Biomedical subjects
G L Collingridge
Publications and source records attributed to G L Collingridge.
Role of excitatory amino acid receptors in synaptic transmission in area CA1 of rat hippocampus.
The new antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), which blocks responses to kainate and quisqualate, has been used in conjunction with D-2-amino-5-phosphonovalerate (APV), which blocks selectively responses to N-methyl-D-aspartate (NMDA), to determine the role of excitatory amino acid receptors in synaptic transmission. An excitatory postsynaptic potential (EPSP)-inhibitory postsynaptic potential (IPSP) sequence was evoked in CA1 neurons by stimulation of the Schaffer collateral-commissural pathway in rat hippocampal slices. CNQX (10 microM) substantially reduced the EPSP without having any effect on input resistance or membrane potential. The IPSP was also reduced provided that the stimulating electrode was place approximately 1 mm from the recording electrode. The EPSP that remained in the presence of CNQX had characteristics of an NMDA receptor-mediated potential; it had a slow timecourse, summated at high frequencies, was blocked reversibly by APV, increased greatly in size in Mg2+-free medium, and showed an anomalous voltage dependence in Mg2+-containing medium. In the presence of CNQX, an APV-sensitive polysynaptic GABAergic IPSP could be evoked, indicating that NMDA receptors can mediate suprathreshold EPSPS in inhibitory interneurons. It is suggested that either NMDA or non-NMDA receptors can, under different circumstances, mediate the synaptic excitation of pyramidal neurons and inhibitory interneurons in area CA1 of the hippocampus.
Temporally distinct pre- and post-synaptic mechanisms maintain long-term potentiation.
Long-term potentiation (LTP) in the hippocampus is widely studied as the mechanisms involved in its induction and maintenance are believed to underlie fundamental properties of learning and memory in vertebrates. Most synapses that exhibit LTP use an excitatory amino-acid neurotransmitter that acts on two types of receptor, the N-methyl-D-aspartate (NMDA) and quisqualate receptors. The quisqualate receptor mediates the fast synaptic response evoked by low-frequency stimulation, whereas the NMDA receptor system is activated transiently by tetanic stimulation, leading to the induction of LTP. The events responsible for maintaining LTP once it is established are not known. We now demonstrate that the sensitivity of CA1 neurons in hippocampal slices to ionophoretically-applied quisqualate receptor ligands slowly increases following the induction of LTP. This provides direct evidence for a functional post-synaptic change and suggests that pre-synaptic mechanisms also contribute, but in a temporally distinct manner, to the maintenance of LTP.
6-Cyano-7-nitroquinoxaline-2,3-dione as an excitatory amino acid antagonist in area CA1 of rat hippocampus.
1. A quantitative pharmacological investigation of the excitatory amino acid antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) has been made in area CA1 of rat hippocampal slices bathed in 1 mM Mg2+ containing medium. 2. At a concentration of 10 microM, CNQX reversibly antagonized responses to alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), quisqualate and kainate; it produced a parallel shift in their log dose-response curves. Responses to N-methyl-D-aspartate (NMDA) were not antagonized by 10 microM CNQX (dose-ratio: 1.04 +/- 0.06, n = 3). 3. Schild plots (constructed over the range 1-100 microM) yielded the following estimated pA2 values, AMPA 5.8, quisqualate 5.9, and kainate 5.9. NMDA was antagonized by 100 microM CNQX, giving an apparent log K of 4.44 +/- 0.06. 4. The slopes (+/- s.e. mean) of the Schild plots were for AMPA 0.84 +/- 0.06, quisqualate 0.79 +/- 0.04 and kainate 0.68 +/- 0.07. These were all significantly less than unity. 5. Synaptic responses elicited by low frequency activation of the Schaffer collateral-commissural pathway were blocked completely by CNQX (10 microM) providing that a low stimulus intensity was used. With high intensity stimulation a small component remained that was blocked by the selective NMDA antagonist D-2-amino-5-phosphonovalerate (APV). 6. These results suggest that CNQX does not differentially affect the responses of CA1 neurones to AMPA, quisqualate and kainate. It does, however, depress responses to these agonists to a greater degree than it does responses to NMDA and it is a highly effective synaptic antagonist.
Excitatory amino acid receptors in the vertebrate central nervous system.
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Ketamine blocks an NMDA receptor-mediated component of synaptic transmission in rat hippocampus in a voltage-dependent manner.
We have examined the voltage dependence of the effects of ketamine on synaptic currents in hippocampal CA1 neurons in vitro under conditions where there is a large N-methyl-D-aspartate (NMDA) receptor mediated component of the response. Ketamine reduced inward currents to a greater extent than outward currents of a corresponding size. D-2-Amino-5-phosphonovalerate (APV) substantially reduced the residual outward currents recorded in ketamine, but had only a small effect on the residual inward ones. It is concluded that in this system the action of ketamine in blocking synaptically evoked NMDA receptor-mediated currents shows some voltage dependence.
CNQX blocks acidic amino acid induced depolarizations and synaptic components mediated by non-NMDA receptors in rat hippocampal slices.
6-Cyano-2,3-dihydroxy-7-nitro-quinoxaline (CNQX; FG 9065) is a new excitatory amino acid antagonist. In the spinal cord it has been reported to selectively block responses to acidic amino acids acting at receptors of the non-N-methyl-D-aspartate (non-NMDA) type. Here we report that in rat hippocampal slices bathed in Mg2+-free medium 10 microM CNQX reversibly blocks responses to alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA), quisqualate and kainate but not NMDA. The synaptic response evoked by low frequency stimulation of Schaffer collateral-commissural fibres in 1 mM Mg2+-containing medium is completely blocked by this concentration of CNQX. In contrast the synaptic response evoked in Mg2+-free medium is not fully blocked by CNQX. The CNQX-insensitive component is, however, abolished by addition of a selective NMDA antagonist. The use of CNQX has allowed for the first time selective synaptic activation of NMDA receptors in the hippocampus.
A quantitative study of the actions of excitatory amino acids and antagonists in rat hippocampal slices.
1. A quantitative pharmacological investigation of the actions of excitatory amino acids on hippocampal CA1 neurones has been made using a new slice preparation developed for grease gap recording; d.c. potential was measured across a grease barrier placed between alvear fibres and the bathing medium. 2. In Mg2+-free perfusate, N-methyl-D-aspartate (NMDA, 1-100 microM), quisqualate (1-500 microM), kainate (1-200 microM) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA, 1-100 microM) caused dose-dependent depolarizations. 3. The dose-response relationships were fitted to logistic expressions. The maximum responses to AMPA, NMDA and kainate were similar; their respective EC50 values were 5, 13 and 23 microM. Quisqualate had a smaller maximum; its EC50 value was 10 microM. The slopes of the dose-response relationships were different for the 4 agonists; the order of steepness of the slopes was NMDA greater than AMPA greater than kainate greater than quisqualate. 4. Similar amino acid-induced depolarizations were observed in slices of just the CA1 region or in whole slices bathed in tetrodotoxin. Isolated alvear fibres, however, were insensitive to the excitatory amino acids. 5. D-2-Amino-5-phosphonovalerate (APV, 50 microM) selectively and reversibly antagonized responses induced by NMDA (apparent pA2 = 5.21). 6. Kynurenic acid (1 mM) reversibly depressed responses to the three agonists tested. The dose-ratios for antagonism of AMPA, kainate and quisqualate were 6.9, 5.6 and 4.6 respectively. 7. This preparation has a different sensitivity profile to agonists from those of previously reported preparations of spinal cord, neocortex and cerebellum. The greater sensitivity to NMDA may be due to the higher density of NMDA receptors in the hippocampus. The effects of the antagonists, APV and kynurenate, are similar to those found in other brain areas.
Synaptic activation of N-methyl-D-aspartate receptors in the Schaffer collateral-commissural pathway of rat hippocampus.
1. The involvement of N-methyl-D-aspartate (NMDA) receptors in the response to single-shock (0.033 Hz) stimulation of the Schaffer collateral-commissural pathway in hippocampal slices has been investigated using current- and voltage-clamp techniques. 2. In the presence of Mg2+ (1 or 2 mM) at membrane potentials near rest, the selective NMDA antagonist D-2-amino-5-phosphonovalerate (APV) had no effect on the excitatory postsynaptic potential (EPSP) and the biphasic inhibitory postsynaptic potential (IPSP) evoked by Schaffer collateral-commissural stimulation. The recurrent IPSP evoked by antidromic stimulation of alvear fibres was also unaffected by APV. 3. The introduction of a Mg2+-free perfusate led, at high stimulus intensity, to an orthodromically evoked epileptiform discharge but little change in the recurrent IPSP. APV suppressed a large proportion of the enhanced response in Mg2+-free perfusate. 4. EPSPs and excitatory postsynaptic currents (EPSCs) evoked in Mg2+-free perfusate invariably had both APV-resistant and APV-sensitive components. Both synaptic components had similar thresholds and latencies to onset. The APV-sensitive component had a long time to peak and long duration. 5. Under current-clamp conditions in Mg2+-containing medium, an APV-sensitive component was recorded at membrane potentials of between -30 and -10 mV, but not at potentials more negative than -55 mV. 6. Under voltage-clamp, but not current-clamp, conditions in Mg2+-containing medium, a small APV-sensitive component was recorded at resting membrane potentials and increased with membrane depolarization. The difference between the current- and voltage-clamp data is attributed to the hyperpolarizing influence of conjointly activated IPSPs. 7. In the presence of Mg2+ and picrotoxin, a dual-component EPSC was recorded between -30 and +30 mV in all cells examined. The APV-resistant and APV-sensitive components had similar latencies to onset. They both had reversal potentials of between -8 and 0 mV. The APV-sensitive component had a longer latency to peak and duration than the APV-resistant component. 8. It is suggested that NMDA receptors can contribute a low-threshold and long-duration monosynaptic component of the response evoked by low-frequency stimulation of the Schaffer collateral-commissural pathway. However, under physiological conditions significant expression of this component is prevented by concurrently activated IPSPs which rapidly hyperpolarize neurones into a region where Mg2+ substantially blocks NMDA channels.
Frequency-dependent N-methyl-D-aspartate receptor-mediated synaptic transmission in rat hippocampus.
1. The effects of the N-methyl-D-aspartate (NMDA) antagonist, D-2-amino-5-phosphonovalerate (APV) were examined on synaptic responses evoked by high-frequency stimulation of the Schaffer collateral-commissural pathway, in the presence of Mg2+ (1 or 2 mM) and functional synaptic inhibition. 2. The synaptic response evoked by 100 Hz stimulation comprised fast excitatory postsynaptic potentials (EPSPs) evoked by each shock and a slow depolarization. APV reduced the size of the depolarization without depressing the fast EPSPs. 3. The mean (+/- 1 S.E.) amplitude of the APV-sensitive component (3.0 +/- 0.3 mV), evoked by 100 Hz stimulation at membrane potentials near rest, was invariably smaller than the first fast EPSP (9.8 +/- 0.7 mV). Both of these synaptic components had similar thresholds and increased in amplitude as the stimulus intensity was raised. There was a positive correlation between the amplitude of the two components (r = 0.57, P less than 0.01). 4. The amplitude of the APV-sensitive component was positively correlated (r = 0.97, P less than 0.05) with the frequency of stimulation during the trains (between 10 and 100 Hz). The threshold frequency for evoking an APV-sensitive component was approximately 10 Hz. 5. In contrast to the fast EPSPs the amplitude of the APV-sensitive component increased with depolarization, and decreased with hyperpolarization, of a neurone from its resting membrane potential. The component was no longer present in some cells which had been hyperpolarized sufficiently. 6. It is suggested that during high-frequency stimulation a neurone may become depolarized for a sufficient time to reduce the Mg2+ block of NMDA channels. This enables the NMDA receptor system to contribute transiently to the synaptic response, despite the inhibitory synaptic mechanisms which prevent its activation during single-shock stimulation. The characteristics of the NMDA receptor-mediated synaptic response may explain properties relating to the induction of long-term potentiation (LTP).
MK-801 blocks NMDA receptor-mediated synaptic transmission and long term potentiation in rat hippocampal slices.
The effect of the anticonvulsant and neuroprotective agent (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclo-hepten-5,10-imine maleate (MK-801) has been studied on synaptic events in the CA1 region of rat hippocampal slices. MK-801 blocked selectively the N-methyl-D-aspartate receptor-mediated component of synaptic transmission, which can be recorded in response to single shock stimulation of the Schaffer collateral-commissural pathway in the absence of added Mg2+ to the perfusate. MK-801 also prevented the induction of long term potentiation, which is normally produced in this pathway by high frequency stimulation in the presence of Mg2+.
Characterization of an N-methyl-D-aspartate receptor component of synaptic transmission in rat hippocampal slices.
The involvement of N-methyl-D-aspartate receptors in synaptic transmission from Schaffer collateral-commissural fibres to CA1 neurons has been investigated in rat hippocampal slices. When the perfusion medium was changed from one containing 1 mM Mg2+ to one with no added Mg2+ there was a pronounced increase in the amplitude of the population spike, the appearance of secondary population spikes and in some slices spontaneous epileptiform discharges developed. The secondary and spontaneous population spikes were abolished by the selective N-methyl-D-aspartate antagonist, D-2-amino-5-phosphonovalerate. The effects on the primary population spike depended on the strength of synaptic activation. At low intensities, the N-methyl-D-aspartate antagonist reduced or abolished this response whereas at high intensities the primary population spike was slightly increased in amplitude by this compound. Mg2+ had dose-dependent (20-500 microM) effects on synaptic responses which were identical to those of D-2-amino-5-phosphonovalerate. Increasing the Ca2+ concentration over a range of 1-3 mM also reduced or abolished secondary population spikes and, at low stimulus intensities, the primary population spike. At higher stimulus intensities, however, the primary population spike was insensitive to the Ca2+ concentration over this range. These results demonstrate the major extent to which N-methyl-D-aspartate receptors can contribute to synaptic transmission and epileptiform activity in the CA1 region of the hippocampus. They also show that an important role of Mg2+ in this region is to prevent significant activation of this receptor system during low-frequency synaptic transmission.
Effects of phencyclidine, SKF 10,047 and related psychotomimetic agents on N-methyl-D-aspartate receptor mediated synaptic responses in rat hippocampal slices.
The effects of representative drugs from three classes of psychotomimetic compounds (arylcyclohexylamines, benzomorphan opioids and dioxolanes) have been examined on synaptic transmission at an identified monosynaptic pathway in rat hippocampal slices. The compounds tested were phencyclidine (PCP) and ketamine, the racemate and isomers of SKF 10,047 (N-allylnormetazocine), and the isomers of dioxadrol (dexoxadrol and levoxadrol). In the absence of added magnesium ions (Mg) in the perfusion medium low frequency stimulation of the Schaffer collateral-commissural pathway evoked a burst of population spikes in the CA1 cell body region. The secondary components of this response could be abolished by the selective N-methyl-D-aspartate (NMDA) antagonist D-2-amino-5-phosphonovalerate (APV). PCP (1 microM) or ketamine (10 microM) selectively blocked the secondary components of the synaptic response. The effect of PCP was neither mimicked nor prevented by hexamethonium and atropine, phentolamine and propranolol, or clonidine and was therefore unlikely to involve cholinergic or adrenergic neurotransmitter systems. The sigma opiate, (+/-)-SKF 10,047 (10 microM) also abolished selectively the secondary components of the synaptic response. There was no apparent difference between the potency of the stereoisomers of this compound. The action of (+/-)-SKF 10,047 was not affected by either naloxone or haloperidol, indicating that this effect did not involve opioid receptors or the haloperidol-sensitive sigma site. Dexoxadrol (10 microM), but not levoxadrol (10 microM), also selectively blocked the secondary components of the synaptic response. It is concluded that these psychotomimetic agents can block an NMDA receptor-mediated component of synaptic transmission in the hippocampus and that this effect is mediated by a specific PCP/sigma site.
Voltage-clamp analysis of somatic gamma-aminobutyric acid responses in adult rat hippocampal CA1 neurones in vitro.
1. The response of CA1 pyramidal neurones to somatic application of gamma-aminobutyric acid (GABA) was studied in adult hippocampal slices using single-electrode voltage-clamp techniques. 2. Small ionophoretic applications of GABA produced a pure outward current at the cell resting potential when recording with potassium-acetate-filled microelectrodes. This response reversed at a membrane potential of -69 +/- 5 mV (mean +/- 1 S.D.; n = 20). In recordings made with caesium-chloride-filled electrodes the GABA response reversed at -24 +/- 12 mV (n = 9). 3. The effect of different holding potentials on the size of the GABA response was examined in the range of -100 to -40 mV in twenty neurones using potassium-acetate-filled electrodes. In every case outward rectification of the response was observed. For twelve neurones the mean ratio (+/- 1 S.D. of the mean) of the conductance increase produced by GABA at -55 mV compared to -85 mV was 1.9 +/- 0.5. 4. Step changes in holding potential resulted in shifts in chloride equilibrium potential (ECl), as determined by time-dependent changes in the size of GABA-induced currents. The new value of ECl was generally reached within a few seconds of altering the membrane potential. Shifts in ECl did not appear to affect the extent of rectification but would cause underestimates of conductance measurements unless these were 'instantaneous'. The mean ratio (+/- 1 S.D. of the mean) of the 'instantaneous' conductance increase produced by GABA at 13 mV positive to that at 13 mV negative to ECl was 1.8 +/- 0.3. 5. The outward rectification was greater than that predicted by the constant-field equation. Possible factors that might contribute towards the rectification and its physiological significance are discussed.
A selective N-methyl-D-aspartate antagonist depresses epileptiform activity in rat hippocampal slices.
The sensitivity of convulsant-induced epileptiform activity in the hippocampus to the selective N-methyl-D-aspartate (NMDA) antagonist D-2-amino-5-phosphonovalerate (D-APV) was examined using in vitro electrophysiological techniques. This compound reduced the number and size of the synaptically evoked population spikes recorded in the CA1 region in the presence of the convulsants, pentylenetetrazol, bicuculline or folate. Intracellular recordings in the presence of bicuculline showed that D-APV reduced the late component of the excitatory postsynaptic potential and the number of action potentials evoked synaptically. A mechanism is suggested to explain how NMDA receptors, which are known not to be involved in normal synaptic transmission in hippocampal slices, can contribute to epileptiform activity.
Intracellular demonstration of an N-methyl-D-aspartate receptor mediated component of synaptic transmission in the rat hippocampus.
Rat hippocampal CA1 pyramidal neurones were monosynaptically activated via stimulation of the Schaffer collateral-commissural pathway. On changing from a 1 mM Mg2+-containing to a Mg2+-free medium there was a pronounced prolongation of the intracellularly recorded excitatory postsynaptic potential. This effect was reversibly abolished by the selective N-methyl-D-aspartate (NMDA) antagonist, D-2-amino-5-phosphonovalerate (APV). We propose that Mg2+ normally prevents expression of NMDA receptor-mediated responses during low-frequency stimulation. During a period of tetanic stimulation, however, cells may depolarize sufficiently to allow a significant NMDA component of the response to be manifest. This could then initiate long-term potentiation.
Magnesium ions block an N-methyl-D-aspartate receptor-mediated component of synaptic transmission in rat hippocampus.
The involvement of N-methyl-D-aspartate (NMDA) receptors in synaptic transmission in the Schaffer collateral-commissural pathway of rat hippocampal slices has been examined in the presence and absence of Mg2+. Superfusion of slices with Mg2+ -free medium resulted in an increase in the amplitude of the population spike and the appearance of multiple population spikes. An NMDA antagonist, which had no effect on the synaptic response in the presence of Mg2+, completely prevented the appearance of multiple responses. If added to slices already bathed in Mg2+ -free medium, the antagonist eliminated the multiple responses and partly reduced the amplitude of the initial population spike. These data suggest that the extent to which NMDA receptors participate in synaptic transmission in rat hippocampus depends on the extracellular Mg2+ concentration.
Effects of folic and kainic acids on synaptic responses of hippocampal neurones.
The actions of the neurotoxic amino acids folate and kainate have been compared on ortho-and antidromic responses evoked in CA1, CA3 and the dentate gyrus of slices of rat hippocampus maintained in vitro. Both in CA1 and the dentate gyrus superfusion of these acids caused an increase in amplitude of the population spike discharging from an excitatory postsynaptic potential which either remained unaffected or was reduced. In the CA3 region kainate and folate had broadly similar actions to enhance the probability of cell firing to synaptic excitation, and also caused epileptiform discharges to occur spontaneously or in response to electrical stimulation. Spontaneous and evoked population bursts in CA3 did not persist in low calcium/high magnesium medium indicating their dependence on intact synaptic transmission; spontaneously occurring bursts in CA1 were eliminated with the latter treatment or when the axonal connections between it and CA3 were cut. Following folate superfusion the commissural-evoked response in CA3 showed large and variable shifts of the latency which were dependent on the stimulus intensity and its timing after a spontaneous population discharge. Although all of the effects of folate were reproduced by bicuculline, no evidence for a decreased recurrent inhibition in CA1 was obtained although this was observed with kainate. The finding that folate and kainate produced their effects in the absence of a detectable effect on the antidromic population spike suggests a mechanism of action other than neuronal depolarization. The implications of these data for the neurotoxic mechanism(s) and the receptor homologies of folate and kainate are discussed.