Introduction

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Memory pharmacology that reverses memory decline or blocks the effects of past traumatic experience through the use of pharmacological agents has not yet been well characterized. The efficacy of memory therapeutics depends on our understanding of the basic mechanisms that characterize memory itself. Memories are thought to be due to lasting synaptic modifications in the brain. Synaptic modifications within memory traces have been linked to a number of mechanisms that involve multiple and interacting afferent pathways, neurotransmitters, messenger molecules, and gene products (Kornhauser and Greenberg, 1997; Alkon et al., 1998; Paulsen and Moser, 1998; Xiang et al., 1998). Some studies have correlated synaptic modifications, or biophysical and biochemical correlates, with behavioral learning and memory (Alkon et al., 1982, 1992; Bradford and McCabe, 1994; Xiang et al., 1998) and others identified long-lasting synaptic modifications such as long-term potentiation (LTP) or depression that are induced by electrophysiological stimulation (Christie et al., 1994; Teyler et al., 1995). Classical paradigms for induction of LTP or long-term depression typically involve tetanization or low-frequency stimulation of a single glutamatergic pathway, respectively. Memory impairments, such as in Alzheimer's disease, are, however, generally characterized by multiple deficits of neurotransmitters in the brain.

Targeting transmitter synapses in memory-related structures is among the most attractive ways to directly affect reception of relevant signals and how they are processed and stored as memory traces. The importance of cholinergic and GABAergic systems in hippocampus-dependent memory has been well established (Winkler et al., 1995; Paulsen and Moser, 1998). Acetylcholine (ACh) is crucial to attention, learning, and memory (Bartus et al., 1982; Buccafusco et al., 1995; Ohno et al., 1997; Robbins et al., 1997), and the generation of hippocampal  rhythmic activity. Activation of the medial septal afferents, a major cholinergic pathway to the hippocampus (Cooper and Sofroniew, 1996; Kalman et al., 1997), is thought to accompany associative learning (Day et al., 1991; Inglis et al., 1994; Inglis and Fibiger, 1995). Its disruption blocks spatial memory (Winson, 1978; Winkler et al., 1995). One GABAergic interneuron innervates a population of some 1000 pyramidal cells. These interneurons exert significant control on hippocampal network activity and synchronize the firing of pyramidal cells (Buhl et al., 1995; Cobb et al., 1995). However, functional interactions between cholinergic receptor activation and GABA synaptic modifications have remained somewhat obscure. Here, we investigate the physiological conditions underlying synaptic modification and induction of long-term synaptic transformation (LTT) of GABAergic synapses by cholinergic receptor activation. LTT has previously been reported to be induced by associating postsynaptic depolarization with s. pyr tetanization (Collin et al., 1995) or intracellular administration of calexcitin (Sun et al., 1999), a memory signal protein (Alkon et al., 1998). It appears to depend on intracellular Ca²⁺ release, probably from the ryanodine receptors (for review, see Alkon et al., 1998). In the present study, we found that associated activation of heterosynaptic inputs without postsynaptic depolarization transforms GABAergic inhibition to excitation. Furthermore, the GABAergic synaptic transformation effectively switched an excitatory input filter to an excitatory input amplifier, altering direction of signal transmission through the network. The synaptic transformation and the transformed response were blocked or eliminated by acetazolamide and bicuculline (BIC), respectively, suggesting an involvement of HCO₃ flux through the chloride channels.

	Materials and Methods

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Brain Slices. Male Sprague-Dawley rats (125-150 g) were anesthetized with diethyl ether and decapitated. The hippocampal formation was removed and sliced (400 µm) with a McIllwain tissue chopper (Collin et al., 1995; Sun et al., 1999). Slices were maintained in an interface chamber (Medical Systems Corp., Greenvale, NY) at 32°C with continuous perfusion of artificial cerebrospinal fluid (aCSF). aCSF consisted of 125 mM NaCl, 3 mM KCl, 1.3 mM MgSO₄, 2.4 mM CaCl₂, 26 mM NaCHO₃, 1.25 mM NaH₂PO₄, and 10 mM C₆H₁₂O₆.

Electrophysiology. Intracellular recordings were obtained from CA1 pyramidal neurons using glass micropipette electrodes filled with 2 M potassium acetate or 0.1 M potassium methylsulfate, with measured tip resistance in the range 70 to 200 M. Cells that show obvious accommodation (e.g., as shown in Figs. 1e, and 2, e and f), a key characteristic of pyramidal cells, were used in the study. Labeling the recorded cells exhibiting the characteristic with dye has previously revealed that the recorded cells are indeed pyramidal cells (Sun et al., 1999). Signals were amplified, digitized, and stored using AxoClamp-2B amplifier and DigiData 1200 with the P-clamp data acquisition and analysis software (Axon Instruments, Foster City, CA). S. pyr, stratum radiatum, and stratum oriens (s. oriens) were stimulated (about 200 µm from the recording electrode; Fig. 1a) using bipolar electrodes constructed of Teflon-insulated platinum iridium wire (25 µm in diameter, the approximate thickness of s. pyr; FHC Inc., Bowdoinham, ME). Monophasic hyperpolarizing PSPs were elicited by orthodromic single-pulse stimulation of interneurons in s. pyr (Collin et al., 1995). In some experiments, a stimulating electrode (about 400 µm from the other stimulating electrodes when two stimulating electrodes were placed) was also placed in s. oriens to activate cholinergic terminals and evoke ACh release (Cole and Nicoll, 1984), or in stratum radiatum to evoke glutamatergic PSPs. Tetanization of s. pyr consisted of 10 trains, 10 pulses at control intensity (30-60 µA and 50 µs), 100 Hz, and a 0.5-s intertrain interval. Stimulation of s. oriens consisted of single pulses (20-60 µA and 50 µs), delivered at 1 Hz for 30 s.

View larger version (28K): [in this window] [in a new window]   Fig. 1.   Characteristics of the pyramidal stimulation-induced IPSPs. Schematic drawing of the stimulation and recording setup (a). Single-pulse stimulation of s. pyr activates inhibitory afferents, which may be GABAergic terminals of Bas interneurons, to CA1 pyramidal neurons (Pyr) and elicits a monophasic hyperpolarizing synaptic response (b-d; 50 µs with intensities indicated below arrowheads in µA). Pyramidal neurons also receive glutamatergic excitatory input via the Sch collateral pathway and cholinergic afferent from the medial septum/vertical limb of the diagonal band nucleus (MS/DBv) in s. oriens. Single-pulse stimulation of s. pyr (50 µA, 50 µs, arrowhead) at different membrane potentials, during 700-ms steps of hyperpolarizing or depolarizing current injection into the CA1 pyramidal neuron (e). Two representative traces show that the peak to maximum response (the first dashed line for the bottom trace and the second dashed for the top trace) depends on the membrane potential, i.e., the direction of anion flux (f). The relationship between the PSPs and membrane potential in response to the single-pulse s. pyr stimulation of the pyramidal neuron can be described with a straight line with a single reversal potential (g), determined by the least sum squares criterion. Bath kynurenate (500 µM, 20 min) abolishes Sch stimulation-induced EPSPs (control; h) but does not change the s. pyr stimulation-induced IPSPs (control; i).

Drugs and Ligands. Benzolamide (gift from T. H. Maren, University of Florida, Gainesville, FL) was applied into recorded cells (0.1 mM; 0.5 nA, 500 ms at 50% on cycles for 10 min through the recording electrode). Carbachol (CCH), physostigmine (PHY), BIC, acetazolamide, kynurenic acid, atropine, 6-cyano-7-nitroquinoxaline-2,3-dione, and D-()-2-amino-5-phosphonopentanoic acid (AP5) were from Sigma (St. Louis, MO) and were solubilized in aCSF in the noted concentrations and delivered to the slice chamber from an external reservoir.

	Results

Top Abstract Introduction Materials and Methods Results Discussion References

A single-pulse stimulus delivered to s. pyr elicited a hyperpolarizing IPSP (Fig. 1, b-d), resulting, mainly if not exclusively, from activation of the GABAergic inputs from the Basket interneurons (Bas) whose cell bodies and axons are restricted to s. pyr. The response magnitude depended on intensities of stimulation (Fig. 1, b-d), without revealing any obvious depolarizing excitatory postsynaptic potential (EPSP) components within the stimulation intensities used. The monophasic nature of the elicited PSPs was further indicated by evoking the PSPs over a range of membrane potentials, hyperpolarized or depolarized relative to the resting potential in eight neurons. Figure 1e illustrates the PSPs elicited by single-pulse stimulation of s. pyr over a range of membrane potential steps. The PSP reversed at a membrane potential of 78 mV (Fig. 1g), exhibiting a linear relationship between the membrane potentials and the evoked PSPs. The time to peak of the evoked response depended on the direction of the anion flux, with the maximum response at hyperpolarized membrane potentials (chloride efflux), occurring some 20 ms earlier than that evoked at resting membrane potentials (chloride influx; Fig. 1f). No detectable minor PSP components that exhibit a different reversal potential were observed. Bath application of 500 µM kynurenate (20 min), a broad-spectrum competitive antagonist for both N-methyl-D-aspartate (NMDA) and non-NMDA receptors (Collingridge and Lester, 1989; Sun, 1996), effectively abolished EPSPs of CA1 pyramidal cells evoked by stimulation of the Schaffer collateral pathway (Sch; by 95.8 ± 3.2%, n = 8, p < 0.05; Fig. 1h). This concentration of kynurenate produced no significant changes (8.5 ± 0.7 mV prekynurenate versus 8.5 ± 0.8 mV during the application; n = 8, p > 0.05) in the IPSPs evoked by single-pulse s. pyr stimulation (Fig. 1i). Thus, these kynurenate results suggest that the single-pulse s. pyr stimulation did not evoke a significant glutamatergic EPSP component. The IPSPs, however, were blocked by the GABA_A receptor antagonist BIC (by >95%, n = 8, p < 0.05; 1 µM, 30-min perfusion; Fig. 2a), indicating the involvement of GABA_A receptors.

View larger version (32K): [in this window] [in a new window]   Fig. 2.   Associating single-pulse stimulation of s. oriens with pyramidal tetanization induced long-term synaptic transformation. Single-pulse stimulation of s. pyr (50 µA, 50 µs) evokes a BIC-sensitive IPSP (a). Postsynaptic responses of a pyramidal cell (b) to single-pulse stimulation of s. pyr before (control) and 60 min after s. pyr tetanization (s. oriens-s. pyr Tet; 10 trains of 50 µA, 50 µs, 10 pulses/train at 100 Hz, 0.5-s intertrain interval), associated with single-pulse s. oriens stimulation (50 µA, 50 µs) (at the arrow). Tetanization of s. pyr alone (10 trains of 50 µA, 50 µs, 10 pulses/train at 100 Hz, 0.5-s intertrain interval) does not induce long-term transformation, but potentiates IPSPs (c), as shown with representative IPSPs in response to single-pulse stimulation (50 µA, 50 µs) of s. pyr before (control) and 60 min (s. pyr Tet) after s. pyr tetanization. Group data show that long-term synaptic transformation is induced by stimulating cholinergic pathway in s. oriens and tetanization of s. pyr (s. oriens-s. pyr Tet) but is sensitive to atropine (atropine + s. oriens-s. pyr Tet), whereas s. pyr tetanization alone is not effective (d; as means ± S.E.M.). Associative tetanization of s. pyr with s. oriens single-pulse stimulation (f) shifts the relationship between the evoked Bas-CA1 PSP and membrane potentials to the right (h), compared with that of the control (e).

Single-pulse stimulation of s. oriens (1 Hz, 30 s) coincident with tetanization of s. pyr consistently induced LTT (Fig. 2, b and d, 7.1 ± 1.3 versus 7.4 ± 1.2 mV before the associated stimulation, n = 9, p < 0.05). Tetanization of s. pyr was applied near the end of s. oriens stimulation (i.e., coincident with pulses 25-30 of the 30-s s. oriens stimulation). This LTT does not appear to result from a simple blockade of a receptor-channel complex. Rather, it was associated with a shift (Fig. 2, e, f, and g) of the relationship between Bas-CA1 PSPs and membrane potential to the right, and a shift of the reversal potential to more positive potentials (from 78.5 ± 1.1 to 61.4 ± 1.2 mV, n = 9, p < 0.05). No obvious depolarization of the resting membrane potential was observed during and after the associative stimulation. The synaptic transformation induced by s. oriens stimulation-s. pyr tetanization was prevented by atropine (20 µM, n = 6; Fig. 2d), which did not affect IPSPs elicited by single-pulse stimulation of s. pyr. Tetanization of s. pyr alone, however, did not induce LTT, but significantly increased the evoked IPSPs (Fig. 2, c and d; from 7.6 ± 1.2 to 10.7 ± 1.0 mV, n = 5, p < 0.05). Single-pulse stimulation of s. oriens alone for 30 s did not induce LTT (data not shown). Nor was temporal separation (i.e., termination of s. oriens stimulation 10 s before tetanization of s. pyr at the same intensity and frequency) effective in inducing the synaptic transformation in six cells tested (data not shown).

The involvement of ACh in the synaptic transformation was further examined by preceding s. pyr tetanization with a 20-min period of extracellular perfusion of CCH (10 µM), an ACh receptor agonist. Perfusion with 10 µM CCH alone for 20 min in the interface chamber did not depolarize the membrane resting potential (n = 18), consistent with the observation (Muller et al., 1988) that the occurrence of carbachol-induced depolarization of hippocampal neurons depends on the rate at which carbachol concentration is elevated in the tissue. Nor were epileptiform-like activities ever observed in the experimental period. Depolarization was consistently induced when using higher CCH concentrations (>100 µM), submission-type chamber, more rapid perfusion, or longer perfusion in brain slices (Muller et al., 1988). The CCH-s. pyr tetanization paradigm, however, transformed the IPSPs (7.1 ± 0.6 mV) to depolarizing EPSPs (6.9 ± 0.9 mV, n = 9, p < 0.05; Fig. 3) and shifted the reversal membrane potential from 78.2 mV (n = 9) on average to more positive potentials (56 mV on average; n = 9) for >1 h. We also tested the possibility that sufficient ACh is spontaneously released from an endogenous source(s) coincident with tetanization of s. pyr to induce the synaptic transformation. Consistent with this possibility, 20 min after PHY (an anticholinesterase, 20 µM), tetanization of s. pyr (without CCH) repeatedly induced a lasting synaptic transformation (Fig. 3; 6.9 ± 0.4 versus 2.4 ± 0.4 mV, n = 5, p < 0.05; Fig. 3g).

View larger version (20K): [in this window] [in a new window]   Fig. 3.   Carbachol and physostigmine mimic s. oriens stimulation in inducing long-term synaptic transformation. Single-pulse s. pyr stimulation (50 µA, 50 µs at arrowhead) evoked a monophasic IPSP (a) in the hippocampal CA1 pyramidal neuron. Long-term synaptic transformation is induced by simultaneous CCH perfusion and s. pyr tetanization (b). After induction by the CCH-s. pyr tetanization paradigm, the synaptic transformation persists with CNQX (100 µM) and APV (50 µM) perfusion for 30 min (c). In a different cell, single-pulse s. pyr stimulation (50 µA, 50 µs at arrowhead) evoked a monophasic IPSP (d). Long-term synaptic transformation is induced with simultaneous CCH perfusion (10 µM, 20 min) and s. pyr tetanization (e). After induction by the CCH-s. pyr tetanization paradigm, the synaptic response is eliminated with BIC (1 µM) perfusion for 20 min (f). Group data (g) show that long-term synaptic transformation (as means + S.E.M.) induced by a 20-min perfusion with either CCH or PHY and tetanization of s. pyr (10 trains, 10 pulses at 100 Hz, 0.5-s intertrain interval).

In the presence of CNQX (100 µM) and AP5 (50 µM), the synapse was not transformed by the CCH-s. pyr tetanization. To the contrary, the CCH-s. pyr tetanization paradigm consistently and significantly potentiated the IPSP amplitude (by 40 ± 8.8%, n = 4, p < 0.05) in the presence of CNQX and AP5. This latter result is consistent with the observation that s. pyr tetanization without postsynaptic depolarization potentiated IPSPs (Collin et al., 1995). Blocking the NMDA receptor subtype with AP5 alone was effective in preventing the long-term synaptic transformation (by 90.1 ± 8.5%, n = 5, p < 0.05). Blocking the non-NMDA receptor subtype did not, however, appear to be sufficient to prevent the synaptic transformation. The CCH-s. pyr tetanization consistently induced the transformation in the presence of CNQX alone (7.0 ± 0.39 versus 8.3 ± 0.46 mV, n = 3, p < 0.05). However, once the transformation was induced, it was not changed by CNQX (100 µM) and AP5 (50 µM) (n = 4; Fig. 3c). These results suggest that although induction of the synaptic transformation requires activation of NMDA receptors or a network circuit involving activation of NMDA receptors, maintenance of the transformation does not. BIC (a GABA_A receptor antagonist, 1.0 µM), however, eliminated the transformed PSPs (Fig. 3f; n = 6). Elimination of the depolarizing PSPs after the synaptic transformation by BIC suggests that the underlying current largely involves activation of GABA_A receptor-activated channels.

In the presence of acetazolamide (1 µM, 30 min), a blocker of carbonic anhydrase and thus the synthesis of HCO₃ (Staley et al., 1995), the Bas-CA1 IPSPs did not undergo transformation. Thus, these IPSPs were not altered by single-pulse s. oriens-s. pyr tetanization (Fig. 4a; 98.9 ± 2.7%, 30 min after compared with 100% control value, n = 8, p > 0.05). When perfused externally, nonbicarbonate buffer also prevented induction of the synaptic transformation (Fig. 4c; 98.5 ± 3.2%, 30 min after single-pulse s. oriens-s. pyr tetanization compared with 100% control value, n = 7, p > 0.05). Since carbonic anhydrase was previously shown to exist in the pyramidal cells (Pasternack et al., 1993), we injected the membrane-impermeant carbonic anhydrase inhibitor benzolamide into the recorded pyramidal cells. This injection prevented (Fig. 4b; 99.0 ± 2.5%, 30 min after compared with 100% control value, n = 87, p > 0.05) the single-pulse s. oriens-s. pyr tetanization-induced synaptic transformation (Fig. 4b), indicating that the inhibitory effects on carbonic anhydrase were intracellular.

View larger version (20K): [in this window] [in a new window]   Fig. 4.   Acetazolamide and nonbicarbonate buffer eliminate the synaptic transformation and the transformation converts excitatory input filter into amplifier. Bath acetazolamide (1 µM) eliminates s. oriens single-pulse-s. pyr tetanization-induced synaptic transformation (a). Intracellular benzolamide is also effective (b). The synaptic transformation is not induced in HEPES buffer (c). Single-pulse stimulation (d) of Bas-CA1 evokes an IPSP, which is enhanced by s. pyr tetanization alone. After s. pyr tetanization, single-pulse stimulation (at arrowhead) of Bas-CA1 powerfully filters out Sch excitatory inputs (e; s. pyr Tet, costimulation also at arrowhead), reducing the Sch response from an above-action potential threshold (control) to below the threshold. Single-pulse s. oriens-s. pyr tetanization produces Bas-CA1 synaptic transformation (f). Single-pulse stimulation (g) of Sch at below-threshold intensities evokes an EPSP, which is amplified to above threshold after the synaptic transformation by costimulation of the Bas-CA1 inputs. Action potentials are truncated.

In 10 cells, excitatory Sch input was stimulated at intensities 30% above their action potential threshold. S. pyr tetanization alone enhanced the Bas-CA1 inhibition (Fig. 4d) and blocked (100% of 20 trials; n = 10, p < 0.05) the effects of excitatory Sch input, stimulated at the above the action potential threshold intensities (100% of 20 trials; Fig. 4e) in all the 10 cells tested. The effective signal-filtering period in each single-pulse-evoked Bas-CA1 response was 100 ms, during which no action potential (0% of 20 trials) was evoked by Sch stimulation at the above-threshold intensity. After the synaptic transformation (Fig. 4f), below-threshold Sch stimulation that by itself did not evoke action potentials (0% of 20 trials) became sufficient to evoke action potentials (100% of 20 trials; n = 9, p < 0.05) when delivered during the period of 100 ms of single-pulse Bas-CA1 stimulation (Fig. 4 g; n = 9). Multiple spikes were evoked when above-threshold Sch stimulation was delivered after the synaptic transformation (data not shown). Associating single-pulse s. oriens stimulation with the tetanization amplified excitatory Sch inputs (Fig. 4g). Thus, weak signals are amplified in the cells with the synaptic transformation, whereas strong excitatory signals cannot successfully pass through the network under the enhanced Bas inhibition.

	Discussion

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The present study shows, for the first time, that the hyperpolarizing IPSPs of rat hippocampal field CA1 pyramidal neurons in response to s. pyr stimulation undergo a reversal in polarity (LTT) after the temporally associated activation of cholinergic and GABAergic receptors. The associated activation of multisynaptic inputs is at least as effective in LTT induction as the postsynaptic depolarization-s. pyr tetanization paradigm reported previously (Collin et al., 1995). Temporal association of different inputs thus may code neural information, in line with the view that recognition memory is not mediated by a single neurotransmitter type (Steckler et al., 1998). The synaptic reversal was sensitive to carbonic anhydrase inhibitors. The critical role of cholinergic receptor activation on the synaptic transformation was further demonstrated by the effectiveness of atropine and the anticholinesterase PHY. The results are consistent with an in vivo report that carbonic anhydrase inhibition reduces the  rhythm (Sone et al., 1998).

The observed synaptic transformation does not appear to involve a masked excitatory component, such as glutamatergic EPSPs. Microstimulation was delivered to the area remote to major excitatory terminal inputs and the stimulation at the selected intensity did not directly activate the pyramidal cells. Thus, the evoked IPSPs are monophasic and showed little or no change in magnitude with an effective blockade of the glutamatergic receptors. The lack of a depolarizing component of the IPSP was further confirmed by a single reversal potential for s. pyr-elicited responses. Furthermore, the evoked IPSPs and transformed depolarizing PSPs were abolished by blocking the GABA_A receptors, whereas an effective blockade of the glutamatergic receptors was ineffective after the synaptic transformation.

However, induction of the synaptic transformation was prevented by pretreatment with the glutamatergic receptor antagonists AP5 and CNQX, AP5, or kynurenate, but not with CNQX alone. This indicates that the excitatory amino acid L-glutamate is released in the in vitro preparation and its activation on the NMDA receptors is required for induction of the synaptic transformation. Whether the glutamate release is a result of s. pyr tetanization or occurs spontaneously is unknown at this time. It is probable that some spontaneous glutamatergic activity is present in vivo and sufficient for synaptic transformation to occur. Disinhibition of inhibitory inputs might also result in enhanced activity of the principal cells. Another possibility is that the induction may involve activation of a network circuit, including activation of NMDA receptors although its maintenance does not. Response of the recorded cells to activation of a network circuit, if evoked, however, would be delayed and not jeopardize the immediate synaptic response analyzed in the study. Thus, the synaptic transformation induced in our study differs from an effect reported by Kaila et al. (1997) and Taira et al. (1997). In their studies, tetanic Sch stimulation, instead of the s. pyr tetanization/cholinergic activation in the present study, elicits a lasting GABAergic depolarizing PSP.

Acetazolamide, an inhibitor of carbonic anhydrase, was shown to reduce or eliminate flux of HCO₃ in hippocampal pyramidal neurons underlying a depolarizing PSP (Staley et al., 1995). Acetazolamide also prevents the synaptic transformation. Activity of carbonic anhydrase in the CA1 pyramidal cells is essential since intracellular application of benzolamide, a membrane-impermeant carbonic anhydrase inhibitor, effectively blocked the synaptic transformation. The results of the present study are consistent with an induction of a depolarizing transmembrane HCO₃ flux that underlies the synaptic transformation. The relatively brief time course of the transformed PSPs, compared with the time course of IPSPs before the synaptic transformation, suggests that the HCO₃ flux may be limited by its availability. It is possible, then, that during a transformed PSP, the GABA-activated channel(s) becomes more rapidly inactivated. Indeed, the channel appears to operate with different characteristics when conducting an efflux or influx of anion, perhaps reflecting a flux direction-dependent property.

GABAergic postsynaptic depolarizing responses are observed in neonatal brains (Leinekugel et al., 1999) and could be induced by pronged activation of GABA_A receptors (Staley and Proctor, 1999) or the use of neuroactive steroids (Burg et al., 1998) in the adults. The present results demonstrate a persistent heterosynaptic modification induced by synergy of neurotransmitters. Synergy between cholinergic, GABAergic, and glutamatergic activation is suggested for the synaptic transformation, consistent with the observation that in hippocampal field CA1, application of GABA in association with Sch tetanization also produces a transient depolarizing response (Wong and Watkins, 1982). The transformed synaptic inputs from the Bas interneurons are also shown to provide a mechanism to direct or gate signal flow through the hippocampal network. These interneurons innervate the perisomatic region of the pyramidal cells. Thus, bursting activity from the interneurons in the absence of associated cholinergic activity enhances the inhibitory control of the pyramidal cells, powerfully blocking excitatory signal transfer through the hippocampal circuit. With the temporal association of cholinergic activity, however, the same type of GABAergic activity amplifies excitatory signal strength. The differentiation in responses according to the nature and temporal association of relevant signals enables the network to perform signal processing and gate information flow and direction. The occurrence of the switch is controlled postsynaptically. The formation of a functional "center" of attention (those transformed) and a "surround" (those enhanced) would dramatically increase the signal-to-noise ratio. The in vitro synaptic changes analyzed here are consistent with in vivo studies that implicate ACh in arousal, attention, and memory mechanisms (Dickinson-Anson et al., 1998; Perry et al., 1999; Sun et al., 2001). These results suggest, therefore, that attention and arousal (mediated by ACh) and signaling (via the hippocampal trisynaptic circuit from the entorhinal cortex through the dentate gyrus and field CA3 to field CA1) could interact with GABAergic synaptic activation. This activation could, via transformation of IPSPs to EPSPs, selectively amplify synaptic weights relevant to a particular memory, forming the signal center of attention. In this switching cascade, the highly efficient carbonic anhydrase appears to play an important role. Consistent with the role of carbonic anhydrase in hippocampus-dependent memory are observations that administration of a carbonic anhydrase inhibitor in vivo reduces hippocampal theta activity (Sone et al., 1998), which is believed by many to gate or facilitate memory information processing in the hippocampus, and impairs rat spatial watermaze performance (Sun et al., 2001). Agents that inhibit carbonic anhydrase may have clinical value for temporary suppression of traumatic memories, such as in surgery or post-traumatic stress disorder.