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PubMed2005

Cytoskeletal, synaptic, and nuclear protein changes associated with rat interface organotypic hippocampal slice culture development.

Although organotypic hippocampal slice cultures (OHSCs) are used to study function within the hippocampus, the effect of maintenance in vitro upon protein expression is not fully understood. Therefore, we examined developmental changes in cultures prepared from P8 rats and maintained on porous membranes between medium and atmosphere. Between 7 and 28 days following explantation, altered hippocampal morphology could not be detected despite a significant decrease in both MAP-2c and a mid-range tau isoform by 21 DIV. During the same period, lower GFAP expression was observed, and GFAP labeling suggested a migration of astrocytes to the slice-atmosphere interface. In contrast, levels of the synaptic proteins synaptophysin and PSD-95 were significantly increased, but GAP-43 was not. The preservation of myelinated axons and synapses, along with glial and endothelial cells, was confirmed by ultrastructural analysis. Furthermore, intranuclear inclusion bodies, which are associated with normal aging in vivo, were detected in the CA1 pyramidal layer in cultures older than 14 DIV. When OHSCs were maintained for approximately 3, 4, and 10 weeks, a rise and then fall in the expression of synaptophysin and, especially, PSD-95 were found, and the biphasic trend paralleled by significant changes in Schaffer collateral-evoked excitatory post-synaptic potentials from CA1 neurons. Our data not only describe changes in cytoskeletal, synaptic, and nuclear proteins related to the maintenance of interface OHSCs, but also emphasize the potential of the model for the study of age-related phenomena within the hippocampus.

Animals↗
PubMedDate not supplied

Protein kinase C injection into hippocampal pyramidal cells elicits features of long term potentiation.

Long-term potentiation (LTP) in the hippocampus is an interesting example of synaptic plasticity because of its induction by physiological discharge rates and its long duration. Of the possible biochemical mechanisms that regulate prolonged changes in cell function, protein phosphorylation is a particularly attractive candidate. We have therefore examined the effect of intracellular injection of calcium/diacylglycerol-dependent protein kinase (protein kinase C (PKC] in CA1 pyramidal neurones in hippocampal slices. Injection of the active enzyme elicited long-lasting enhancement of synaptic transmission, similar to LTP, whereas inactivated kinase failed to do so. The observed changes included an increased amplitude of the excitatory post-synaptic potential (e.p.s.p.) and an increased probability of firing and a reduced latency of the associated actin potential.

Action Potentials↗
PubMed2001

Multiple personalities: synaptic target cells as introverts and extroverts.

The intricate process of wiring a neuronetwork requires a high degree of accuracy in the communication between pre- and post-synaptic cells. While presynaptic cells have been widely recognized for their dynamic role in synaptic matchmaking, post-synaptic cells have historically been overlooked as passive targets. Recent studies in the Drosophila embryonic neuromuscular system provide compelling evidence that post-synaptic cells participate actively in the synaptogenic process. Endocytosis allows them to quickly modify the array of molecular cues they provide on their surfaces and the extension of dynamic filopodia allows post-synaptic cells to engage in direct long-distance communication. By making use of familiar cellular mechanisms such as endocytosis and filopodia formation, post-synaptic cells may be able to communicate more effectively with potential synaptic partners.

Animals↗
PubMed1997

Effects of cholinergic modulation on responses of neocortical neurons to fluctuating input.

Neocortical neurons in vivo are spontaneously active and intracellular recordings have revealed strongly fluctuating membrane potentials arising from the irregular arrival of excitatory and inhibitory synaptic potentials. In addition to these rapid fluctuations, more slowly varying influences from diffuse activation of neuromodulatory systems alter the excitability of cortical neurons by modulating a variety of potassium conductances. In particular, acetylcholine, which effects learning and memory, reduces the slow alterhyperpolarization, which contributes to spike frequency adaptation. We used whole-cell patch-clamp recordings of pyramidal neurons in neocortical slices and computational simulations to show, first, that when fluctuating inputs were added to a constant current pulse, spike frequency adaptation was reduced as the amplitude of the fluctuations was increased. High-frequency, high-amplitude fluctuating inputs that resembled in vivo conditions exhibited only weak spike frequency adaptation. Second, bath application of carbachol, a cholinergic agonist, significantly increased the firing rate in response to a fluctuating input but minimally displaced the spike times by < 3 ms, comparable to the spike jitter observed when a visual stimulus is repeated under in vivo conditions. These results suggest that cholinergic modulation may preserve information encoded in precise spike timing, but not in interspike intervals, and that cholinergic mechanisms other than those involving adaptation may contribute significantly to cholinergic modulation of learning and memory.

Animals↗
PubMedDate not supplied

Diagnosis of the level and severity of cord lesion in cervical spondylotic myelopathy. Spinal evoked potentials.

Experimental and clinical studies were performed to measure the segmental spinal evoked potential (SEP) of the cervical cord after stimulation of the median nerve, to determine the location, severity, and outcome of cord lesion in patients who had cervical spondylotic myelopathy. The SEP in control subjects consisted of two waves, the primary reactive R-wave and the secondary reactive N-wave. The R-wave related to both the dorsal root potential and the funiculus posterior potential; the N-wave related to the post-synaptic potential in the spinal cord. In the early stages of cervical spondylotic myelopathy, the N-wave weakens or disappears at the level of lesion. Then, as damage progresses, the R-wave also weakens or disappears.

Adult↗
PubMed2000

Information coding via spontaneous oscillations in neural ensembles.

How information is encoded and decoded via spontaneous oscillations is investigated by using an ensemble of Hodgkin-Huxley neurons. A signal can be encoded in spontaneous and highly irregular spike trains via high-order rate coding with the second-order statistics being relevant, in which the temporal structure and the correlation between the spikes are taken into account. Although the encoded information is implicitly contained in the spike train, it can be retrieved in the post-synaptic potential. The spontaneous oscillation is filtered and the irregularity of the spike train is suppressed. In particular, we show that an arbitrary signal can be transmitted reliably through spontaneous and highly irregular spike trains, and then be reconstructed downstream in the information transmission pathway.

Action Potentials↗
PubMed1976

An electrophysiological study of chemical and electrical synapses on neurones in the parasympathetic cardiac ganglion of the mudpuppy, Necturus maculosus: evidence for intrinsic ganglionic innervation.

1. The cardiac ganglion of the mudpuppy is situated on a thin sheet of tissue. Two nerve cell types can be distinguished readily in the living preparation - principal cells and smaller interneurones which synapse with the principal cells. The purpose of this study was to investigate synaptic transmission and the functional organization of neuronal connections of ganglion cells with intracellular micro-electrodes. 2. Stimulation of the preganglionic, vagus, nerves evoked a large excitatory response in principal cells. About three quarters of these neurones were innervated by a single vagal axon. The remaining cells received two or more preganglionic nerve fibres. 3. The quantum content of vagal excitatory post-synaptic potentials (e.p.s.p.s) was measured. Normally, the e.p.s.p. was suprathreshold and consisted of about twenty-two quanta, whereas only about nine quanta were required to reach threshold and initiate an action potential. 4. Intracellular stimulation of principal cells evoked e.p.s.p.s in neighbouring principal cells. The responses were blocked by cholinergic antagonists. These potentials were caused by excitation of principal cell axon collateral synapses. 5. Principal cells also formed electrical junctions with each other. These electrical junctions were very weak. Although they transmitted slow potential changes, only a small response was recorded in one cell when an electrically coupled neighbouring cell fired an impulse. The resistance of the electrical junction between principal cells was calculated to be about 5-8 X 10(8) omega. 6. Stable penetrations of interneurones were only rarely achieved, making it difficult to study their functional relationship to principal cells. Action potentials were recorded from interneurones in a few instances. 7. These data demonstrate that parasympathetic ganglion cells in the heart of the mudpuppy receive innervation from more than one source involving both chemical and electrical synapses, and that some of the synapses are intrinsic to the ganglion.

Action Potentials↗
PubMed1989

Bicuculline- and phaclofen-sensitive components of N-methyl-D-aspartate-induced hyperpolarizations in rat dorsolateral septal nucleus neurones.

1. Intracellular recordings were made from rat dorsolateral septal nucleus (DLSN) neurones in vitro. Pressure application (puff) of N-methyl-D-aspartate (NMDA) usually produced a hyperpolarization followed by a depolarization in normal Mg2+ solution. 2. A hyperpolarizing response and a depolarizing response could be evoked separately by appropriate positioning of the puff pipette. The NMDA-induced hyperpolarization was blocked by a low-Ca2+ solution or a tetrodotoxin (TTX)-containing solution, but under these conditions the NMDA-induced depolarization was spared. The amplitude of NMDA-induced hyperpolarizations was 7.7 +/- 2.7 mV (n = 13) at the resting membrane potential level. 3. An NMDA-induced hyperpolarization had two components. A fast component reversed at about -76 mV and a slow component reversed at -90 mV. The reversal potential of the slow component shifted in the depolarizing direction of hyperpolarizing direction when the slice was bathed in a high or low-K+ solution, respectively. 4. The reversal potentials of NMDA-induced hyperpolarizations were similar to the synaptic potentials evoked by fimbrial stimulation. The reversal potentials of the fast and slow components were close to the IPSP reversal potential (-70 mV) and the late hyperpolarizing potential (LHP) reversal potential (-95 mV), respectively. 5. NMDA-induced hyperpolarizations were blocked by the specific NMDA receptor antagonist 2-amino-5-phosphonopontanoate (APV). The fast component of an NMDA-induced hyperpolarization was blocked by the specific GABAA receptor antagonist, bicuculline, and the slow component was depressed by the specific GABAB receptor antagonist, phaclofen. 6. Glutamate receptor subtype-specific agonists, such as kainate or quisqualate, could induce similar hyperpolarizations which had bicuculline-sensitive and insensitive components. These non-NMDA-type agonist-induced hyperpolarizations were not affected by APV (50 microM) but were blocked by kynurenic acid (1 mM). 7. We conclude that these excitatory amino acid-induced hyperpolarizations observed in the rat DLSN are mediated by GABAergic interneurones which have both non-NMDA-type and NMDA-type receptors.

2-Amino-5-phosphonovalerate↗
PubMed1985

Use of brain slices in the study of the auditory system: spatial and temporal summation of synaptic inputs in cells in the anteroventral cochlear nucleus of the mouse.

One of the more dramatic technological developments in recent years in neurobiology is the ability to extract a slice of brain tissue and to maintain its viability over extended periods of time. The development of brain slice preparations of the mammalian central nervous system provides a powerful tool for studying the physiology and pharmacology of neurons. In vitro preparations allow stable intracellular recordings to be made from cells. The characteristics of synaptic potentials and the intrinsic electrical properties can be measured while the extracellular environment is controlled and manipulated. Intracellular physiological studies in the anteroventral cochlear nucleus reveal that different morphological classes of cells have characteristic intrinsic electrical properties. Bushy cells are particularly well suited to preserve and convey the temporal firing pattern of inputs from the auditory nerve. Stellate cells can transform inputs from the auditory nerve by summing in time and space.

Animals↗
PubMed1988

An in vitro investigation of the action of ketamine on excitatory synaptic transmission in the hippocampus of the guinea-pig.

The effect of ketamine on excitatory synaptic transmission was examined in vitro in slices of guinea-pig hippocampus. Ketamine (100-500 microM) depressed the amplitude of the field excitatory post-synaptic potential (e.p.s.p.) and the population spike evoked in the dentate gyrus by stimulation of the perforant path. It did not affect the coupling between the e.p.s.p. and the population spike. Ketamine (0.2 mM) had no consistent depolarizing or hyperpolarizing effect on the resting membrane potential of the granule cells nor did it alter their input resistance. It did, however, reduce the amplitude of the intracellularly recorded e.p.s.p. Ketamine (5-200 microM) reduced the sensitivity of the granule cells to 1-glutamate applied iontophoretically but did not significantly depress the potassium-stimulated efflux of [3H]glutamate from synaptosomes. It is concluded that ketamine depresses excitatory synaptic transmission in the dentate gyrus by depressing chemical transmission probably via a post-synaptic mechanism.

Action Potentials↗
PubMed1998

Selective loss of hippocampal long-term potentiation, but not depression, following fluid percussion injury.

We investigated the early effects of in vivo fluid percussion injury (FPI) on hippocampal synaptic potentials and excitability. In vitro field potential recordings and immunocytochemistry were performed in the CA1 region in slices from naïve, post-FPI, or sham-operated rats. The following electrophysiological and morphological parameters were affected following FPI: (1) threshold for population spike generation was increased suggesting that post-FPI neurons were hypoexcitable; (2) long-term potentiation (LTP) could not be induced in injured hippocampi; (3) GFAP and inducible NO synthase (iNOS) immunoreactivity were enhanced post-FPI; and (4) following injury, synaptophysin immunoreactivity was enhanced in CA1 stratum radiatum. The effects of FPI on synaptic plasticity were LTP-specific, since long-term depression (LTD) could be equally induced and maintained in post-FPI, sham-operated and control slices. Sham-operated slices were characterized by synaptic excitability indistinguishable from naïve controls, but displayed decreased ability for LTP production and expressed high levels of iNOS. We conclude that FPI causes a selective loss of LTP, possibly due to a previous potentiation induced by trauma as reflected by the increased expression of synaptic proteins. Sham surgical procedures were, however, not without effects on long-term potentiation itself; the latter effects appear to be mediated by an increased production of NO. Our study demonstrates for the first time that hippocampal slices can be used to investigate the correlates of in vivo FPI. Furthermore, we describe LTP-specific deficits in post-traumatic brain injury, suggesting that FPI can selectively erase one of the two main NMDA-dependent forms of synaptic plasticity in the hippocampus.

Animals↗
PubMed1985

Development of the monosynaptic stretch reflex in the rat: an in vitro study.

The properties and development of the stretch reflex pathway were investigated in new-born and fetal rats using the isolated spinal cord-hind limb preparation. Muscle afferent discharge was elicited by small stretch of the triceps surae muscle in the new-born rat and in the fetus. It appeared as early as embryonic day 18.5. Ramp-and-hold stretch elicited only phasic discharges in most afferent fibres. A phasic reflex response was evoked in the triceps surae muscle by brief or ramp-and-hold stretch of the muscle in the new-born rat. The threshold stretch required for evoking the reflex response was close to that for eliciting the afferent discharge. A reflex response in the triceps surae muscle was also evoked by electrical stimulation of the triceps surae muscle nerve or the sciatic nerve in the new-born rat. Excitatory post-synaptic potentials (e.p.s.p.s) in the triceps surae motoneurones were evoked by stimulation of the muscle nerve in the new-born rat. The amplitude of the e.p.s.p.s was large enough to generate spike potentials. Homonymous e.p.s.p.s were significantly larger than heteronymous e.p.s.p.s. The amplitudes of the e.p.s.p.s were very susceptible to the rate of stimulus repetition. At a stimulus frequency of 10 Hz they were depressed to less than 10% of the control value. Presynaptic impulses evoked by stimulation of afferents in the muscle nerve appear in the motor nucleus less than 1.0 ms before the onset of synaptically evoked field potentials. The interval between the arrival of impulses evoked by dorsal root stimulation and the onset of e.p.s.p.s in motoneurones was 0.56 +/- 0.16 ms, indicating monosynaptic transmission from the primary afferents to the motoneurones. In the fetus, a reflex response in the triceps surae muscle was observed following a small stretch of the muscle (or electrical stimulation of the sciatic nerve) in all preparations at embryonic day 20.5 and in about half of those examined at embryonic day 19.5. Neither stimulation evoked a reflex response at embryonic day 18.5. Latencies of the reflex responses evoked by muscle stretch or by nerve stimulation were similar to those in the new-born rat. It is concluded that the monosynaptically evoked stretch reflex response in the triceps surae muscle first appear at embryonic day 19.5. Natural and electrical stimulation of the plantar skin evoked a reflex response with long latencies in flexor muscles. Such a cutaneous reflex was first present at embryonic day 17.5, two days earlier than the onset of the stretch reflex.

Action Potentials↗
PubMed1984

The contribution of in vitro studies to the understanding of epilepsy.

The slice technique has turned out to be a valuable tool in the study of fundamental epileptic processes. Intracellular recording from brain tissue in vitro may be maintained for hours. The effect of changes in the perfusion medium may be studied easily without interference from the blood-brain barrier and tissue from different brain regions may be studied in isolation. Generally, brain cells in slices behave as brain cells in vivo in spite of the obvious possibilities for distorted metabolism. The cellular basis of the epileptic process is believed to be large depolarizing potentials. Although similar potentials recorded from cells in slices have been shown to be synaptic potentials, abnormal membrane properties are also induced by epileptogenic substances. The spread of the epileptic process may occur by direct electrical coupling via gap junctions, by local increases in extracellular potassium concentration or by synaptic transmission. Reduced GABA'ergic inhibition, account, at least in part, for epileptic discharges induced by epileptogenic drugs. The role for this mechanism in naturally occurring epilepsy is unknown, but it is probably involved somehow. GABA increases the conductance of the neuronal membrane to Cl. In hippocampal dendrites, it probably in addition increases the conductance to Na because here the induced inhibitory response is depolarizing. The somatic and dendritic membranes are also different in their excitatory responses. In the hippocampus, calcium spikes predominate in the dendrites whereas steady discharges consisting of sodium spikes are characteristic of the soma. The same receptor complex seems to be involved in dendrites and soma in the enhancing effect of benzodiazepines on GABA responses. However, GABA-benzodiazepine receptor complexes in other parts of the brain are probably constituted in different ways. We know of many disturbances in the brain which may be involved in epilepsy. A single critical deviation from normal physiology in naturally occurring epilepsy has not been identified. Possibly, the process may be initiated by any one of several factors, which all take part when the disease is established.

Animals↗
PubMed1982

Synaptic excitation of the second and third order auditory neurons in the avian brain stem.

Synaptic potentials were examined in the second- and third-order auditory neurons of nucleus magnocellularis and nucleus laminaris in the chick. Brain stems of mature chick embryos were explanted and maintained in vitro for 4 to 8 h. Field potentials, extracellular spike potentials and intracellular potentials evoked by 8th-nerve stimulation were examined. Eighth-nerve stimulation reliability elicited four identifiable field potentials which could be attributed to: (i) the afferent volley of the 8th-nerve axons, (ii) postsynaptic responses of n. magnocellularis neurons, and (iii) ipsilaterally and, (iv) contralaterally-evoked n. laminaris postsynaptic responses. Intracellular-recorded postsynaptic potentials were characterized by a rapid rise time and short duration. They were apparently monosynaptic with a synaptic delay of 0.4 ms. In each n. magnocellularis neuron the 'fast' excitatory postsynaptic potentials were composed of 1 to 3 all-or-none components. 'Slow' excitatory postsynaptic potentials were characterized by a longer latency, a longer duration and graded amplitude variation in proportion to the intensity of 8th-nerve stimulation. Both 'fast' and 'slow' excitatory postsynaptic potentials had similar reversal potentials. Since the 8th nerve makes monosynaptic connection with n. magnocellularis neurons, it is likely that at this synapse the 'fast' excitatory postsynaptic potentials were produced, while the 'slow' potential may be attributable to the convergence of many boutonal synapses of unknown origin. Intracellular injections of horseradish peroxidase into n. magnocellularis revealed that its efferents bifurcate below the nucleus and send one axon to the contralateral n. laminaris while the other axon forms a highly divergent projection to the ipsilateral laminar nucleus. The intracellular records obtained from n. laminaris are consistent with this anatomical finding in that graded excitatory postsynaptic potentials were elicited by 8th-nerve stimulation.

Animals↗
PubMed1984

[Interaction between primary segmental afferents and motor neurons in the spinal cord of the lamprey].

Direct connections between dorsal cell axons and dendrites of motoneurons and giant interneurons in the isolated spinal cord of the lamprey, are described. Single dorsal cells were stimulated through a separate intracellular microelectrode, and unitary synaptic potentials in the motoneuron were recorded. Intracellular injection of HRP into the synaptically connected cells was performed via microelectrodes with 10% HRP solution. The synaptic contacts formed by injected cells were identified under the light microscope. Only dorsal cells and motoneurons in the spinal cord were stained with HRP retrograde transportation from segmental muscles. Similarity of primary afferents and motoneurons intrasegmental connections in the spinal cord of lamprey and some lower vertebrates, is discussed.

Animals↗
PubMed1994

Active propagation of somatic action potentials into neocortical pyramidal cell dendrites.

The dendrites of neurons in the mammalian central nervous system have been considered as electrically passive structures which funnel synaptic potentials to the soma and axon initial segment, the site of action potential initiation. More recent studies, however, have shown that the dendrites of many neurons are not passive, but contain active conductances. The role of these dendritic voltage-activated channels in the initiation of action potentials in neurons is largely unknown. To assess this directly, patch-clamp recordings were made from the dendrites of neocortical pyramidal cells in brain slices. Voltage-activated sodium currents were observed in dendritic outside-out patches, while action potentials could be evoked by depolarizing current pulses or by synaptic stimulation during dendritic whole-cell recordings. To determine the site of initiation of these action potentials, simultaneous whole-cell recordings were made from the soma and the apical dendrite or axon of the same cell. These experiments showed that action potentials are initiated first in the axon and then actively propagate back into the dendritic tree.

Action Potentials↗
PubMed2006

17beta-Estradiol potentiates field excitatory postsynaptic potentials within each subfield of the hippocampus with greatest potentiation of the associational/commissural afferents of CA3.

We sought to determine the impact of 17beta-estradiol throughout the hippocampal trisynaptic pathway and to investigate the afferent fiber systems within CA1 and CA3 in detail. To achieve this objective, we utilized multielectrode arrays to simultaneously record the field excitatory postsynaptic potentials from the CA1, dentate gyrus, and CA3 of rat hippocampal slices in the presence or absence of 100 pM 17beta-estradiol. We confirmed our earlier findings in CA1, where 17beta-estradiol significantly increased field excitatory postsynaptic potentials amplitude (20%+/-3%) and slope (22%+/-7%). 17beta-Estradiol significantly potentiated the field excitatory postsynaptic potentials in dentate gyrus, amplitude (15%+/-4%) and slope (17%+/-5), and in CA3, amplitude (15%+/-4%) and slope (19%+/-5%). Using a high-density multielectrode array, we sought to determine the source of potentiation in CA1 and CA3 by determining the impact of 17beta-estradiol on the apical afferents and the basal afferents within CA1 and on the mossy fibers and the associational/commissural fibers within CA3. In CA1, 17beta-estradiol induced a modest increase in the amplitude (7%+/-2%) and slope (9%+/-3%) following apical stimulation with similar magnitude of increase following basal stimulation amplitude (10%+/-2%) and slope (12%+/-3%). In CA3, 17beta-estradiol augmented the mossy fiber amplitude (15%+/-3%) and slope (18%+/-6%) and the associational/commissural fiber amplitude (31%+/-13%) and slope (40%+/-15%). These results indicate that 17beta-estradiol potentiated synaptic transmission in each subfield of the hippocampal slice, with the greatest magnitude of potentiation at the associational/commissural fibers in CA3. 17beta-Estradiol regulation of CA3 responses provides a novel site of 17beta-estradiol action that corresponds to the density of estrogen receptors within the hippocampus. The implications of 17beta-estradiol potentiation of the field potential in each of the hippocampal subfields and in particular CA3 associational/commissural fibers for memory function and clinical assessment are discussed.

Afferent Pathways↗
PubMed1984

The synaptic nature of the paroxysmal depolarizing shift in hippocampal neurons.

It is well known that there are two types of neuronal burst discharges: endogenous and network driven. Frequently, the two types are similar in appearance. But careful analysis has shown them to be quite different mechanistically and to share few common properties. Endogenous bursts are a common feature of certain hippocampal neurons. Application of a convulsant agent to a hippocampal slice, however, results in the appearance of interictal discharges whose intracellular correlate is the paroxysmal depolarizing shift (PDS). In contrast to the normal endogenous burst, the PDS is a network-driven burst. It has been proposed that the underlying depolarization of the PDS constitutes a large, compound, excitatory synaptic potential. In support of this hypothesis, recent voltage clamp experiments have demonstrated that a large synaptic conductance underlies the PDS. In this report we review those experiments and others that have established the synaptic nature of epileptiform activity in the hippocampus.

Animals↗