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R Dingledine

Publications and source records attributed to R Dingledine.

At least 91 records · Page 5Linked to original sources
PubMed1983

N-methyl aspartate activates voltage-dependent calcium conductance in rat hippocampal pyramidal cells.

The depolarizing actions of N-methyl-DL-aspartate (NMA) and L-glutamate on pyramidal neurones were compared in a hippocampal slice preparation. Tetrodotoxin (1 microM) was added to the perfusion solution to suppress regenerative Na conductances. Depolarization evoked by ionophoretic application of NMA triggered slow, high-threshold regenerative spikes. These are considered to be Ca spikes since the amplitude and rate of rise could be reduced by verapamil, D-600, Co2+ and Mn2+, and increased by Ba2+. Multiple Ca-spike thresholds could be demonstrated in the same cell. In contrast, depolarizations evoked by L-glutamate only rarely triggered Ca-spikes. The minimum latency to the onset of depolarization evoked by NMA was less than 20 ms. The latency and amplitude of NMA-evoked responses were highly dependent on the position of the ionophoretic pipette; movements of the pipette by as little as 10-50 micron could markedly change the size of the response. Spatially separate hot spots for NMA and glutamate were not found. Depolarizations evoked by small to moderate ionophoretic currents of NMA were usually associated with an apparent rise in input resistance, as tested by the response to transmembrane current pulses. Ionophoresis of L-glutamate, or high NMA doses, however, usually caused a fall in input resistance. Both the depolarization and the conductance change evoked by NMA were highly voltage-dependent within the approximate range -50 to -80 mV; they could be increased by modest depolarization and reduced by hyperpolarization of the membrane. No reversal potential could be demonstrated in the hyperpolarizing direction. Rather, the NMA response approached zero asymptotically at sufficiently hyperpolarized membrane potentials. Subthreshold depolarizations and conductance changes elicited by NMA could be blocked by Co2+, Mn2+ and Cd2+, and reduced by D-600 and verapamil. These Ca2+ antagonists had little or no effect on resting membrane potential or input resistance, or on responses to L-glutamate. Ba2+ increased the amplitude of subthreshold NMA responses. Intracellular injection of Cs+ plus tetraethylammonium caused cells to fire large, prolonged (up to 15 s) Ca spikes, presumably because most K+ conductances were blocked. Under these conditions the effect of NMA was unchanged or enhanced. Raising [K+]o to 10.5 mM (from the normal 3.5 mM) caused a depolarization and fall in input resistance, but did not change the amplitude or voltage dependence of the NMA response. Reducing [Na+]o caused an initial increase, then usually a delayed decrease in the amplitude of the NMA response.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗
PubMed1983

Excitatory amino acids: modes of action on hippocampal pyramidal cells.

Recent pharmacological and biochemical evidence supports the idea that acidic amino acids act as neurotransmitters at several excitatory synapses in the hippocampus. In this paper I review work comparing certain physiological actions of N-methyl-DL-aspartate (NMA) and L-glutamate in a hippocampal slice preparation. Intracellular recordings were made from pyramidal neurons bathed in 1 microM tetrodotoxin; agonists were applied by focal ionophoresis. NMA evoked calcium spikes and produced an apparent increase in the input resistance of pyramidal cells, whereas glutamate was very weak in these respects. The depolarization and conductance change caused by NMA were voltage dependent: both could be abolished by hyperpolarizing the cell to -70 to -90 mV, but no reversal potential could be demonstrated. The results of pharmacological and ionic manipulations suggest that the primary action of NMA does not involve reduction of a conventional potassium conductance. It is suggested that N-methyl-D-aspartate (NMDA) receptor activation increases a voltage-sensitive calcium conductance leading to a transient rise in cytoplasmic calcium concentration. The significance of this event is discussed with respect to the possible synaptic functions of chemically gated, voltage-sensitive calcium channels, and in particular with respect to the possible roles that NMDA receptors might serve in the genesis of long-term potentiation of excitatory synapses in the hippocampus.

Action Potentials↗
PubMed1982

Pharmacological characterization of opioid effects in the rat hippocampal slice.

The potencies of several opiates and opioid peptides for potentiating the synaptic activation of CA1 pyramidal cells were compared in the rat hippocampal slice preparation. Morphine and the opioid peptides [D-Ala2, D-Leu5]-enkephalin (DADL), beta-endorphin and Tyr-D-Ser-Gly-Phe-Leu-Thr (a delta agonist) caused a concentration-dependent shift to the left in the input-output curve constructed by plotting population spike amplitude (a measure of evoked firing) as a function of the dendritic field excitatory postsynaptic potential. The concentration-response curves for DADL and morphine had similar slopes and maxima, although the curve for morphine was biphasic due to the addition of a nonopiate effect that became apparent at higher concentrations (greater than or equal to 20 microM). The EC50 values were 68 nM for DADL and 3000 nM for morphine. The IC50 values of naloxone against equieffective concentrations of DADL and morphine were not significantly different. Perfusion of slices with a combination of nearly maximally effective concentrations of DADL and morphine resulted in an effect that was no greater than the maximum effect obtained by either drug alone. The results suggest that these opioids produce their actions through a common pathway. The rank order of potency to produce identical effects was DADL greater than Tyr-D-Ser-Gly-Phe-Leu-Thr greater than beta-endorphin greater than morphine. The kappa agonist ethylketocyclazocine was inactive at concentrations up to 10 microM. The data suggest that delta opioid receptors play a key role in the epileptiform action of these opiates in the CA1 region of the rat hippocampus. However, this opioid response may be different from those characterized in peripheral preparations because ethylketocyclazocine appears to be inactive in the hippocampal CA1 region.

Animals↗
PubMed1981

The excitatory action of acetylcholine on hippocampal neurones of the guinea pig and rat maintained in vitro.

In preliminary experiments on 39 identified pyramidal cells in the in vitro slice preparation of the guinea-pig hippocampus the depolarization evoked by acetylcholine (ACh) applied by microiontophoresis was always associated with an increase in membrane resistance. In 9 slices cut from the rat hippocampus similar results were obtained from 24 cells. In a more detailed analysis on 13 cells from the rat hippocampus, whose mean resting potential was -74 mV and mean resting input resistance 33 M omega, the mean peak depolarization evoked by ACh was 11.6 mV and the mean increase in membrane resistance 12 M omega. The reversal potential for the excitatory action of ACh was 29 mV more hyperpolarizing than the resting membrane potential. The depolarization evoked by ACh was linearly related to the corresponding increase in membrane resistance expressed as a fraction of the resting membrane resistance determined before and after the application of ACh. This was true throughout each of the individual applications of ACh and of the peak response evoked by each of the 13 applications. The constancy of this relationship is compatible with the usual model used to describe synaptic events thought to be mediated by the closure of ionic channels which are open in the absence of the transmitter. The onset of the response to ACh was always approximately 4 times slower than that evoked by a near equipotent microiontophoretic application of glutamate from an adjacent barrel of the same multibarrelled micropipette. Following the application of ACh, recovery was also slow and, on average, was approximately 10 times longer than that following a near equipotent application of glutamate. It is suggested that the slow onset and offset of the responses evoked by ACh are not compatible with models based on diffusion and are best explained by postulating a sequential generation of one or more intermediates.

Acetylcholine↗
PubMed1981

Calcium dependence of synaptic transmission in the hippocampal slice.

'Population' afferent spike and 'population' EPSP were recorded with extracellular microelectrodes in slices of hippocampal tissue maintained in vitro. Calcium concentration was changed in the bathing solution, and calcium activity ([Ca2+]0) was measured in interstitial fluid of the slice with ion-selective microelectrodes. Synaptic transfer was a non-linear continuous function of [Ca2+]0. Deviation of [Ca2+]0 by 0.1 mM from the 1.2 mM control level caused a change of approximately 15% in the slope of the input-output function.

Animals↗
PubMed1981

Presynaptic inhibitory effect of acetylcholine in the hippocampus.

(1) In order to investigate the effects of acetylcholine (ACh) on synaptic transmission in the rat hippocampus, extracellular and intracellular recordings were made from pyramidal neurons in an in vitro slice preparation while synaptic inputs to the cell population were stimulated. ACh was applied ionophoretically into somatic and dendritic layers of the slice. (2) ACh applied into the apical dendritic layer of the CA1 region reduced the size of the locally evoked field excitatory postsynaptic potential (EPSP) without altering the size of the afferent fiber volley. Likewise, dendritically applied ACh reduced the size of intracellularly recorded EPSPs. This effect of ACh appeared to be muscarinic since it was not affected by hexamethonium (up to 3 X 10-5 M) but was antagonized by atropine in a dose-dependent manner. (3) The distribution of Ach-sensitive sites matched closely the spatial distribution of activated synapses on the pyramidal cell dendrites as shown by ionophoretic mapping experiments. (4) In contrast to the effects of dendritic applications of ACh, ionophoresis of ACh into the cell layer resulted in an increase and prolongation of EPSPs and a transient decrease in the size of recurrent somatic inhibitory postsynaptic potentials (IPSPs). These effects on synaptic potentials could not be explained by the observed changes in membrane potential and input resistance following somatic application of ACh. (5) Short dendritic applications of ACh had no consistent effect on the membrane potential or slope conductance of pyramidal neurons and did not attenuate the depolarization evoked by brief dendritic applications of glutamate. In addition, the time course of ACh-reduced EPSPs was not different from control. (6) We conclude that ACh exerts a presynaptic inhibitory effect on both excitatory and inhibitory afferents to hippocampal pyramidal neurons. This effect of ACh is widespread, occurring in all regions of Ammon's horn tested as well as in stratum moleculare of fascia dentata.

Acetylcholine↗
PubMed1981

Possible mechanisms of enkephalin action on hippocampal CA1 pyramidal neurons.

(1) Intracellular and extracellular recordings were made from CA1 pyramidal neurons in an in vitro rat hippocampal slice preparation, while [D-Ala2, D-Leu5]enkephalin (DADL) was applied by perfusion at a known concentration (1 to 5 X 10-7 M), in a small droplet, or by iontophoresis into the cellular and dendritic layers of the slice. The effects of DADL on synaptic potentials and membrane properties were studied in an effort to determine the mechanisms underlying its epileptogenic action in the hippocampus. (2) DADL increased the size and often the duration of excitatory postsynaptic potentials (EPSPs) generated on either the apical or basal dendrites; this resulted in an increased discharge probability for a constant orthodromic stimulus. Extracellular field potential recordings showed a larger population spike for a given size field EPSP. These effects of DADL could be reversed substantially by perfusion with naloxone (1 to 5 X 10-7 M) and appeared qualitatively different from the epileptiform actions of penicillin. (3) DADL did not appear to increase the intrinsic excitability of the soma membrane, since membrane potential, input resistance, spike threshold, and antidromic field potentials all were unchanged. In addition, the shape of the membrane charging curve during hyperpolarizing current injection was not changed noticeably by DADL. (4) At the concentrations tested, DADL did not attenuate recurrent inhibition in the CA1 region, as evaluated by comparing in the absence and presence of DADL: (a) antidromically evoked recurrent inhibitory postsynaptic potentials (IPSPs) and their dependence of membrane potential, (b) the reduction of a synaptically driven population spike by a prior antidromic volley, (c) iontophoretic GABA (gamma-aminobutyric acid) responses. Similarly, IPSPs evoked by orthodromic stimulation appeared either unaffected or occasionally enhanced by DADL. (5) By iontophoretic mapping, it was shown that the DADL-sensitive sites are limited to stratum oriens and stratum pyramidale. Local application of DADL into stratum radiatum was relatively ineffective in enhancing the efficacy of synapses located in this region. (6) The dendritic input-output relationship between the presynaptic fiber volley and the field EPSP was not changed by DADL. This finding and the results of the iontophoretic mapping experiments suggest that increased excitatory transmitter release was not involved. (7) The data are consistent with the proposal that DADL selectively attenuates a dendritic IPSP which is virtually invisible to the soma, although the possibility cannot be ruled out that DADL may, in addition, act to enhance the responsiveness of pyramidal dendritic membrane to excitatory synaptic activation.

Animals↗
PubMed1980

Conductance changes and inhibitory actions of hippocampal recurrent IPSPs.

Intracellular recordings were obtained from CA1 pyramidal neurons in obliquely cut in vitro hippocampal slices. Recurrent IPSPs were elicited by antidromic stimulation of alvear fibers. The mechanisms by which IPSPs depress pyramidal cell excitability were investigated. Recurrent IPSPs could be reversed in sign by small hyperpolarizing currents applied through the recording electrode, indicating an increased membrane conductance. By using an AC bridge circuit it was found that the maximum impedance decrease usually occurred slightly before the peak of the IPSP. Otherwise the time course of the impedance change matched that of the IPSP itself. Inhibitory actions of the conductance increase were studied by adjusting the membrane potential to the IPSP equilibrium potential, thus allowing only the IPSP conductance to play an inhibitory role. Under these conditions non-linear summation of recurrent IPSPs with EPSPs originating in the apical dendrites could be demonstrated only during the initial 15--25 msec ofthe IPSP, which is the period of maximum conductance increase. The inhibition afforded by the hyperpolarization of the recurrent IPSP far outlasts the period of effective EPSP shunting by the inhibitory synaptic currents. The mechanisms of recurrent inhibition in the hippocampus thus appear similar to those operating in spinal motoneuron IPSPs.

Animals↗
PubMed1980

Two different responses of hippocampal pyramidal cells to application of gamma-amino butyric acid.

1. Extra- and intracellular recordings were made from CA1 cells in hippocampal slices in vitro. The effects of ionophoretically applied GABA on somatic and dendritic regions were studied. 2. Ionophoresis of GABA at dendritic sites gave a reciprocal effect by inhibiting the effect of excitatory synapses close to the dendritic application, while facilitating those lying further away. For example, GABA delivered to the mid-radiatum dendritic region reduced the population spike generated by a radiatum volley, while facilitating the population spike evoked by oriens fibre stimulation. Similarly, when single cells were recorded from, mid-apical dendritic delivery of GABA abolished the synaptically driven discharges evoked by fibres terminating at this part of the dendritic tree, but facilitated the responses to input from fibres terminating on the basal dendrites of the same cell. 3. With intracellular recording two effects were observed. Applied near the soma, GABA induced a hyperpolarization associated with an increased membrane conductance. When applied to dendrites, GABA caused a depolarization also associated with an increased membrane conductance. Both types of GABA applications could inhibit cell discharges, although in some cases the depolarizing response could facilitate other excitatory influences or cause cell firing by itself. 4. Both the hyperpolarizing and depolarizing GABA responses persisted after blockade of synaptic transmission by applying a low calcium high magnesium solution, indicating mediation via a direct effect upon the cell membrane. 5. The reversal potential for the hyperpolarizing GABA effect was similar to the equilibrium potential for the i.p.s.p. evoked from alveus or orthodromically, and was 10-12 mV more negative than the resting potential. The size of the depolarizing response was also dependent upon the membrane potential. By extrapolation an estimated equilibrium potential was calculated as about -40 mV. 6. Our results support the idea that the hyperpolarizing basket cell inhibition at the soma is mediated by the release of GABA. This hyperpolarizing response causes a general inhibition of firing. The dendritic effects of GABA, however, seem to represent another type of inhibition, which by shunting synaptic currents makes possible a selective inhibitory influence on afferents synapsing locally while facilitating more remotely placed excitatory synapses. We propose the term discriminative inhibition for this postulated new type of control of pyramidal cell discharges.

Animals↗
PubMed1980

Reduced inhibition during epileptiform activity in the in vitro hippocampal slice.

1. Intracellular recordings were made from CA1 pyramidal cells in the hippocampal slice in vitro. The responses to orthodromic and antidromic activation and to ionophoretically applied GABA were studied. 2. The epileptogenic agent sodium benzyl penicillin reduced the recurrent i.p.s.p. evoked by subthreshold antidromic stimulation. Reversal potential studies of the i.p.s.p. and resistance measurements showed that this reduction was mainly due to a decrease in i.p.s.p. conductance. 3. Penicillin also reduced the conductance and associated membrane potential changes induced by ejecting GABA near the soma or into the apical dendritic region. 4. The mixed e.p.s.p.-i.p.s.p. evoked by orthodromic stimulation was converted to a pure depolarizing potential as the i.p.s.p. was blocked. Concurrently the probability of discharge to a constant orthodromic stimulus was increased. Similar changes were seen in a low chloride solution. 5. The time course of the reduction of inhibition was similar to that of the enhanced orthodromic response seen after penicillin treatment. 6. We conclude that reduction of postsynaptic inhibition is partly responsible for the increased probability of orthodromic discharge caused by penicillin. The longer latency all-or-nothing burst seen in some cells, however, seems to require an additional mechanism, although reduced inhibition may facilitate the triggering of this burst.

Animals↗
PubMed1978

Naloxone as a GABA antagonist: evidence from iontophoretic, receptor binding and convulsant studies.

From the following three lines of evidence, it is proposed that at least part of the convulsant activity of naloxone is a result of GABA receptor blockade. Firstly, iontophoretic naloxone reversibly antagonized GABA-evoked depression of firing rate in 21 of 27 neurons tested in the rat olfactory tubercle-nucleus accumbens region, without blocking inhibition evoked in the same cells by glycine (15 cells) or morphine (6 cells). Secondly, i.p. naloxone in high doses caused convulsions in mice, and potentiated the convulsant activity of bicuculline, but not that of strychnine. Diazepam, which protected mice against convulsions elicited by bicuculline, but not by strychnine, also protected mice against naloxone. Thirdly, naloxone, morphine, levorphanol and its non-analgesic enantiomer dextrorphan displaced 3H-GABA from GABA receptor sites in homogenates of human cerebellum, all with comparable low potencies (IC50 = 250--400 micron). There was no correlation with affinities at the stereospecific receptor sites that mediate opiate-induced analgesia, since the potent opiates etorphine and diprenorphine were relatively inactive (IC50 greater than 3 mM). In addition naloxone displaced 3H-GABA from receptor sites in rate forebrain and cerebellum, with similar low potency.

Action Potentials↗
PubMed1977

Distribution of endorphins (peptides with morphine-like pharmacological activity) in pituitary.

Endorphin was found in the pituitary gland of rat, sheep, pig, cow and man. In bovine pituitary, highest concentrations have been found in that part of the posterior lobe containing the pars intermedia, although endorphin was also present in the pars distalis. Analysis by gel filtration and adsorption chromatography suggested that the major endorphins from rat, pig and cow pituitary were similar, with apparent molecular weights in the range 3000-3500 daltons.

Animals↗
PubMed1977

Brain stem stimulation and the acetylcholine-evoked inhibition of neurones in the feline nucleus reticularis thalami.

1. In cats anaesthetized with halothane and nitrous oxide, the responses to iontophoretically applied acetylcholine (ACh) and to high-frequency stimulation of the mid-brain reticular formation (MRF) were tested on spontaneously active neurones in the nucleus reticularis thalami and underlying ventrobasal complex.2. The initial response to MRF stimulation of 90% of the ACh-inhibited neurones found in the region of the dorsolateral nucleus reticularis was an inhibition. Conversely, the initial response of 82% of the ACh-excited neurones in the ventrobasal complex was an excitation. Neurones in the rostral pole of the nucleus reticularis were inhibited by both ACh and RMF stimulation.3. The mean latency (and s.e. of mean) for the MRF-evoked inhibition was 13.7 +/- 3.2 ms (n = 42) and that for the MRF-evoked excitation, 44.1 +/- 4.2 ms (n = 35).4. The ACh-evoked inhibitions were blocked by iontophoretic atropine, in doses that did not block amino acid-evoked inhibition. In twenty-four ACh-inhibited neurones the effect of iontophoretic atropine was tested on MRF-evoked inhibition. In all twenty-four neurones atropine had no effect on the early phase of MRF-evoked inhibition but weakly antagonized the late phase of inhibition in nine of fourteen neurones.5. Interspike-interval histograms showed that the firing pattern of neurones in the nucleus reticularis was characterized by periods of prolonged, high-frequency bursting. Both the ACh-evoked inhibitions and the late phase of MRF-evoked inhibitions were accompanied by an increased burst activity. In contrast, iontophoretic atropine tended to suppress burst activity.6. The possibility is discussed that electrical stimulation of the MRF activates an inhibitory cholinergic projection to the nucleus reticularis. Since neurones of the nucleus reticularis have been shown to inhibit thalamic relay cells, activation of this inhibitory pathway may play a role in MRF-evoked facilitation of thalamo-cortical relay transmission and the associated electrocortical desynchronization.

Acetylcholine↗