Spatial Distribution of Potentiated Synapses in

Spatial Distribution of Potentiated Synapses in Hippocampus:Dependence on Cellular Mechanisms and Network Properties

M. F. Yeckel and T. W. Berger

Department of Biomedical Engineering, Program in Neuroscience, University of Southern California, Los Angeles,California 90089-1451

Long-term potentiation (LTP) of synaptic transmission, studiedintensively in reduced brain preparations such as hippocampalbrain slices, is the leading candidate for the cellular/molecularbasis of learning and memory. Serious consideration of LTP asunderlying information storage in the intact brain, however,requires understanding how LTP can be induced selectively atspecific synaptic sites in a neural system when the mechanismsunderlying LTP are regulated by other structural and functionalproperties of the same neural system. In the studies reportedhere, we tested the hypothesis that different patterns of activitywithin the same population of entorhinal cortical afferents couldlead to a selective potentiation of spatially distinct populationsof synapses across different regions of the hippocampus, in-cluding those activated multisynaptically. We focused specifi-cally on potentiation of direct, monosynaptic entorhinal input todentate granule cells, which expresses an NMDA receptor-

dependent LTP, and on potentiation of indirect, disynaptic en-torhinal input to CA3 pyramidal cells, which is transmitted bythe mossy fiber projection of dentate granule cells and ex-presses an NMDA receptor-independent LTP. The principalfindings of these experiments show that lower stimulation fre-quencies (10–20 Hz) of entorhinal cortical axons selectivelyinduce LTP of mossy fiber input to CA3 transsynaptically viaexcitation of dentate granule cells, and that patterns of stimu-lation of that mimic neuronal firing in the entorhinal cortexduring endogenous theta rhythm (five-impulse bursts at 200 Hz,interburst intervals of 200 msec) induce LTP both monosynap-tically for input to dentate granule cells and transsynaptically formossy fiber input to CA3.

Key words: LTP; CA3; CA1; pyramidal cell; dentate gyrus;granule cell; mossy fiber; perforant path; entorhinal cortex;transsynaptic; in vivo; learning; memory

The most promising mechanism for the cellular/molecularbasis of learning and memory is long-term potentiation (LTP)of glutamatergic synaptic transmission (Berger, 1984; Land-field and Deadwyler, 1988; Morris, 1989; Bliss and Col-lingridge, 1993). Intensive experimental study of this phenom-enon has revealed that LTP can be expressed by each of thesynaptic populations forming the intrinsic, input–output path-way of the hippocampus (entorhinal cortex to dentate granulecells, granule cells to CA3 pyramidal cells, and CA3 pyramidalcells to CA1 pyramidal cells), and that the induction of LTP atany of these synaptic sites is achieved when excitatory activityincreases to a critical level (McNaughton et al., 1978; Wigstromand Gustafsson, 1983; Kelso et al., 1986). In addition, thereappear to be different “forms” of LTP based on evidence thatmore than one set of mechanisms can lead to synaptic poten-tiation. For example, LTP of entorhinal cortical input to den-tate granule cells requires activation of the NMDA subtype ofglutamate receptor, whereas LTP of dentate granule cell inputto pyramidal cells of the CA3 region can be induced indepen-dently of NMDA receptor-mediated activity (Harris and Cot-man, 1986; Wigstrom et al., 1986). The dynamics of the under-lying cellular/molecular mechanisms place constraints on the

patterns of afferent activity, which optimally induce synapticpotentiation. For example, NMDA receptor-dependent LTP ispreferentially induced by high-frequency (100 – 400 Hz) burstsof excitatory activity, because temporal summation during thebursts is an efficient means for providing the depolarization ofthe postsynaptic neuron required to relieve the Mg 21 block ofthe NMDA channel (Collingridge et al., 1988).

To date, electrophysiological studies of LTP have focused pri-marily on identifying the cellular/molecular mechanisms underly-ing potentiation of synapses activated monosynaptically, which hasnecessitated the use of reduced preparations, such as in vitro slicesand tissue cultures (Andersen et al., 1977; Malenka et al., 1988;Davies et al., 1989; Bekkers and Stevens, 1990; Foster and Mc-Naughton, 1991). When considered in the context of the intactbrain, however, additional factors related specifically to networkproperties may play essential roles in achieving the specific induc-tion requirements for LTP (Berger and Sclabassi, 1988; Berger etal., 1997). For example, the level of depolarization that results froman excitatory event after its propagation through a multisynapticpathway will be influenced substantially by the degree of progres-sive convergence or divergence in the series of anatomical projec-tions comprising that circuit. Analogously, the temporal pattern ofaction potential activity expressed by any one neuron in a circuitwill be determined strongly by the extent of nonlinear transforma-tions that occur at preceding synaptic junctions. These consider-ations raise the possibility that patterns of activity that are ineffec-tive for inducing LTP monosynaptically may be effective for other,transsynaptically activated, cells. This possibility is strengthened bythe fact that different mechanisms underlie LTP of synaptic con-nections between different subregions of hippocampus, strongly

Received Sept. 5, 1997; revised Oct. 21, 1997; accepted Oct. 23, 1997.This work was supported by National Institute of Mental Health Grants MH51722

and MH00343, the Human Frontiers Science Organization, and a National ResearchService Award fellowship to M.F.Y. We thank Christian Perron for assistance withCSD analysis and Dr. Nicholas Poolos for a critical reading of this manuscript.

Correspondence should be addressed to Dr. Mark F. Yeckel, Division ofNeuroscience, Baylor College of Medicine, Houston, TX 77030. E-mail:[email protected] © 1997 Society for Neuroscience 0270-6474/97/180438-13$05.00/0

The Journal of Neuroscience, January 1, 1998, 18(1):438–450

suggesting that the most effective pattern of activity for inducingLTP varies for different populations of synapses.

In the studies reported here, we test the hypothesis that differ-ent patterns of activity within the same population of entorhinalcortical afferents can lead to selective potentiation of spatiallydistinct populations of synapses with the hippocampus, includingLTP of synapses activated multisynaptically. Our studies of thehippocampus in vivo show that selective potentiation of differentsynaptic sites reflect different mechanisms of LTP induction, andthat a combination of anatomical and physiological factors deter-mines the spatiotemporal propagation of excitatory activitythrough intrinsic hippocampal pathways.

MATERIALS AND METHODSSurg ical preparation. Experiments were performed on adult, male NewZealand White rabbits anesthetized continuously with gaseous halothane(1.5–2.0%) while body temperature was maintained at 37–39°C. Allexperimental procedures were consistent with those outlined in NationalInstitutes of Health publication 91-3207, Preparation and Maintenance ofHigher Animals During Neuroscience Experiments.

The skull and dura overlying the parietal cortex, dorsal to the hip-pocampus, were removed, and bipolar stimulating microelectrodes(Epoxylite-insulated stainless steel 00 insect pins, with 250 mm exposedtips and spaced 500–1000 mm apart) were positioned stereotaxically intomedial perforant path fibers of the angular bundle. When examining themossy fiber pathway, a concentric bipolar stimulating electrode (RhodesMedical Instruments; 100 mm diameter, modified so that the total ex-posed length was ;350 mm) was lowered into the ventral two-thirds ofthe hilus of the ipsilateral dentate gyrus adjacent to the CA3c region.

Stainless steel recording microelectrodes (etched and insulated insectpins; 10 mm exposed tip; 1 MV measured at 135 Hz in vitro) or tungstenrecording microelectrodes (A-M Systems; 50–100 mm; 4–12 MV mea-sured at 1 kHz in vitro) were placed stereotaxically into the cell bodylayers or the dendritic layers of the dentate gyrus and the CA3 subfieldso that evoked responses could be recorded simultaneously from thesetwo subfields. In some experiments, single recording electrodes wereglued to concentric bipolar stimulating electrodes (horizontal tip sepa-ration, 100–200 mm; vertical tip separation, 500 mm) and placed such thatperforant path-evoked granule cell responses could be evaluated andmossy fibers in the hilus could be excited. In general, perforant path-to-dentate responses and mossy fiber-to-CA3c responses were evaluated fora 1 mm cross-section of the hippocampus, perpendicular to the long axis,roughly corresponding to the anatomically identified mossy fiber trajec-tory (Claiborne et al., 1986). At the completion of each experiment,recording and stimulation loci were marked by passing 100 mA of currentfor 8–15 sec through the metal electrodes; animals were deeply anesthe-tized with pentobarbital sodium and perfused transcardially with 0.9%formalin. The brains were removed and stored in 10% formalin for atleast 2 d, and frozen sections 50 mm thick were taken using a slidingmicrotome. The sectioned tissue then was stained for metal deposits with10% potassium ferrocyanide and counterstained with 0.2% saffranin-O.

In some experiments, small quantities (nanoliters per minute) ofpharmacological agents were delivered with a micropressure ejectionsystem (15 msec impulses; 5–15 psi; 1 psi 5 6.9 kPa) into the cell body ordendritic regions.

Data collection and analysis. Stimulation pulses (0.1–0.2 msec) weredelivered by using constant low frequencies (,0.2 Hz) or pairs of pulseswith interpulse intervals that varied between 10 and 200 msec. In cases inwhich two pathways were being assessed (e.g., perforant path input to thedentate and mossy fiber input to CA3), stimulation pulses were givenalternately to each pathway (0.1 Hz/pathway). In one series of experi-ments, 10–30 Hz stimulation (,30 pulses) was delivered to the perforantpath while recording in both the cell layers of the dentate gyrus and CA3.Three primary patterns of stimulation trains were tested for induction ofLTP: (1) 400 Hz trains, 10 impulses per train, one train/10 sec; (2) 100Hz trains, 50 impulses per train, one train/10 sec; and (3) 200 Hz burstsat a 10 Hz frequency (theta burst), five impulses per burst, one burst /100msec. Unless indicated differently, a total of 100 impulses were given. Insome experiments, lower frequencies of stimulation (10–50 Hz) wereused to induce LTP. The stimulation patterns of these trains were similarto the 100 Hz pattern: one to two trains of 50 pulses and one train/10 sec.Long-term potentiation was operationally defined as an increase in

population spike amplitude .120% of control response amplitude for aminimum of 20 min.

Evoked field potentials were bandpass-filtered using low- and high-frequency limits of 0.003 and 10 kHz, respectively. Unitary spike eventsgenerated by single cells were recorded simultaneously and differentiatedfrom population field potentials using low- and high-frequency limits of0.1–1.0 and 10 kHz, respectively. Data were digitized at 10–20 kHz(2400/E Series data acquisition processor, Microstar Laboratories) on aSeries 3500 HP-Apollo workstation and stored for later analysis.

Laminar analysis was performed by lowering a recording electrodeinto the dorsal hippocampus parallel to the dendritic axis of the pyra-midal and granule neurons. Recordings were started in stratum oriens ofCA1, and the electrode was lowered at increments of 50 mm until eitherthe ventral blade of the dentate gyrus or stratum oriens of CA3c wasreached. The tissue was allowed to stabilize for 1–2 min at each recordingsite, and then three to five responses were evoked at 0.1 Hz. Furtheranalysis was performed on averages of the responses. In some experi-ments, up to three stimulation intensities were given (intensity rangedfrom threshold for activation of single action potentials in stratumpyramidale to intensities supramaximal for evoked field responses).Additionally, in cases in which 10 Hz stimulation trains were given, theintertrain interval was 1 min. The one-dimensional current–source den-sity (CSD) at each site (x) was calculated using the following formula:

Ix 5 2s~Ex2h 2 2Ex 1 Ex1h!/4h2,

where Ix is the current at location x, h is the sampling distance (50 mm),Ex is the extracellular voltage at location x, Ex2h is the extracellularvoltage at location x 2 h, Ex1h is the extracellular voltage at location x 1h, and s is the tissue conductivity tensor (for a complete treatment of thetheoretical basis of CSD analysis, see Freeman and Nicholson, 1975).The tissue (i.e., extracellular space) was assumed to be isotropic for thepyramidal cell regions of the hippocampus.

Because the stability of brain tissue is critical for an accurate renderingof laminar profiles and calculated CSDs, basic physiological function ofthe animal was carefully monitored, and any aberration in breathing,heart rate, or EEG resulted in an end to the experiment.

RESULTSFrequency-dependent excitation of hippocampalgranule and pyramidal neuronsThe primary excitatory afferent to hippocampus originates inlayer II of the entorhinal cortex and projects monosynaptically tothe dentate gyrus and the CA3 pyramidal cell region (Lorente deNo, 1933; Steward and Scoville, 1976; Tamamaki and Nojyo,1993; Witter, 1993). Intrinsic projections from dentate granulecells to CA3 pyramidal neurons provide an additional cascade ofexcitatory input (Swanson et al., 1978; Amaral and Witter, 1989).Thus, entorhinal cortical and intrinsic hippocampal afferentsform the basis for a feedforward excitation of the pyramidal cellfields, such that an excitatory volley from the entorhinal cortexexcites both dentate granule cells and hippocampal pyramidalcells simultaneously (Yeckel and Berger, 1995b) and subse-quently induces longer latency excitation of CA3 and CA1 neu-rons (Yeckel and Berger, 1990, 1995b; Berger and Yeckel, 1991).

Consistent with sequential excitation through the trisynapticpathway, latencies to population and single-cell responses wereprogressively longer for each of the three subfields: (1) entorhinalexcitation of the dentate granule cells, 4.5–5.5 msec; (2) disynap-tic excitation of CA3, 8–13 msec; and (3) trisynaptic excitation ofCA1, 16–21 msec. The longer latency excitations of CA3 and CA1neurons could not be maintained in response to continuous stim-ulation with frequencies .25 Hz because of failure of propaga-tion of the response across synapses, and therefore, suggestsmultisynaptic excitation. The latencies of pyramidal cell dis-charge also were consistent with the cumulative monosynapticlatencies for each of the individual pathways constituting thetrisynaptic circuit: 4–6 msec for medial perforant pathway inputto granule cells, 4–7 msec for activation of CA3 by mossy fiber

Yeckel and Berger • Induction of Transsynaptic Hippocampal LTP J. Neurosci., January 1, 1998, 18(1):438–450 439

stimulation, and 5–8 msec for responses of CA1 cells evoked bySchaffer collateral stimulation. Disynaptic excitation of CA1 wasconsistent with monosynaptic excitation of CA3, followed bymonosynaptic excitation via the Schaffer collaterals (Fig. 1). In-hibition of perforant path-evoked granule cell activity by pressureejection of the GABAA agonist muscimol (100–500 mM) locallyinto the dentate gyrus was accompanied by a suppression oflong-latency responses observed in CA3 (Fig. 2). Conversely,local application of the GABAA antagonist bicuculline methio-dide (100–200 mM) increased granule cell responsiveness to en-torhinal input and selectively increased the magnitude of long-latency CA3 responses (Yeckel and Berger, 1990; Berger andYeckel, 1991).

On the basis of previous reports that stimulation of perforant

path fibers with frequencies of ;10 Hz leads to a progressiverecruitment in the number of active granule cells (i.e., frequencyfacilitation; Andersen et al., 1971; Munoz et al., 1991), and on thebasis of quantitative anatomical findings that granule cell outputconverges substantially in its projection to CA3 (estimates asgreat as 12:1; Gaarskjaer, 1978; Amaral et al., 1990), we tested thehypothesis that the relative strengths of monosynaptic and mul-tisynaptic entorhinal inputs to hippocampal pyramidal cells arefrequency-dependent. Using stimulation frequencies rangingfrom 1 to 30 Hz (total number of pulses, 20–30), we found thatmonosynaptic excitation of pyramidal cells by perforant pathfibers occurs preferentially in response to frequencies of electricalstimulation ,5 Hz. In contrast, frequencies of 10–15 Hz greatlyenhances polysynaptic excitation of hippocampal pyramidal neu-rons through the intrinsic pathways (Yeckel and Berger, 1990;Berger and Yeckel, 1991) (Fig. 3).

To resolve the propagation of excitation through the intrinsichippocampal pathways better, CSD analysis was performed onlaminar profiles of field potentials evoked by 10 Hz stimulation ofperforant path fibers. A recording electrode was lowered into thedorsal hippocampus at increments of 50 mm, and three 10 Hztrains of 15 impulses were delivered (intertrain interval was 1min). Evoked responses were averaged for each site, and esti-mates of the one-dimensional CSD were generated. The results ofthese experiments showed that current sinks generated by the firstcouple of stimulation pulses in a 10 Hz train were short in latency(typically ,7 msec), and limited to the stratum moleculare of thedentate gyrus and stratum lacunosum–moleculare of both theCA3 and CA1 subfields. Additional stimulation pulses in a 10 Hz

Figure 1. Serial propagation through the trisynaptic pathway. Schematicdiagram of the intrinsic organization of hippocampus and associatedphysiological responses (data are from different preparations). Top, En-torhinal innervation of the dentate gyrus via the perforant path ( pp).Perforant path-evoked population spike in the dentate gyrus (arrowindicates a unitary discharge). Second from top, Mossy fiber (mf ) input tothe CA3 pyramidal cell region originating from dentate granule cells.Stimulation of granule cell axons evokes single units (arrows) and theassociated population spike. Second from Bottom, Innervation of the CA1pyramidal cell region by CA3 via the Schaffer collaterals (sch). Stimula-tion of CA3 axons within stratum radiatum of CA1 resulted in a mono-synaptically evoked population spike. Bottom, Trisynaptic excitation ofCA1 by 10 Hz stimulation of perforant path fibers. Single action potentialswere excited at latencies corresponding to monosynaptic, disynaptic, andtrisynaptic excitation. Disynaptic excitation of CA1 resulted from mono-synaptic excitation of CA3, which in turn excited CA1 monosynaptically.

Figure 2. Characterization of disynaptically evoked responses in CA3 bystimulation of entorhinal afferents. The presence of disynaptic input toCA3, via mossy fiber (mf ) input, was confirmed by selectively alteringgranule cell responsiveness to perforant path ( pp) stimulation and ob-serving changes in longer-latency components of CA3 field responses.Local application of the GABAA agonist muscimol (100–500 mM) into thedentate gyrus suppressed granule cell responsiveness (arrow) to perforantpath stimulation (the same stimulation intensity was used for both controland muscimol conditions) and concomitantly increased suppression of thelonger-latency component of the CA3 response (arrow). Waveforms rep-resent averages of five responses. Calibration bar, 5 msec and 1 mV.

440 J. Neurosci., January 1, 1998, 18(1):438–450 Yeckel and Berger • Induction of Transsynaptic Hippocampal LTP

train resulted in disynaptic and trisynaptic excitation of the py-ramidal cell regions via the trisynaptic pathway, as evidenced bylonger-latency current sinks (12–18 msec) in stratum lucidum ofthe CA3 subfield and stratum radiatum of the CA1 subfield (Fig.4). These findings reveal that the magnitude of disynaptic andtrisynaptic excitatory input to CA3 and CA1 pyramidal neurons ispredominantly determined by the frequency of perforant pathinput, with 10–15 Hz resulting in the maximum excitation.

Frequency-dependent induction of perforant path andmossy fiber LTPWe next examined the relationship between frequency of afferentstimulation and the probability and magnitude of LTP inductionof monosynaptic, perforant path input to dentate granule cellsand monosynaptic, mossy fiber input to CA3 pyramidal cells.Field potentials evoked in CA3, in vivo, have not been character-ized adequately, however, so it was first necessary to establishcriteria for identification of monosynaptically evoked mossy fiberresponses (Yeckel and Berger, 1995a; Berger et al., 1997).

Distinguishing CA3 responses to mossy fiber excitation versusother afferents is technically difficult and is an issue that remains

controversial (Williams and Johnston, 1991; Claiborne et al.,1993; Langdon et al., 1993). Pyramidal neurons of CA3 receivefour major sets of synaptic input: monosynaptic perforant pathinput, mossy fiber input, innervation by other CA3 pyramidalneurons via recurrent collaterals, and inhibitory input fromGABAergic interneurons local to the CA3 subfield. In addition tocriteria such as the presence of orthodromic, short-latency unitarydischarges in stratum pyramidale of CA3, laminar and CSDanalyses were used to identify stratum lucidum as the location forshort-latency current sinks generated in response to hilar stimu-lation and to distinguish these mossy fiber-evoked responses fromother afferents such as the recurrent collaterals.

Although recordings were made from the CA3c region (prox-imal to the dentate gyrus)—a subregion of CA3 with a lowerdensity of recurrent innervation than has been reported for theCA3a and CA3b subfields (Lorente de No, 1933; Ishizuka et al.,1990)—we still found considerable contamination of monosynap-tically evoked mossy fiber field responses by polysynaptic activa-tion via the recurrent collaterals. The difficulty of interpretingCA3 field responses evoked by stimulating presumed mossy fibers

Figure 3. A, Diagrammatic representation of the relative strengths of monosynaptic and polysynaptic excitation of the pyramidal cell regions as afunction of the frequency of entorhinal input. Top, Stimulation frequencies ,5 or .20 Hz result predominantly in monosynaptic activation of CA3 andCA1 pyramidal neurons. Bottom, Stimulation frequencies between 5 and 15 Hz result in enhanced excitation of pyramidal cells transsynaptically. B,Frequency-dependent excitation of dentate granule cells. Recruitment of granule cell activity with 10 Hz stimulation of entorhinal afferents. Populationfield responses (top) and multiunit recordings (bottom) recorded from the granule cell layer of the dentate gyrus show progressive facilitation of evokedresponses during a 10 Hz stimulation train (20–30 impulses). Facilitation was observed only during the stimulation train. Calibration bar, 50 mV and 1mV for units and fields, respectively, and 5 msec. C, Frequency-dependent disynaptic excitation of hippocampal CA3 pyramidal cells. Trains ofstimulation were given at frequencies of 1, 10, and 20 Hz (30 impulses total) to perforant path fibers. Frequency-dependent excitation of direct perforantpath input to CA3 (similar to facilitation of perforant path-to-dentate responses above) and indirect (i.e., disynaptic) input via the mossy fiber projection(arrows identify monosynaptically and disynaptically evoked population spikes). Calibration bar, 1 mV and 5 msec.



in the hilus (i.e., distinguishing the onset and slope of monosyn-aptic mossy fiber activation from that of population EPSPs evokedby recurrent collaterals) is apparent with laminar analysis of fieldpotentials and estimations of one-dimensional CSDs (n 5 5). Theresults of laminar analysis showed that mossy fiber-evoked pop-ulation EPSPs recorded in stratum lucidum of CA3 were oftenirregular in appearance because of the presence of numeroussingle action potentials generated in nearby stratum pyramidaleof CA3 (,75 mm away). Although relatively low stimulationintensities were used, we were not able to evoke a populationEPSP subthreshold for generation of single action potentials instratum pyramidale of CA3 that was detectable above backgroundnoise (Fig. 5A). Similarly, the proximity of evoked synaptic re-sponses to associated action potential generation made it difficultto resolve population spikes recorded in stratum pyramidale frompopulation EPSPs generated in stratum lucidum. In contrast, fieldresponses recorded in stratum radiatum, reflecting disynapticexcitation by recurrent collaterals, were smooth in appearance,similar to population EPSPs recorded in stratum moleculare ofthe dentate gyrus or population EPSPs recorded in stratum ra-diatum of CA1. Current–source density analysis confirmed stra-tum lucidum and stratum pyramidale as the locations for short-latency current sinks generated in response to hilar stimulationand showed a longer-latency current sink in stratum radiatum(and corresponding current source in stratum pyramidale) gen-erated by disynaptic recurrent excitation (Fig. 5B).

Although field potentials recorded in stratum pyramidale rep-resent a composite of the population spike and the populationEPSP, isolation of monosynaptically evoked mossy fiber re-sponses was performed most accurately by recording in stratumpyramidale, where contamination attributable to disynaptic exci-tation via the recurrents occurred later in time and, as sourcecurrent, could be distinguished more easily from the current sinkgenerated by monosynaptic mossy fiber excitation. Mossy fiber-evoked responses were quantified by measuring the amplitude ofthe negative peak of the short-latency component of the response.

As we have shown previously (Berger and Yeckel, 1991), and ashas been confirmed by others (Derrick and Martinez, 1994), high-frequency stimulation (.100 Hz) is less optimal for inducing mossyfiber LTP. As shown in Figure 6, comparison of the effects of 100and 400 Hz stimulation (50–100 impulses) revealed that 100 Hz wassignificantly more efficient at inducing mossy fiber LTP than 400 Hztetanization, both in terms of probability of induction and magnitudeof potentiation (n 5 5 for each frequency; unpaired t test, p , 0.001).This is in contrast to perforant path input to the dentate (n 5 5 for

each frequency), in which the probability of induction and magni-tude of potentiation were not significantly different when 100 and400 Hz frequencies were used to induce LTP (Fig. 6A). Additionalexperiments revealed that the probability of inducing mossy fiberLTP was significantly greater for stimulation frequencies of 10–50Hz (50–100 pulses total) than for higher frequencies of 100–400 Hz(67 and 33%, respectively); the opposite trend was found for theprobability of LTP induction of the dentate gyrus (.100 Hz, 80%;,25 Hz, 25%). Consistent with previous reports, it also was foundthat stimulation frequencies as low as 10 Hz were capable of induc-ing mossy fiber LTP (Zalutsky and Nicoll, 1990), whereas the lowestfrequency capable of inducing perforant path LTP was 25 Hz (Fig.6B).

The difference in optimal stimulation parameters for LTPinduction of perforant path and mossy fiber synapses may reflectthe known differences in the cellular mechanisms underlyingpotentiation of the two pathways (Harris and Cotman, 1986;Wigstrom et al., 1986). Application of the NMDA receptor an-tagonist D-APV (50–100 mM) blocked the induction of perforantpath LTP (12 of 12 experiments, 400 Hz); recovery occurred;30–40 min after D-APV administration, as assessed by theability to induce LTP (n 5 4). This is in contrast to LTP of mossyfiber input to CA3, in which LTP was induced despite pressureejection of D-APV (50–100 mM) into the CA3 subfield (6 of 15experiments, 100 Hz stimulation trains used). The lack of NMDAreceptor dependence for mossy fiber LTP confirms the interpre-tation that responses evoked in CA3 by hilar stimulation repre-sent excitation of mossy fibers and not inadvertent activation ofperforant path or Schaffer collaterals, for which LTP induction isNMDA receptor-dependent (Fig. 6C).

Transsynaptic induction of mossy fiber LTP afterstimulation of perforant pathThe experiments presented thus far demonstrate that 10 Hz stim-ulation of entorhinal afferents greatly enhances disynaptic excita-tion of CA3 pyramidal cells, and that comparable stimulationfrequencies are capable of inducing LTP of mossy fiber input toCA3 cells. These findings suggest that when entorhinal activityapproaches this frequency range, there may be sufficient recruit-ment of granule cell output to CA3 to reach the critical level ofexcitation required for induction of LTP at mossy fiber synapses.We tested this hypothesis by investigating whether 10–15 Hzstimulation of perforant path fibers was capable of inducing LTP ofmossy fiber input to CA3 transsynaptically via monosynaptic exci-tation of the dentate gyrus and disynaptic excitation of CA3. In one

4

Figure 4. Top. Transsynaptic excitation of CA1 pyramidal cells with 10 Hz stimulation. Contour plots of one-dimensional CSDs computed from fieldresponses evoked by 10 Hz stimulation of perforant path fibers. Each panel shows the spatial and temporal distribution of current sinks (2 values) andcurrent sources (1 values) for a given impulse in the stimulation train (impulse number listed above the plot). The presence of short-latency current sinksin the distal dendritic region of CA1 and the middle- to outer-third of dentate granule cells is consistent with monosynaptic excitation by entorhinalafferents (mono; monosynaptically evoked population discharges are identified with an asterisk). A longer-latency current sink, present in stratumradiatum of CA1, is consistent with trisynaptic excitation of CA1 via Schaffer collaterals (tri).

Figure 5. Bottom. Laminar analyses of mossy fiber-evoked responses in CA3. Data show the difficulty in distinguishing monosynaptic and polysynapticresponses evoked by stimulation of mossy fiber axons in the hilus. A, Laminar profile of field potentials recorded from the proximal two-thirds of theCA3c subfield (the region bound by the white dashed lines) while stimulating mossy fibers in the hilus. Vertical lines identify various components of thewaveform: (1) the line on the lef t (blue) shows the onset of the mossy fiber-evoked population EPSP; (2) the middle line (red) identifies the latency foractivation of the population spike (and contamination of the population EPSP recorded in the dendrites); (3) the line on the right (green) shows the latencyto peak of disynaptic activation of CA3 via the recurrent collaterals. Waveforms are averages of five responses recorded at 50 mm increments. Calibrationbar, 0.6 mV and 5 msec. B, Current-source density representation of mossy fiber excitation. Contour plot of one-dimensional CSD identifies the spatialand temporal distribution of current sinks (2 values) and current sources (1 values). The presence of a short-latency current sink in stratum lucidumof CA3 (proximal apical dendrites) is consistent with monosynaptic excitation (mono) by mossy fiber input. Longer-latency current sinks, present instratum radiatum of CA3 and CA1, are consistent with disynaptic excitation (di) via the recurrent collaterals and Schaffer collaterals, respectively.


series of experiments, stimulating electrodes were placed both inthe perforant path and in the hilus of the dentate gyrus to activatemossy fiber axons of granule cells; the possibility of mossy fiberLTP was assessed by examining CA3 responses evoked by hilarstimulation before and after stimulating the perforant path with10–15 or 400 Hz (Fig. 7). In support of our hypothesis, resultsrevealed that 10–15 Hz stimulation (50–100 impulses) of entorhi-nal cortical axons induced LTP of mossy fiber input to CA3transsynaptically via the dentate gyrus (n 5 6; paired t test, p ,0.01). LTP was not induced transsynaptically when 400 Hz stimu-lation was used (eight of eight experiments). As described previ-ously, stimulation frequencies ,25 Hz did not induce LTP of

perforant path input to the dentate gyrus (see Fig. 6B). These dataindicate that long-lasting synaptic potentiation occurs preferen-tially within different subregions of hippocampus depending on thefrequency of entorhinal cortical input, and that potentiation canoccur selectively for transsynaptically activated synapses.

Modulation of transsynaptic LTPResults of the previous experiments predict that the probability ofentorhinal cortical input inducing LTP will depend in part on thelevel of synaptic potentiation at the time of that input. Partialsupport for this prediction was obtained from experiments inwhich polysynaptic responses were recorded from CA3 pyramidal

Figure 6. Optimal stimulation patterns differ for induction of perforant path and mossy fiber LTP. A, Data from representative experiments (lef t) inwhich 100 Hz stimulation (or 400 Hz) was given to either the mossy fiber pathway or the perforant path, followed by 400 Hz stimulation (or 100 Hz).Regardless of the order in which the two different stimulation frequencies were delivered to mossy fibers, 100 Hz stimulation induced a greater-magnitudeLTP than 400 Hz stimulation. Group data (right) show that the 100 Hz stimulation protocol induces significantly greater-magnitude mossy fiber LTP thanthe 400 Hz stimulation protocol (top); similar differences were not found for 100 and 400 Hz stimulation of perforant path fibers (bottom) (waveformsare averages of five responses). Calibration bar, 1 mV and 5 msec. Graphs summarize results from five experiments for each stimulation frequency. Errorbars indicate SEM. B, The probability of inducing mossy fiber LTP was significantly greater for lower stimulation frequencies (,50 Hz) than for higher(.100 Hz) frequencies. In contrast, there was a greater probability of inducing LTP of perforant path input at higher frequencies of stimulation (.50Hz; LTP was never induced for perforant path input with stimulation frequencies ,25 Hz). C, LTP of mossy fiber input to CA3 is NMDAreceptor-independent. Pressure ejection of the NMDA antagonist D-APV (50–100 mM; ;70 nl/min for 15–30 min) into the apical dendrites of CA3 anddentate granule cells resulted in the selective blockade of perforant path LTP (for both monosynaptic input to CA3 and the dentate gyrus; dentatepopulation spikes shown here) but not mossy fiber LTP (waveforms represent averages of five responses). Calibration bar, 1 mV and 5 msec.


neurons after monosynaptic LTP of perforant path axons. Resultsshowed that increasing the synaptic strength of monosynapticentorhinal afferents significantly decreased the number of im-pulses necessary to evoke suprathreshold disynaptic and trisyn-aptic responses of CA3 and CA1 and significantly increased themaximum amplitude of the evoked polysynaptic response (Bergerand Yeckel, 1991). Changes in propagation through the hip-pocampus during 10–15 Hz frequency trains was examined bycomputing the integrated area of the long-latency component ofCA3 potentials both before and after LTP of perforant path inputwith 400 Hz stimulation. The region of the evoked responsecorresponded to previously identified disynaptic activation (seeFig. 2). Results showed that LTP of perforant path input signif-icantly changed propagation to CA3 (Fig. 8); polysynaptic activa-tion increased in 8 of 11 experiments in which LTP was induced(one-way repeated measures ANOVA, F 5 5.532; p , 0.05). Twoexperiments were excluded from analysis because of interictalbursting that occurred during and after the stimulation trains.

These data suggest that potentiation of perforant path/dentatesynapses will increase the likelihood of transsynaptic LTP induc-tion of mossy fiber input to CA3 pyramidal cells. This hypothesiswas tested by first stimulating perforant path fibers at a lowfrequency (10–15 Hz, 50–100 impulses) and with stimulationintensities subthreshold for transsynaptic induction of mossy fiberLTP (i.e., very little disynaptic activation of CA3 was evidentduring the stimulation train). After induction of monosynapticLTP of perforant path input with 400 Hz stimulation, the iden-tical low-frequency, low-intensity stimulation parameters resultedin significantly greater disynaptic excitation of CA3 during the

stimulation train, and consequently, transsynaptic LTP of mossyfiber input (n 5 5; one-way repeated measures ANOVA, F 57.656; p , 0.01, using Gieser–Greenhouse correction for repeatedmeasures) (Fig. 9).

Multisynaptic induction of hippocampal LTP withtheta-patterned stimulationThe selective induction of perforant path and mossy fiber LTPwith high- and low-frequency inputs, respectively, strongly sug-gests that input signals incorporating induction parameters opti-mal for both pathways will potentiate both populations of syn-apses. To test this possibility, perforant path fibers werestimulated with high-frequency “bursts” of impulses (each burstconsisted of five pulses delivered at 200 Hz), applied at a lowfrequency (interburst intervals of 100 msec), a pattern of activityobserved in layer II of the entorhinal cortex during behaviorsassociated with theta rhythm generation (Alonso and Garcia-Austt, 1987), and shown previously to induce NMDA receptor-dependent LTP maximally (Larson and Lynch, 1986; Rose andDunwiddie, 1986). The high frequency of activity within bursts isoptimal for LTP of the perforant path; the lower frequencycharacterized by the interburst interval is optimal for LTP ofmossy fibers. Stimulating the perforant path with this theta-patterned input (theta burst) resulted in the induction of LTP bothmonosynaptically for perforant path input to dentate and trans-synaptically for mossy fiber input to CA3 (n 5 5; two-way re-peated measures ANOVA, F 5 67.393; p , 0.001) (Fig. 10). Inadditional experiments it was found that giving theta burst stim-ulation at a lower intensity, or with fewer bursts (,20), could

Figure 7. Transsynaptic induction of mossy fi-ber LTP with 10 –15 Hz stimulation of entorhinalafferents. Top, Schematic of experimental proce-dure: (1) baseline recordings were obtained bystimulating, alternately, the perforant path ( pp)and recording evoked dentate population spikes(right, pre) and mossy fibers (mf ) while recordingevoked CA3 pyramidal cell responses ( far right,pre); (2) stimulating the perforant path at a lowfrequency (10 –15 Hz; 50 –100 impulses); and (3)repeating part 1 to determine whether there wasa change in the magnitude of evoked responses( post). Stimulation of perforant path fibers at10 –15 Hz resulted in LTP of mossy fiber input toCA3 transsynaptically via monosynaptic excita-tion of the dentate gyrus and disynaptic excita-tion of CA3. Direct, monosynaptic input to thedentate gyrus never exhibited LTP with low-frequency stimulation trains (waveforms repre-sent averages of five waveforms). Calibrationbar, 1 mV and 5 msec. Bottom, Summary plot ofexperiments in which transsynaptic LTP ofmossy fiber input was induced with low-frequency stimulation of perforant path fibers(n 5 6). Error bars indicate SEM.


result in monosynaptic LTP but not transsynaptic LTP. Aftermonosynaptic perforant path LTP, however, it was found thatgiving the identical theta burst stimulation (previously subthresh-old for induction of transsynaptic LTP) could induce LTP trans-synaptically for mossy fiber input to CA3 (n 5 4). LTP induced bytheta burst stimulation was NMDA receptor-dependent for per-forant path input but not for LTP of transsynaptic input to CA3,as assessed by the application of D-APV (n 5 2); this againconfirmed that transsynaptic LTP occurred via the mossy fiberprojection (Fig. 11).

DISCUSSIONIn the studies reported here, we tested the hypothesis that differ-ent patterns of activity within the same population of entorhinalcortical afferents could lead to a selective potentiation of spatiallydistinct populations of synapses within hippocampus, includingthose activated transsynaptically. We focused specifically on po-tentiation of direct, monosynaptic entorhinal input to dentategranule cells, which expresses an NMDA receptor-dependentLTP, and on potentiation of indirect, disynaptic entorhinal input

to CA3 pyramidal cells, which is transmitted by the mossy fiberprojection of dentate granule cells and expresses an NMDAreceptor-independent LTP. The principal findings of these exper-iments are as follows: (1) the stimulus prerequisites for theinduction of LTP differ for entorhinal afferents to the dentategyrus and mossy fiber afferents to CA3, such that higher stimu-lation frequencies (.100 Hz) are more effective at inducing LTPof perforant path input to granule cells of the dentate, and lowerstimulation frequencies (,100 Hz) induce monosynaptic mossyfiber LTP of CA3 pyramidal cells more efficiently; (2) consistentwith the findings that propagation of activity through the trisyn-aptic pathway predominantly results from electrical stimulation ofentorhinal afferents at 10–15 Hz, and that mossy fiber LTP can beinduced with stimulation frequencies as low as 10 Hz, we foundthat low frequencies of activity of entorhinal cortical axons(10–20 Hz) induces transsynaptic LTP of dentate granule cellmossy fiber input to CA3 selectively; (3) patterns of stimulationthat mimic neuronal firing in the entorhinal cortex during endog-enous theta rhythm induce LTP both monosynaptically for input

Figure 8. Propagation of excitation topyramidal cells after LTP of entorhinalafferents. A, Disynaptic responses wereevoked in CA3 with 10 Hz stimulation ofentorhinal afferents (30 pulses total). Af-ter induction of perforant path-to-dentate LTP (data not shown), propaga-tion to CA3 increased based on the sizeof the late component [disynaptic (di)] ofthe evoked response (waveforms are sin-gle evoked responses recorded during a10 Hz stimulation train). Calibrationbar, 0.4 mV and 4 msec. B, Disynapticpropagation to CA3 increased signifi-cantly after perforant path LTP ( post ppLTP), as assessed by the integrated areaof longer-latency responses correspond-ing to disynaptic excitation (n 5 9). Er-ror bars indicate SEM.


to dentate granule cells and transsynaptically for mossy fiberinput to CA3; and (4) the ability to induce transsynaptic LTP ofmossy fiber input to CA3 with stimulation of entorhinal afferentsusing either low-frequency or theta burst stimulation depends onthe synaptic strength of monosynaptic input to the dentate, suchthat LTP of perforant path synapses increases the likelihood oftranssynaptic induction of mossy fiber LTP.

The results of these experiments show that in the intact hip-pocampus, the constraints on perforant path and mossy fiber LTPinduction imposed by differences at the cellular/molecular levelinteract with additional constraints imposed by other properties ofthe network in which those synapses are embedded. Both perforantpath and mossy fiber synapses require a critical level of excitatoryinput from the entorhinal cortex, either monosynaptically or trans-

synaptically. Although sharing this common requirement, it hasbeen demonstrated previously, and confirmed here in vivo, that thecellular/molecular mechanisms triggered when that critical level ofexcitatory input is reached are different for the two populations ofsynapses; perforant path LTP is NMDA receptor-dependent,whereas mossy fiber LTP is not (Harris and Cotman, 1986; Wig-strom et al., 1986). The cellular/molecular differences underlyingthe induction of LTP for these different subsets of synapses alsomay account for the differences in the optimal tetanization param-eters necessary for inducing LTP. At lower frequencies of stimu-lation, LTP of perforant path input to the dentate was not inducedbecause stimulation at 10–15 Hz does not result in sufficient de-polarization to relieve the Mg21 block of NMDA channels (Blan-pied and Berger, 1992). When the frequency of entorhinal input

Figure 9. Induction of transsynaptic mossy fiber LTP is enabled byprevious induction of perforant path LTP. Top, Schematic of stimulationparadigm: (1), alternate stimulation of the perforant path ( pp) and mossyfiber (mf ) path while recording in the dentate and CA3, respectively; (2),evoked responses after low-frequency stimulation (10–15 Hz, 50–100impulses) of perforant path fibers at intensities subthreshold for inductionof transsynaptic mossy fiber LTP; (3), high-frequency stimulation (400Hz, 50–100 impulses) of perforant path fibers resulted in LTP of thedentate gyrus but not mossy fiber input to CA3; (4), propagation throughthe trisynaptic pathway was sufficiently enhanced after induction of per-forant path LTP so that low-frequency stimulation [identical to thatdelivered in (1)] induced mossy fiber LTP transsynaptically. Antidromicactivation of dentate granule cells was not effected. Arrows point toexamples of mossy fiber-evoked single units. Bottom, Summary data (n 55) Error bars indicate SEM.


increases to $50–100 Hz, temporal summation of perforant pathinput increases sufficiently for NMDA receptor activation and,thus, for the induction of LTP (Collingridge et al., 1988). At thelevel of mossy fiber input to CA3, frequencies of $50–100 Hzwithin the perforant path are not efficiently transmitted transsyn-aptically, and as a result, the induction of LTP is restricted tomonosynaptically activated synapses. Although entorhinal activityin the range of 10–15 Hz does not lead to LTP of perforant pathsynapses, it does lead to frequency facilitation, i.e., a progressiveincrease in the number of active granule cells distributed over awider spatial extent of the dentate gyrus. Because of the highdegree of convergence of granule cell projections onto CA3 pyra-midal cells, the increased number of active granule cells leads togreater levels of excitatory input sufficient to induce LTP of mossyfiber synapses within CA3. Theta burst stimulation incorporatesinput parameters optimal for potentiation of both perforant pathand mossy fiber synapses, and as a result, LTP is induced mono-synaptically and transsynaptically. Thus, potentiation of perforantpath and mossy fiber synapses is achieved not only through twodifferent mechanisms at the cellular/molecular level but alsothrough two different mechanisms at the network level; the criticallevel of excitatory perforant path input to granule cells is reached

as a consequence of temporal coincidence, whereas the criticallevel of mossy fiber excitation to CA3 is reached as a consequenceof spatial convergence.

More generally, the interaction between cellular/molecular con-straints and network constraints illustrated by these results pro-vides an empirical basis for the possibility that the spatial distribu-tion of potentiated synapses within a neural network variesdynamically as a function of the temporal properties of afferentactivity. The relevance of this finding to hippocampal function isbased on the strong correspondence between the stimulation pa-rameters used here and the temporal characteristics of actionpotential activity of hippocampal and entorhinal cortical neuronsobserved in the behaving animal (Ranck, 1973; Berger et al., 1983).Further elaboration of this basic principle may provide a basis forunderstanding several well documented properties of hippocampalneuronal population activity that evolve as a consequence of asso-ciative learning in the intact animal: the nonuniform spatial distri-bution of hippocampal neurons expressing learning-enhanced ac-tivity (i.e., “neural correlates”) (O’Keefe, 1979; Berger et al., 1983;Eichenbaum et al., 1989), the dependence of a particular spatialdistribution of neural correlates on task requirements and behav-ioral context (Christian and Deadwyler, 1986; Wiener et al., 1989),

Figure 10. Multisynaptic induction of hippocam-pal LTP with theta-patterned stimulation. Sche-matic representation of the hypothesized patternof input to granule cells of the dentate gyrus andCA3 pyramidal neurons using a stimulation pat-tern incorporating both high- and low-frequencyimpulses (200 Hz bursts of 5 impulses at 10 Hz; 50bursts). Data show the consequences of theta-likepatterns of perforant path ( pp) activation (thetaburst) on the induction of LTP (n 5 5). Error barsindicate SEM. LTP is induced both monosynapti-cally and transsynaptically (i.e., multisynaptically)for perforant path input to the dentate gyrus(monosynaptic LTP; top) and for mossy fiber (mf )input to CA3 (transsynaptic LTP; bottom). Wave-forms are averages of all responses evoked in thelast 5 min of a sample period. Calibration bars, 1mV and 5 msec.


Figure 11. NMDA-independent induction of transsynaptic LTP. Pressure ejection of the NMDA receptor antagonist D-APV (100 mM; ;70 nl/min for15–30 min) into the apical dendrites of the CA3 and the dentate gyrus before theta burst stimulation blocked the induction of perforant path ( pp) LTP(for both monosynaptic input to CA3 and the dentate) but not transsynaptic mossy fiber (mf ) LTP (waveforms are averages of responses collected duringlast 5 min of a given sample period). Calibration bars, 1 mV and 5 msec.


and the functional importance of the characteristic theta rhythmic-ity of hippocampal neural activity (Berry and Thompson, 1978;Otto et al., 1991).

REFERENCESAlonso A, Garcia-Austt E (1987) Neuronal sources of theta rhythm in

the entorhinal cortex of the rat. II. Phase relations between unitdischarges and theta field potentials. Exp Brain Res 67:502–509.

Amaral DG, Witter MP (1989) The three-dimensional organization ofthe hippocampal formation: a review of anatomical data. Neuroscience31:571–591.

Amaral DG, Ishizuka N, Claiborne B (1990) Neurons, numbers and thehippocampal network. In: Understanding the brain through the hip-pocampus: the hippocampal region as a model for studying structuresand function (Storm-Mathisen J, Zimmer J, Ottersen OP, eds), pp1–11. Amsterdam: Elsevier.

Andersen P, Bliss TVP, Skrede KK (1971) Unit analysis of hippocampalpopulation spikes. Exp Brain Res 13:208–221.

Andersen P, Sundberg SH, Sveen O (1977) Specific long-lasting potentia-tion of synaptic transmission in hippocampal slices. Nature 266:736–737.

Bekkers JM, Stevens CF (1990) Presynaptic mechanism for long-termpotentiation in the hippocampus. Nature 346:724–729.

Berger TW (1984) Long-term potentiation of hippocampal synaptictransmission affects rate of behavioral learning. Science 627–630.

Berger TW, Sclabassi RJ (1988) Long-term potentiation and its relation tohippocampal pyramidal cell activity and behavioral learning during clas-sical conditioning. In: Long-term potentiation: from biophysics to behav-ior (Landfield PW, Deadwyler SA, eds), pp 467–497. New York: Liss.

Berger TW, Yeckel MF (1991) Long-term potentiation of entorhinalafferents to the hippocampus: enhanced propagation of activity throughthe trisynaptic pathway. In: Long-term potentiation: a debate of currentissues (Baudry M, Davis JL, eds), pp 327–356. Cambridge, MA: MIT.

Berger TW, Rinaldi P, Weisz DJ, Thompson RF (1983) Single unit analysisof different hippocampal cell types during classical conditioning of therabbit nictitating membrane response. J Neurophysiol 50:1197–1221.

Berger TW, Yeckel MF, Thiels E (1997) Network determinants of hip-pocampal synaptic plasticity. In: Long-term potentiation, Vol 3 (BaudryM, Davis JL, eds), pp 233–267. Cambridge, MA: MIT.

Berry SD, Thompson RF (1978) Prediction of learning rate from thehippocampal electroencephalogram. Science 200:1298–1300.

Blanpied TA, Berger TW (1992) Characterization in vivo of the NMDAreceptor-mediated component of dentate granule cell population syn-aptic responses to perforant path input. Hippocampus 2:373–388.

Bliss TVP, Collingridge GL (1993) A synaptic model of memory—long-term potentiation in the hippocampus. Nature 361:31–39.

Christian EP, Deadwyler SA (1986) Behavioral functions and hip-pocampal cell types: evidence for two nonoverlapping populations inthe rat. J Neurophysiol 55:331–348.

Claiborne BJ, Amaral DG, Cowan WM (1986) A light and electronmicroscope analysis of the mossy fibers of the rat dentate gyrus. J CompNeurol 246:435–458.

Claiborne BJ, Xiang Z, Brown TH (1993) Hippocampal circuitry com-plicates analysis of long-term potentiation in mossy fiber synapses.Hippocampus 3:115–122.

Collingridge GL, Herron CE, Lester RAJ (1988) Frequency-dependentN-methyl-D-aspartate receptor-mediated transmission in rat hippocam-pus. J Physiol (Lond) 399:301–312.

Davies SN, Lester RA, Reymann KG, Collingridge GL (1989) Tempo-rally distinct pre- and post-synaptic mechanisms maintain long-termpotentiation. Nature 338:500–503.

Derrick BE, Martinez JL (1994) Frequency-dependent associative long-term potentiation at the hippocampal mossy fiber-CA3 synapse. ProcNatl Acad Sci USA 91:10290–10294.

Eichenbaum H, Wiener SI, Shapiro ML, Cohen NJ (1989) The organi-zation of spatial coding in the hippocampus: a study of neural ensembleactivity. J Neurosci 9:2764–2775.

Foster TC, McNaughton BL (1991) Long-term potentiation of CA1 syn-aptic transmission is due to increased quantal size, not quantal content.Hippocampus 1:79–91.

Freeman JA, Nicholson C (1975) Experimental optimization of currentsource-density technique for anuran cerebellum. J Neurophysiol38:369–382.

Gaarskjaer FB (1978) Organization of the mossy fiber system of the ratstudied in extended hippocampi. I. Terminal area related to number ofgranule and pyramidal cells. J Comp Neurol 178:49–72.

Harris EW, Cotman CW (1986) Long-term potentiation of guinea pigmossy fiber responses is not blocked by N-methyl-D-aspartate antago-nists. Neurosci Lett 70:132–137.

Ishizuka N, Weber J, Amaral DG (1990) Organization of intrahip-pocampal projections originating from CA3 pyramidal cells in the rat.J Comp Neurol 295:580–623.

Kelso SR, Ganong AH, Brown TH (1986) Hebbian synapses in thehippocampus. Proc Natl Acad Sci USA 83:5326–5330.

Landfield PW, Deadwyler SA, eds (1988) Long-term potentiation: frombiophysics to behavior. New York: Liss.

Langdon RB, Johnson JW, Barrionuevo G (1993) Asynchrony of mossyfibre inputs and excitatory postsynaptic currents in rat hippocampus.J Physiol (Lond) 472:157–176.

Larson J, Lynch G (1986) Induction of synaptic potentiation in hippocam-pus by patterned stimulation involves two events. Science 232:985–988.

Lorente de No R (1933) Studies on the structure of cerebral cortex. II.Continuation of the study of the ammonic system. J Psychol Neurol45:381–438.

Malenka RC, Kauer JA, Zucker RS, Nicoll RA (1988) Postsynapticcalcium is sufficient for potentiation of hippocampal synaptic transmis-sion. Science 242:81–84.

McNaughton BL, Douglas RM, Goddard GV (1978) Synaptic enhance-ment in fascia dentata: cooperativity among coactive afferents. BrainRes 157:277–293.

Morris RGM (1989) Synaptic plasticity and learning: selective impair-ment of learning in rats and blockade of long-term potentiation in vivoby the N-methyl-D-aspartate receptor antagonist AP5. J Neurosci9:3040–3057.

Munoz MD, Nunez A, Garcia-Austt E (1991) Frequency potentiation ingranule cells in vivo at theta frequency perforant path stimulation. ExpNeurol 113:74–78.

O’Keefe J (1979) A review of the hippocampal place cells. Prog Neuro-biol 13:419–439.

Otto T, Eichenbaum H, Weiner SI, Wible CG (1991) Learning-relatedpatterns of CA1 spike trains parallel stimulation parameters optimal forinducing hippocampal long-term potentiation. Hippocampus 1:181–192.

Ranck Jr JB (1973) Studies on single neurons in dorsal hippocampalformation and septum in unrestrained rats. Part I. Behavioral correlatesand firing repertoires. Exp Neurol 41:461–531.

Rose GM, Dunwiddie TV (1986) Induction of hippocampal long-termpotentiation using physiologically patterned stimulation. Neurosci Lett69:244–248.

Steward O, Scoville SA (1976) Cells of origin of entorhinal corticalafferents to the hippocampus and fascia dentate of the rat. J CompNeurol 169:347–370.

Swanson LW, Wyss JM, Cowan WM (1978) An autoradiographic studyof the organization of intrahippocampal association pathways in the rat.J Comp Neurol 181:681–716.

Tamamaki N, Nojyo Y (1993) Projection of the entorhinal layer II neu-rons in the rat as revealed by intracellular pressure-injection of neuro-biotin. Hippocampus 3:471–480.

Wiener SI, Paul CA, Eichenbaum H (1989) Spatial and behavioral cor-relates of hippocampal neuronal activity. J Neurosci 9:2737–2763.

Wigstrom H, Gustafsson B (1983) Facilitated induction of hippocampallong-lasting potentiation during blockade of inhibition. Nature301:603–604.

Wigstrom H, Gustafsson B, Huang YY (1986) Mode of action of exci-tatory amino acid receptor antagonists on hippocampal long-lastingpotentiation. Neuroscience 17:1105–1115.

Williams SH, Johnston D (1991) Kinetic properties of two anatomicallydistinct excitatory synapses in hippocampal CA3 pyramidal neurons.J Neurophysiol 66:1010–1020.

Witter MP (1993) Organization of the entorhinal hippocampal system -a review of current anatomical data. Hippocampus 3:33–44.

Yeckel MF, Berger TW (1990) Feedforward excitation of the hippocam-pus by afferents from the entorhinal cortex: redefinition of the role ofthe trisynaptic pathway. Proc Natl Acad Sci USA 87:5832–5836.

Yeckel MF, Berger TW (1995a) Characterization of mossy fiber-evokedresponses in the CA3 subfield, in vivo. Soc Neurosci Abstr 21:1437.

Yeckel MF, Berger TW (1995b) Monosynaptic excitation of hippocam-pal CA1 pyramidal cells by afferents from the entorhinal cortex. Hip-pocampus 5:108–114.

Zalutsky RA, Nicoll RA (1990) Comparison of two forms of long-termpotentiation in single hippocampal neurons. Science 248:1619–1624.


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Spatial Distribution of Potentiated Synapses in