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Excitatory amino acid antagonists depress transmission in hippocampal projections to the lateral septum
Brain Research, 382 (1986) 437-440 Elsevier
437
BRE 21781
Excitatory amino acid antagonists depress transmission in hippocampal projections to the lateral septum DAVID R. STEVENS and CARL W. COTMAN Department of Psychobiology, University of California, lrvine, CA 92717 (U.S.A.) (Accepted May 27th, 1986) Key words: excitatory amino acid - - lateral septum - - kynurenic acid --p-bromobenzoyl-piperazine-2,3-dicarboxylic acid - intracellular recording - - synaptic transmission - - fimbria
Antagonists of excitatory amino acid neurotransmission were tested as antagonists of septal excitatory postsynaptic potentials (EPSPs) generated by stimulation of the fimbria. Septal EPSPs were depressed in a concentration-dependent manner by kynurenic acid and P-bromobenzoyl piperazine-2,3-dicarboxylic acid but not by L-2-amino-4-phosphonobutyric acid or D-2-amino-5-phosphonopentanoic acid. These results indicate that the hippocampal projection to the lateral septum is similar to the Schaffer collateral system and is mediated by an excitatory amino acid or a similar derivative. One of the outputs of the hippocampus terminates in the dorsolateral septal area 17'22. The majority of these fibers originate in hippocampal area CA3, enter the septum via the fimbria, and are branches of the Schaffer collateral pathway x'12'2e. Since transmission in the Schaffer collateral system is thought to be mediated by an excitatory amino acid 2'15'19, it has been proposed that the hippocampal projection to the septum is also mediated by an excitatory amino acid 21. The studies described below indicate the presence, release, and uptake of glutamate, and the presence of binding sites for excitatory amino acids associated with the hippocampal projection to the lateral septum. The levels of glutamate decrease following kainic acid lesions of the hippocampus or knife cuts of the fimbria 5, and glutamate uptake decreases in the lateral septum following fimbria section 2° or kainic acid lesions of the hippocampus 23. Malthe-S0renssen and co-workers 13 have demonstrated that [3H]D-aspartate is released in a calciumdependent manner in the lateral septum following stimulation of the fimbria/fornix. These authors interpreted this result as indicating that an excitatory amino acid (probably glutamate) is the excitatory neurotransmitter in this pathway. The lateral septal
area has been shown to contain relatively high numbers of binding sites for N-methyl-D-aspartate ( N M D A ) and quisqualic acid 14 that probably represent postsynaptic receptors for glutamate, aspartate, or a related derivative. To date however, there is little evidence addressi n g the pharmacological characterization of this pathway. Zhadina and co-workers 24 have reported that firing rates of septal neurons are increased in the presence of glutamate. Also, JoEls and Urban 9 have reported that glutamate increases firing rates of septal neurons and that glutamate-diethyl-ester, ( G D E E ) but not 2-amino-5-phosphonopentanoate (AP5), applied iontophoretically, depresses potentials elicited in the septum by stimulation of the fimbria. It was concluded that an excitatory amino acid acting as a quisqualate receptor may mediate transmission of this pathway. However, the reported specificity of G D E E has not been seen in studies of the hippocampus 3. In this paper, we have used intracellular recording techniques to examine the actions of known concentrations of excitatory amino acid antagonists which have been characterized in the hippocampus, on septal excitatory postsynaptic potentials (EPSPs) resulting from stimulation of the fimbria/ fornix. The results of these experiments are com-
Correspondence: D.R. Stevens, Department of Psychobiology, University of California, Irvine, CA 92717, U.S.A. 0006-8993/86/$03.50 © 1986 Elsevier Science Publishers B.V. (Biomedical Division)
438 pared to the results of similar experiments carried out in the hippocampal slice preparation. Male Sprague-Dawley albino rats weighing 160-220 g were decapitated, the brains were rapidly removed, and slices prepared using a Vibratome. Details of the dissection have been reported previously 18. The slice was then transferred to a recording chamber for intracellular recording, lntracellular recordings were carried out using glass microelectrodes filled with 4 M K-acetate and having resistances of 80-160 Mr2. Cell input resistance was monitored continually using current injection via the bridge circuit of the intracellular amplifier (Biodyne model
A2) The recording chamber was maintained at 33 + 1 °C and the slice was constantly superfused with an artificial cerebrospinal fluid (ACSF) of the following composition (in mM): NaC1 117, KCI 2.5, CaC12 2.0 K2HPO 4 1.2, MgSO42.0, NaH2CO3 25, and D-glucose 10. The medium was bubbled with 95% 02/5% CO2 to maintain pH. The fimbria/fornix was stimulated using bipolar tungsten electrodes. Antagonists (kynurenic acid, bicuculline methiodide, L-2-amino-4-phosphonobutyric acid (AP4), Sigma; p-bromobenzoyl piperazine2,3-dicarboxylic acid (pBBPzDA) and D-amino-5phosphonovaleric acid (AP5), Tocris, England, were applied as part of the normal superfusate. Control EPSPs resulting from single stimuli (mean of 10 stimuli) were compared to the means of EPSPs recorded in the presence of antagonist (or other treatment). Following treatments, the response was followed for observation of recovery after washout of the antagonist. Stimulation of the fimbria/fornix produces short latency (2-3 ms) graded EPSPs which reach action potential threshold with increasing stimulus intensity (Fig. 1A). Using short current pulses of both polarities, membrane resistance was monitored during the EPSP, demonstrating that the EPSP is associated with an increase in conductance (Fig. 1B). Superfusion of the slice with medium containing 0.25 mM Ca 2÷ and 3.45 mM Mg 2÷ results in a depression (80%) of the EPSP amplitude (Fig. 1C). EPSPs typically rise rapidly and decline with a half-time near 20 ms. The time course of the EPSP is highly variable with durations as long as 50 ms. With increasing stimulus intensity the duration of the
A
B
l!
....... low c a l c i u m
kynurenate
5 0 0 uM
bicucullJne 10 uM
Fig. 1. lntracellular recordings from lateral septal neurons. A: septal EPSPs and an orthodromic action potential resulting from stimulation of the fimbria/fornix. B: a representative oscilloscope tracing shows that tonic potentials resulting from intracellular injection of current of either polarity are reduced during the EPSP indicating that membrane conductance is increased. C: treatment of the septal slice with low calcium-high magnesium acsf results in depression of the EPSP amplitude (78 + 5%, mean + S.D., 't' indicates the treatment response). D: kynurenic acid (500ktM) treatment results in a depression of EPSP amplitude and prolongation of the EPSP (t, treatment response). E: treatment of the septal slice with bicuculline methiodide (10/~M) results in a prolongation of the EPSP (t, 3-min superfusion, 't', 6-min superfusion).
EPSP decreases. Treatment of the slice with the excitatory amino acid antagonist kynurenic acid 16 results in depression of EPSP amplitude and an increase in the duration of the EPSP (Fig. 1D). Under similar stimulus conditions, treatment of the slice with the 7aminobutyric acid (GABA) antagonist bicuculline methiodide (10 #M) results in prolongation of the time course of the EPSP (Fig. 1E). This result indicates that the time course of the EPSP is decreased by a GABA-mediated inhibitory postsynaptic potential. The sensitivity of septal EPSPs to antagonists of excitatory amino acid transmission was tested in order to demonstrate that this pathway was sensitive to amino acid antagonists, and to compare their effectiveness in this region to that in the hippocampus. The results of treatment with a series of excitatory amino acid antagonists is shown in Fig. 2. Septal slices were treated with kynurenic acid, a recently described antagonist at hippocampal excitatory amino acid receptors 6. Kynurenic acid depressed the amplitude of the septal EPSP at a concentration of 100-500/~mol, with no noticeable effects on membrane potential or input resistance (Fig. 2A). pBBPzDA has been shown to be an effective antagonist of excitatory amino acid responses in the hippocampus 7. Treatment of the septal preparation with this agent also resulted in depression of the septal EPSP (Fig. 2A). pBBPzDA was slightly more potent
439 A
CONTROL
"IRF_ATIh~NIT
B LOG-CONCENTRATION/RESPONSE RELATION
~
loo
• •
KYN pBBPzDA
KYN200 ulM
s
~
~mSec
t
o lOO
~ooo CONCENTRATION (uM~
Fig. 2. The actions of selected excitatory amino acid antagonists on septal EPSPs. A: pairs of records demonstrating representative EPSP's during control recordings and the effect of treatment of the same neuron with the indicated agent. Kynurenic acid and pBBPzDA depressed septal EPSPs while AP4 and AP5 had no effect on septal EPSP's. B: log concentrationresponse relations for Kynurenic acid and pBBPzDA, both of which depressed EPSPs in a concentration-dependent manner with IC50values near 200pM. The bars on the plotting symbols represent one standard deviation. The two agents exhibited parallel concentration-response curves and similar potencies.
than kynurenate (Fig. 2B). AP4 (10-20 pM), an agent which acts presynaptically to depress transmission in mossy fiber terminals in the guinea pig hippocampus s, was tested and found to have no effect on membrane properties or on septal EPSP amplitude (Fig. 2A). The specific NMDA antagonist, AP5 (ref. 4), was tested on the septal EPSP also, and had no effect on membrane properties or on the EPSP at concentrations of 5 50pM. These results demonstrate that the potential generated by stimulation of fimbria/fornix is graded, calcium-dependent, and associated with a conductance increase. Our observations are in accord with the proposed synaptic nature of this response. The increase in the duration of the EPSP following depression by kynurenic acid indicates that the EPSP decay is not passive and may be altered by the presence of an inhibitory synaptic component. This result suggests the presence of a feed-forward inhibitory circuit similar to that reported in the hippocampus I. The increase in the duration of the EPSP following treatment with bicuculline methiodide, a G A B A antagonist, supports this suggestion. Bicuculline methiodide treatment increased the duration of the EPSP at low stimulus intensities well below the threshold for action potentials. These results are readily explained
by the orthodromic activation of GABAergic interneurons by fimbrial stimulation. We cannot rule out the activation of some other pathway or recurrent inhibition following the activation of other septal principal neurons, although the short latency of the bicuculline methiodide sensitive component of the synaptic response does not support the latter alternative. The actions of kynurenic acid and p B B P z D A as antagonists of the EPSP support the proposal that septal EPSPs are mediated by excitatory amino acid receptors, p B B P z D A is slightly more potent than is kynurenic acid. Both agents have IC50 values near 200 /~M, consistent with those reported for these agents on commissural/associational synapses of the hippocampus 6,7. The lack of effect of AP4 is not inconsistent with the proposal that either glutamate or aspartate is the transmitter in this pathway since AP4 appears to act on a subset of excitatory amino acid receptors 8:°. AP4 is also ineffective at the Schaffer collateral synapse 2. The failure of D-AP5 (50/~M) to antagonize the septal EPSP indicates that in this pathway, as in the Schaffer collateral projection, NMDA receptors do not normally contribute to the synaptic response II. In physiological experiments the apparent dissociation constant of AP5 for the N M D A binding site is near 5 /~M H. While NMDA receptors have been reported to be present in this area 14, their role remains unclear. The possibility that these sites play a role in synaptic plasticity needs to be examined 2:~. Our results support the proposal that glutamate or a related compound acts as the neurotransmitter of the hippocampal projection to the septal area and that the postsynaptic response is mediated by an excitatory amino acid receptor. They also indicate a high degree of similarity between the hippocampal projection to the septum and that of the Schaffer collateral pathway. This similarity is not unexpected since these two projections arise from the same neurons 22. The potencies of kynurenic acid and pBBPzDA on the septal EPSP are similar to their potencies against kainic acid responses recorded from hippocampal pyramidal cells of area C A I and on Schaffer collateral/ commissural EPSPs. These results support the suggestion that the hippocampal projection to the lateral septum is mediated by an excitatory amino acid acting at a kainic acid binding site. However, a role for
440 q u i s q u a l i c acid b i n d i n g sites c a n n o t b e r u l e d o u t w i t h o u t f u r t h e r e x a m i n a t i o n o f t h e a c t i o n s of e x o g e -
n o u s l y a p p l i e d a g o n i s t s a n d t h e i r i n t e r a c t i o n w i t h antagonists.
1 Andersen, P., Blackstad, T.W. and L0mo, T., Location and identification of excitatory synapses on hippocampal pyramidal cells, Exp. BrainRes., 1 (1966) 236-248. 2 Collingridge, G.L., Kehl, S.J. and McLennan, H., Excitatory amino acids in synaptic transmission in the Schaffer collateral-commissural pathway in the rat hippocampus, J. Physiol. (London), 334 (1983) 33-46. 3 Collingridge, G.L., Kehl, S.J. and McLennan, H., The antagonism of amino acid induced excitations of rat hippocampal CA t neurones in vitro, J. Physiol. (London), 334 (1983) 19-31. 4 Davies, J., Francis, A.A., Jones, A.W. and Watkins, J.C., 2-amino-5-phosphonovalerate (2APV), a potent and selective antagonist of amino acid induced and synaptic excitation, Neurosci. Lett., 21 (1981) 77-81. 5 Fonnum, F. and Walaas, I., The Effect of intrahippocampal kainic acid injections and surgical lesions on neurotransmitters in hippocampus and septum, J. Neurochem., 31 (1978) 1173-1187. 6 Ganong, A.H., Lanthorn, T.H. and Cotman, C.W., Kynurenic acid inhibits synaptic and acidic amino acid-induced responses in the rat hippocampus and spinal cord, Brain Research, 273 (1983) 170-174. 7 Ganong, A.H., Jones, A.W., Watkins, J.C. and Cotman, C.W., Parallel antagonism of synaptic transmission and Kainate/quisqualate responses in the hippocampus by piperazine-2,3-dicarboxylic acid analogs J. Neurosci., in press. 8 Ganong, A.H., Perkins, M.N., Flatman, J.N. and Cotman, C.W., Differential effect of excitatory amino acid antagonists on evoked and spontaneous excitatory potentials in guinea pig hippocampus, J. Physiol. (London), in press. 9 JoEIs, M. and Urban, I.L.A., Electrophysiological and pharmacological evidence in favor of amino acid neurotransmission in the fimbria-fornix fibers innervating the lateral septal complex of rats. Exp. Brain Res., 54 (1984) 455-462. 10 Koerner, J.F. and Cotman, C.W., Micromolar L-2-amino4-phosphonobutyric acid selectively inhibits perforant path synapses from lateral entorhinal cortex, Brain Research, 216(1981) 192-198. 11 Harris, E.H., Ganong, A.H. and Cotman, C.W., Longterm potentiation in the hippocampus involves activation of N-methyl-D-aspartate receptors, Brain Research 323 (1984) 132-137. 12 Lorente de No', R., Studies on the structure of the cerebral
cortex. II. Continuation of the study of the ammonic system, J. Psychol. Neurol., 46 (1934) 113-177. 13 Malthe-Sorenssen, D., Skrede, K.K. and Fonnum, F., Release of D-[H3]-aspartate from the dorsolateral septum after electrical stimulation of the fimbria in vitro, Neuroscience 5 (1980) 127-133. 14 Monaghan, D.T. and Cotman, C.W., Anatomical organization of NMDA, Kainate, and Quisqualate receptors. In E.J., Roberts, J. Storm-Mathisen and H.F. Bradford, (Eds.), Excitatory Amino Acids, MacMillan Press, New York, 1986. 15 Nadler, J.V., White, W.F., Vaca, K.W., Perry, B.W. and Cotman, C.W., Biochemical correlates of transmission mediated by glutamate and aspartate, J. Neurochem., 133 (1978) 147-155. 16 Perkins, M.N. and Stone, T.W., An iontophoretic investigation of the actions of convulsant kynurenines and their interaction with the endogenous excitant, quinolinic acid, Brain Research 247 (1982) 184-187. 17 Raisman, G., The connexions of the septum, Brain, 89 (1966) 317-341. 18 Stevens, D.R., Gallagher, J.P. and Shinnick-Gallagher, P., Intracellular recordings from rat dorsolateral septal neurons, in vitro, Brain Research, 305 (1984) 353-356. 19 Storm-Mathisen, J., Localization of transmitter candidates in the brain; the hippocampal formation as a model, Prog. Neurobiol. (N.Y.), 8 (1977) 119-185. 20 Storm-Mathisen, J., Glutamic acid and excitatory nerve endings: reduction of glutamic acid uptake after axotomy, Brain Research, 120 (1977) 379-386. 21 Storm-Mathisen, J. and Woxen-Opsahl, M., Aspartate and or glutamate may be transmitters in hippocampal efferents to septum and hypothalamus, Neurosci. Lett., 9 (1978) 65-70. 22 Swanson, L.W. Sawchenko, P.E. and Cowan, W.M., Evidence that the commissural, associational and septal projections of the regio inferior of the hippocampus arise from the same neurons, Brain Research 197 (1980) 207-212. 23 Zaczek, R., Hedreen, J.C. and Coyle, J.T., Evidence for a hippocampal-septal glutaminergic pathway in the rat, Exp. Neurol., 65 (1979) 145-156. 24 Zhadina, S.D., Vinogradova, O.S. and Bragin, A.C., Effect of acetylcholine and glutamate on septal neurons in vitro. In I.P. Pavlova (Ed.), Zhurnal Vysshi Nervnoi De Yatel'nosti lmeni Vol. 30, no. 2, Plenum Press, 1983, pp. 392-397 (translation).
437
BRE 21781
Excitatory amino acid antagonists depress transmission in hippocampal projections to the lateral septum DAVID R. STEVENS and CARL W. COTMAN Department of Psychobiology, University of California, lrvine, CA 92717 (U.S.A.) (Accepted May 27th, 1986) Key words: excitatory amino acid - - lateral septum - - kynurenic acid --p-bromobenzoyl-piperazine-2,3-dicarboxylic acid - intracellular recording - - synaptic transmission - - fimbria
Antagonists of excitatory amino acid neurotransmission were tested as antagonists of septal excitatory postsynaptic potentials (EPSPs) generated by stimulation of the fimbria. Septal EPSPs were depressed in a concentration-dependent manner by kynurenic acid and P-bromobenzoyl piperazine-2,3-dicarboxylic acid but not by L-2-amino-4-phosphonobutyric acid or D-2-amino-5-phosphonopentanoic acid. These results indicate that the hippocampal projection to the lateral septum is similar to the Schaffer collateral system and is mediated by an excitatory amino acid or a similar derivative. One of the outputs of the hippocampus terminates in the dorsolateral septal area 17'22. The majority of these fibers originate in hippocampal area CA3, enter the septum via the fimbria, and are branches of the Schaffer collateral pathway x'12'2e. Since transmission in the Schaffer collateral system is thought to be mediated by an excitatory amino acid 2'15'19, it has been proposed that the hippocampal projection to the septum is also mediated by an excitatory amino acid 21. The studies described below indicate the presence, release, and uptake of glutamate, and the presence of binding sites for excitatory amino acids associated with the hippocampal projection to the lateral septum. The levels of glutamate decrease following kainic acid lesions of the hippocampus or knife cuts of the fimbria 5, and glutamate uptake decreases in the lateral septum following fimbria section 2° or kainic acid lesions of the hippocampus 23. Malthe-S0renssen and co-workers 13 have demonstrated that [3H]D-aspartate is released in a calciumdependent manner in the lateral septum following stimulation of the fimbria/fornix. These authors interpreted this result as indicating that an excitatory amino acid (probably glutamate) is the excitatory neurotransmitter in this pathway. The lateral septal
area has been shown to contain relatively high numbers of binding sites for N-methyl-D-aspartate ( N M D A ) and quisqualic acid 14 that probably represent postsynaptic receptors for glutamate, aspartate, or a related derivative. To date however, there is little evidence addressi n g the pharmacological characterization of this pathway. Zhadina and co-workers 24 have reported that firing rates of septal neurons are increased in the presence of glutamate. Also, JoEls and Urban 9 have reported that glutamate increases firing rates of septal neurons and that glutamate-diethyl-ester, ( G D E E ) but not 2-amino-5-phosphonopentanoate (AP5), applied iontophoretically, depresses potentials elicited in the septum by stimulation of the fimbria. It was concluded that an excitatory amino acid acting as a quisqualate receptor may mediate transmission of this pathway. However, the reported specificity of G D E E has not been seen in studies of the hippocampus 3. In this paper, we have used intracellular recording techniques to examine the actions of known concentrations of excitatory amino acid antagonists which have been characterized in the hippocampus, on septal excitatory postsynaptic potentials (EPSPs) resulting from stimulation of the fimbria/ fornix. The results of these experiments are com-
Correspondence: D.R. Stevens, Department of Psychobiology, University of California, Irvine, CA 92717, U.S.A. 0006-8993/86/$03.50 © 1986 Elsevier Science Publishers B.V. (Biomedical Division)
438 pared to the results of similar experiments carried out in the hippocampal slice preparation. Male Sprague-Dawley albino rats weighing 160-220 g were decapitated, the brains were rapidly removed, and slices prepared using a Vibratome. Details of the dissection have been reported previously 18. The slice was then transferred to a recording chamber for intracellular recording, lntracellular recordings were carried out using glass microelectrodes filled with 4 M K-acetate and having resistances of 80-160 Mr2. Cell input resistance was monitored continually using current injection via the bridge circuit of the intracellular amplifier (Biodyne model
A2) The recording chamber was maintained at 33 + 1 °C and the slice was constantly superfused with an artificial cerebrospinal fluid (ACSF) of the following composition (in mM): NaC1 117, KCI 2.5, CaC12 2.0 K2HPO 4 1.2, MgSO42.0, NaH2CO3 25, and D-glucose 10. The medium was bubbled with 95% 02/5% CO2 to maintain pH. The fimbria/fornix was stimulated using bipolar tungsten electrodes. Antagonists (kynurenic acid, bicuculline methiodide, L-2-amino-4-phosphonobutyric acid (AP4), Sigma; p-bromobenzoyl piperazine2,3-dicarboxylic acid (pBBPzDA) and D-amino-5phosphonovaleric acid (AP5), Tocris, England, were applied as part of the normal superfusate. Control EPSPs resulting from single stimuli (mean of 10 stimuli) were compared to the means of EPSPs recorded in the presence of antagonist (or other treatment). Following treatments, the response was followed for observation of recovery after washout of the antagonist. Stimulation of the fimbria/fornix produces short latency (2-3 ms) graded EPSPs which reach action potential threshold with increasing stimulus intensity (Fig. 1A). Using short current pulses of both polarities, membrane resistance was monitored during the EPSP, demonstrating that the EPSP is associated with an increase in conductance (Fig. 1B). Superfusion of the slice with medium containing 0.25 mM Ca 2÷ and 3.45 mM Mg 2÷ results in a depression (80%) of the EPSP amplitude (Fig. 1C). EPSPs typically rise rapidly and decline with a half-time near 20 ms. The time course of the EPSP is highly variable with durations as long as 50 ms. With increasing stimulus intensity the duration of the
A
B
l!
....... low c a l c i u m
kynurenate
5 0 0 uM
bicucullJne 10 uM
Fig. 1. lntracellular recordings from lateral septal neurons. A: septal EPSPs and an orthodromic action potential resulting from stimulation of the fimbria/fornix. B: a representative oscilloscope tracing shows that tonic potentials resulting from intracellular injection of current of either polarity are reduced during the EPSP indicating that membrane conductance is increased. C: treatment of the septal slice with low calcium-high magnesium acsf results in depression of the EPSP amplitude (78 + 5%, mean + S.D., 't' indicates the treatment response). D: kynurenic acid (500ktM) treatment results in a depression of EPSP amplitude and prolongation of the EPSP (t, treatment response). E: treatment of the septal slice with bicuculline methiodide (10/~M) results in a prolongation of the EPSP (t, 3-min superfusion, 't', 6-min superfusion).
EPSP decreases. Treatment of the slice with the excitatory amino acid antagonist kynurenic acid 16 results in depression of EPSP amplitude and an increase in the duration of the EPSP (Fig. 1D). Under similar stimulus conditions, treatment of the slice with the 7aminobutyric acid (GABA) antagonist bicuculline methiodide (10 #M) results in prolongation of the time course of the EPSP (Fig. 1E). This result indicates that the time course of the EPSP is decreased by a GABA-mediated inhibitory postsynaptic potential. The sensitivity of septal EPSPs to antagonists of excitatory amino acid transmission was tested in order to demonstrate that this pathway was sensitive to amino acid antagonists, and to compare their effectiveness in this region to that in the hippocampus. The results of treatment with a series of excitatory amino acid antagonists is shown in Fig. 2. Septal slices were treated with kynurenic acid, a recently described antagonist at hippocampal excitatory amino acid receptors 6. Kynurenic acid depressed the amplitude of the septal EPSP at a concentration of 100-500/~mol, with no noticeable effects on membrane potential or input resistance (Fig. 2A). pBBPzDA has been shown to be an effective antagonist of excitatory amino acid responses in the hippocampus 7. Treatment of the septal preparation with this agent also resulted in depression of the septal EPSP (Fig. 2A). pBBPzDA was slightly more potent
439 A
CONTROL
"IRF_ATIh~NIT
B LOG-CONCENTRATION/RESPONSE RELATION
~
loo
• •
KYN pBBPzDA
KYN200 ulM
s
~
~mSec
t
o lOO
~ooo CONCENTRATION (uM~
Fig. 2. The actions of selected excitatory amino acid antagonists on septal EPSPs. A: pairs of records demonstrating representative EPSP's during control recordings and the effect of treatment of the same neuron with the indicated agent. Kynurenic acid and pBBPzDA depressed septal EPSPs while AP4 and AP5 had no effect on septal EPSP's. B: log concentrationresponse relations for Kynurenic acid and pBBPzDA, both of which depressed EPSPs in a concentration-dependent manner with IC50values near 200pM. The bars on the plotting symbols represent one standard deviation. The two agents exhibited parallel concentration-response curves and similar potencies.
than kynurenate (Fig. 2B). AP4 (10-20 pM), an agent which acts presynaptically to depress transmission in mossy fiber terminals in the guinea pig hippocampus s, was tested and found to have no effect on membrane properties or on septal EPSP amplitude (Fig. 2A). The specific NMDA antagonist, AP5 (ref. 4), was tested on the septal EPSP also, and had no effect on membrane properties or on the EPSP at concentrations of 5 50pM. These results demonstrate that the potential generated by stimulation of fimbria/fornix is graded, calcium-dependent, and associated with a conductance increase. Our observations are in accord with the proposed synaptic nature of this response. The increase in the duration of the EPSP following depression by kynurenic acid indicates that the EPSP decay is not passive and may be altered by the presence of an inhibitory synaptic component. This result suggests the presence of a feed-forward inhibitory circuit similar to that reported in the hippocampus I. The increase in the duration of the EPSP following treatment with bicuculline methiodide, a G A B A antagonist, supports this suggestion. Bicuculline methiodide treatment increased the duration of the EPSP at low stimulus intensities well below the threshold for action potentials. These results are readily explained
by the orthodromic activation of GABAergic interneurons by fimbrial stimulation. We cannot rule out the activation of some other pathway or recurrent inhibition following the activation of other septal principal neurons, although the short latency of the bicuculline methiodide sensitive component of the synaptic response does not support the latter alternative. The actions of kynurenic acid and p B B P z D A as antagonists of the EPSP support the proposal that septal EPSPs are mediated by excitatory amino acid receptors, p B B P z D A is slightly more potent than is kynurenic acid. Both agents have IC50 values near 200 /~M, consistent with those reported for these agents on commissural/associational synapses of the hippocampus 6,7. The lack of effect of AP4 is not inconsistent with the proposal that either glutamate or aspartate is the transmitter in this pathway since AP4 appears to act on a subset of excitatory amino acid receptors 8:°. AP4 is also ineffective at the Schaffer collateral synapse 2. The failure of D-AP5 (50/~M) to antagonize the septal EPSP indicates that in this pathway, as in the Schaffer collateral projection, NMDA receptors do not normally contribute to the synaptic response II. In physiological experiments the apparent dissociation constant of AP5 for the N M D A binding site is near 5 /~M H. While NMDA receptors have been reported to be present in this area 14, their role remains unclear. The possibility that these sites play a role in synaptic plasticity needs to be examined 2:~. Our results support the proposal that glutamate or a related compound acts as the neurotransmitter of the hippocampal projection to the septal area and that the postsynaptic response is mediated by an excitatory amino acid receptor. They also indicate a high degree of similarity between the hippocampal projection to the septum and that of the Schaffer collateral pathway. This similarity is not unexpected since these two projections arise from the same neurons 22. The potencies of kynurenic acid and pBBPzDA on the septal EPSP are similar to their potencies against kainic acid responses recorded from hippocampal pyramidal cells of area C A I and on Schaffer collateral/ commissural EPSPs. These results support the suggestion that the hippocampal projection to the lateral septum is mediated by an excitatory amino acid acting at a kainic acid binding site. However, a role for
440 q u i s q u a l i c acid b i n d i n g sites c a n n o t b e r u l e d o u t w i t h o u t f u r t h e r e x a m i n a t i o n o f t h e a c t i o n s of e x o g e -
n o u s l y a p p l i e d a g o n i s t s a n d t h e i r i n t e r a c t i o n w i t h antagonists.
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