agonist 8-hydroxy-dipropylaminotetralin (8-OH-DPAT) and was blocked by the selective 5-HT1A antagonist 1-(2-methoxyphenyl)-4[4-(2-phthalimido)butyl]piperazine oxadiazol-3-yl]methyl]phenyl]-methanesulphonamide. In contrast, the selective 5-HT2 receptor antagonist ketanserin failed to affect the action of 5-HT. The effects of 5-HT and 8-OH-DPAT on the high K+-induced increase in [Ca2+]i were studied in acutely dissociated basolateral amygdala neurons. High K+-induced increase in [Ca2+]i was blocked by Ca2+-free solution and Cd2+ suggesting that Ca2+ entry responsible for the depolarization-evoked increase in [Ca2+]i occurred through voltage-dependent Ca2+ channels. Application of 5-HT and 8-OH-DPAT reduced the K+-induced Ca2+ influx in a concentration-dependent manner. The effect of 5-HT was completely abolished in slices pretreated with Rp-cyclic adenosine 3',5'-monophosphothioate (Rp-cAMP), a regulatory site antagonist of protein kinase A, suggesting that 5-HT may act through a cAMP-dependent mechanism. Taken together, these results suggest that functional 5-HT1A receptors are present in the excitatory terminals and mediate the 5-HT inhibition of synaptic transmission in the amygdala.     

Glutamate currents in morphologically identified human dentate granule cells in temporal lobe epilepsy

Glutamate-receptor-mediated synaptic transmission was studied in morphologically identified hippocampal dentate granule cells (DGCs; n = 31) with the use of whole cell patch-clamp recording and intracellular injection of biocytin or Lucifer yellow in slices prepared from surgically removed medial temporal lobe specimens of epileptic patients (14 specimens from 14 patients). In the current-clamp recording, low-frequency stimulation of the perforant path generated depolarizing postsynaptic potentials that consisted of excitatory postsynaptic potentials and phase-inverted inhibitory postsynaptic potentials mediated by the gamma-aminobutyric acid-A (GABA(A)) receptor at a resting membrane potential of -62.7 +/- 2.0 (SE) mV. In the voltage-clamp recording, two glutamate conductances, a fast alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-receptor-mediated excitatory postsynaptic current (EPSC; AMPA EPSC) and a slowly developing N-methyl-D-aspartate (NMDA)-receptor-mediated EPSC (NMDA EPSC), were isolated in the presence of a GABA(A) receptor antagonist. NMDA EPSCs showed a voltage-dependent increase in conductance with depolarization by exhibiting an N-shaped current-voltage relationship. The slope conductance of the NMDA EPSC ranged from 1.1 to 9.4 nS in 31 DGCs, reaching up to twice the size of the AMPA conductance. This widely varying size of the NMDA conductance resulted in the generation of double-peaked EPSCs and a nonlinear increase of the slope conductance of up to 37.5 nS with positive membrane potentials, which resembled "paroxysmal currents," in a subpopulation of the neurons. In contrast, AMPA EPSCs, which were isolated in the presence of an NMDA receptor antagonist (2-amino-5-phosphonovaleric acid), showed voltage-independent linear changes in the current-voltage relationship and were blocked by 6-cyano-7-nitroquinoxaline-2,3-dione. The AMPA conductance showed little variance, regardless of the size of the NMDA conductance of a given neuron. The average AMPA slope conductance was 5.28 +/- 0.65 (SE) nS in 31 human DGCs. This value was similar to AMPA EPSC conductances in normal rat DGCs (5.35 +/- 0.52 nS, mean +/- SE; n = 55). Dendritic morphology and spine density were quantified in the individual DGCs to assess epileptic pathology. Dendritic spine density showed an inverse correlation (r2 = 0.705) with a slower rise time and a longer half-width of the excitatory postsynaptic potentials mediated by the NMDA receptor. It is concluded that both AMPA and NMDA EPSCs contribute to human DGC synaptic transmission in epileptic hippocampus. However, a wide range of changes in the slope conductance of the NMDA EPSCs suggests that the NMDA-receptor-mediated conductance could be altered in human epileptic DGCs. These changes may influence the generation of chronic subthreshold epileptogenic synaptic activity and give rise to pathological excitation leading to epileptic seizures and dendritic pathology.     

Comparative electrophysiological aspects of aluminum actions on central neurons and neuronal synapses of invertebrate and vertebrate animals

Aluminum compounds (Al) increased the membrane potential (Em) and decreased the input resistance (Rin) of identified snail neurons. The neuronal excitability increased after Al withdrawal in the washing period. Cumulative Al applications caused dose-dependent modulation of Em and Rin in most of the studied neurons. Two phase actions were recorded on stimulus evoked excitatory postsynaptic potentials (EPSPs) or currents (EPSCs) at pH 6.5-6.9. The Al induced facilitation followed by attenuation were statistically significant, time- and dose-dependent events. They could be recorded at each Em. Low affinity and slow binding kinetics could characterize the Al actions on the neurons. Al showed pH- dependent suppression of EPSPs or EPSCs in some neurons. Our findings are partially comparable with Al induced electrophysiological and pharmacological modifications reported on vertebrate neurons.     

M100907, a selective 5-HT2A receptor antagonist and a potential antipsychotic drug, facilitates N-methyl-D-aspartate-receptor mediated neurotransmission in the rat medial prefrontal cortical neurons in vitro

The technique of intracellular recording was used to examine the effect of M100907 (formerly MDL 100907), a highly selective 5-HT2A receptor antagonist and a potential antipsychotic drug (APD), on N-methyl-D-aspartate (NMDA) and (+/-)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor-mediated responses in pyramidal cells of the rat medial prefrontal cortex in in vitro brain slice preparations. Bath administration of M100907, but not its inactive stereoisomer M100009, produced a 350% to 550% increase of NMDA-induced responses in a concentration-dependent manner with an EC50 value of 14 nmol/L, reminiscent of the action of clozapine. M100907 did not alter AMPA responses. Moreover, M100907 significantly increased the amplitude and duration of excitatory postsynaptic potentials and currents evoked by electrical stimulation of the forceps minor. We have generated several lines of evidence indicating that M100907 enhances glutamate receptor-mediated neurotransmission in pyramidal cells of the medial prefrontal cortex by facilitating NMDA-induced release of excitatory amino acids. The robust potentiation of NMDA receptor-mediated neurotransmission may explain, at least partly, the potential antipsychotic action of this compound. Furthermore, if M100907 proves to be an effective APD and if our findings can be extended to other atypical APDs, which are known to possess a relatively high affinity to 5-HT2A receptors, they may account for the purported efficacy of atypical APDs in alleviating some negative symptoms such as cognitive and executive functions.     

Electrophysiological and pharmacological characterization of the direct perforant path input to hippocampal area CA3

Monosynaptic perforant path responses evoked by subicular stimulation were recorded from CA3 pyramidal cells of rat hippocampal slices. These monosynaptic responses were isolated by using low intensities of stimulation and by placing a cut through the mossy fibers. Perforant path-evoked responses consisted of both excitatory and inhibitory components. Excitatory postsynaptic currents (EPSCs) were mediated by both alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acidreceptors (AMPAR) and N-methyl--aspartate receptors (NMDAR). Inhibitory postsynaptic currents consisted of gamma-aminobutyric acid-A (GABAA-) and -B (GABAB)-receptor-mediated components. At membrane potentials more positive than -60 mV and at physiological [Ca2+]/[Mg2+] ratios, >30% of perforant path evoked EPSC was mediated by NMDARs. This value varied as a function of the membrane voltage and external [Mg2+]. Two types of responses were observed after low-intensity stimulation of the perforant path. The first type of response showed paired-pulse facilitation and was reduced by 2-amino-4-phosphonobutyric acid (AP4). The second type of response showed paired-pulse depression and was reduced by baclofen. Electrophysiological and pharmacological characteristics of these two types of responses are similar to the properties of lateral and medial perforant path-evoked EPSPs in the dentate gyrus.     

Monosynaptic GABA-mediated inhibitory postsynaptic potentials in CA1 pyramidal cells of hyperexcitable hippocampal slices from kainic acid-treated rats

To examine the mechanisms underlying chronic epileptiform activity, field potentials were first recorded to identify hyperexcitable hippocampal slices from kainic acid-treated rats. Intracellular recordings were then obtained from CA1 pyramidal cells in the hyperexcitable areas. Twenty-two of the 47 cells responded to electrical stimulation of the stratum radiatum with a burst of two or more action potentials and reduced early inhibitory postsynaptic potentials, and were considered hyperexcitable. The remaining 25 cells were not hyperexcitable, displaying a single action potential and biphasic inhibitory postsynaptic potentials after stimulation, like control cells (n = 20). A long duration, voltage-sensitive component was associated with subthreshold excitatory postsynaptic potentials in the majority of hyperexcitable (12/15) and non-hyperexcitable (3/5) cells examined from kainic acid-treated animals, but not from cells (1/10) of control animals. Stimulation of stratum radiatum during pharmacological blockade of ionotropic excitatory amino acid synaptic transmission elicited biphasic monosynaptic inhibitory postsynaptic potentials in all hyperexcitable (n = 9) and non-hyperexcitable (n = 9) cells tested from kainate-treated animals, as well as in control cells (n = 8). The mean amplitude, latency to peak, equilibrium potential, and conductance changes of early and late monosynaptic inhibitory postsynaptic potentials were not different between cells of kainic acid-treated and control animals. In seven hyperexcitable cells tested, the early component of monosynaptic inhibitory postsynaptic potentials was significantly reduced by the GABAA receptor antagonist bicuculline (100-200 microM). The late component was significantly decreased by the GABAB receptor antagonist 2-hydroxysaclofen (1-2 mM; n = 3). Comparable effects were observed on early and late monosynaptic inhibitory postsynaptic potentials in non-hyperexcitable cells (n = 4) from kainic acid-treated animals and control cells (n = 5). These results suggest that GABAergic synapses on hyperexcitable hippocampal pyramidal cells of kainate-treated rats are intact and functional. Therefore, epileptiform activity in the kainate-lesioned hippocampus may not arise from a disconnection of GABAergic synapses made by inhibitory interneurons on pyramidal cells. The hyperexcitability may be due to underactivation of inhibitory interneurons and/or reorganization of excitatory inputs to pyramidal cells since, in kainate-treated animals, pyramidal cells appear to express additional excitatory mechanisms.     

Effect of piribedil and its metabolite, S584, on brain lipid peroxidation in vitro and in vivo

We have previously reported that neonatal (P3) serotonin (5-HT) depletion results in a significant decrease in the number of dendritic spines per 50 microns of dendritic length on dentate granule cells. This effect is specific and permanent. Neither total dendritic length nor the number of dendritic segments is affected by 5-HT depletion. The area dentata contains a dense 5-HT1a receptor population that is present in the at birth. Therefore, 5-HT1a receptors represented a likely candidate for the mediation of the effects of 5-HT on developing granule cells. The present study used the drugs buspirone and NAN-190, which have been shown to be an agonist and antagonist respectively at postsynaptic 5-HT1a receptors in vivo, to test the idea that neurotrophic actions of 5-HT result from 5-HT1a receptor stimulation. Following 5-HT depletion with PCA, pups received daily injections of buspirone (1.0 mg/kg) from P5 to P14. Granule cell morphology was then studied using intracellular filling with Neurobiotin on P14, P21 and P60. Buspirone treatment prevented the loss of dendritic spines previously shown to follow 5-HT depletion with PCA. No other morphological parameters were significantly changed by buspirone treatment. Naive pups received daily injections of NAN-190 from P3 to P14. One group received 1.0 mg/kg while a second group received 3.5 mg/kg. Both doses of NAN-190 resulted in dendritic spine loss comparable to that obtained with neonatal PCA treatment. This loss was permanent suggesting that the first two postnatal weeks may represent a critical period for the action of 5-HT on developing granule cells. Significant, dose-dependent changes in total dendritic length and number of dendritic segments reminiscent of the effects of norepinephrine depletion were also observed in NAN-190-treated rats. We suspect that this change is the result of the action NAN-190 at alpha receptors and is therefore distinct from the specific effect of 5-HT on the number of dendritic spines. The NAN-190 experiment also shows that the loss of dendritic spines is a function of decreased stimulation of 5-HT1a receptors and not the loss of 5-HT terminal membrane.     

Cell-specific alterations in synaptic properties of hippocampal CA1 interneurons after kainate treatment

Cell-specific alterations in synaptic properties of hippocampal CA1 interneurons after kainate treatment. J. Neurophysiol. 80: 2836-2847, 1998. Hippocampal sclerosis and hyperexcitability are neuropathological features of human temporal lobe epilepsy that are reproduced in the kainic acid (KA) model of epilepsy in rats. To assess directly the role of inhibitory interneurons in the KA model, the membrane and synaptic properties of interneurons located in 1) stratum oriens near the alveus (O/A) and 2) at the border of stratum radiatum and stratum lacunosum-moleculare (LM), as well as those of pyramidal cells, were examined with whole cell recordings in slices of control and KA-lesioned rats. In current-clamp recordings, intrinsic cell properties such as action potential amplitude and duration, amplitude of fast and medium duration afterhyperpolarizations, membrane time constant, and input resistance were generally unchanged in all cell types after KA treatment. In voltage-clamp recordings, the amplitude and conductance of pharmacologically isolated excitatory postsynaptic currents (EPSCs) were significantly reduced in LM interneurons of KA-treated animals but were not significantly changed in O/A and pyramidal cells. The rise time of EPSCs was not significantly changed in any cell type after KA treatment. In contrast, the decay time constant of EPSCs was significantly faster in O/A interneurons of KA-treated rats but was unchanged in LM and pyramidal cells. The amplitude and conductance of pharmacologically isolated gamma-aminobutyric acid-A (GABAA) inhibitory postsynaptic currents (IPSCs) were not significantly changed in any cell type of KA-treated rats. The rise time and decay time constant of GABAA IPSCs were significantly faster in pyramidal cells of KA-treated rats but were not significantly changed in O/A and LM interneurons. These results suggest that complex alterations in synaptic currents occur in specific subpopulations of inhibitory interneurons in the CA1 region after KA lesions. A reduction of evoked excitatory drive onto inhibitory cells located at the border of stratum radiatum and stratum lacunosum-moleculare may contribute to disinhibition and polysynaptic epileptiform activity in the CA1 region. Compensatory changes, involving excitatory synaptic transmission on other interneuron subtypes and inhibitory synaptic transmission on pyramidal cells, may also take place and contribute to the residual, functional monosynaptic inhibition observed in principal cells after KA treatment.