Evoked Potentials from the Dentate Gyrus during Auditory Stimulus Generalization in the Rat 1

Transcription

1 EXPERIMEN1AL NEUROLOGY 71, (1981) Evoked Potentials from the Dentate Gyrus during Auditory Stimulus Generalization in the Rat 1 SAM A. DEADWYLER, MARK O. WEST, AND JOHN H. ROBINSON Department of Physiology and Pharmacology. Bowman Gray School of Medicine. Winston-Salem. North Carolina Received August Averaged evoked potentials were recorded from the outer molecular layer of the denate gyrus in freely moving rats undergoing tests of stimulus generalization during performance of an auditory discrimination task. Results indicated that the two major negative components (N 1 and N 2 ) were reciprocally related to the degree of stimulus generalization as indicated by the behavioral response gradient. The N. component was larger when stimuli were least similar in frequency to the originally trained stimulus, whereas the N 2 component was smaller under those same conditions. Destruction of either the entorhinal cortex or the medial and lateral septal nuclei resulted in differential alterations in both stimulus generalization behavior and components of the averaged potentials from the outer molecular layer. The results suggest differential functional roles for the septal and entorhinal projections to the dentate gyrus of the rodent hippocampal formation. INTRODUCTION In previous publications from this laboratory we described the existence of conditioned, sensory-specific evoked potentials confined to the molecular layer of the dentate gyrus of the hippocampal formation in the freely moving rat (2,3). Auditory evoked potentials of large amplitude (0.5 to 1.0 mv) recorded from this region were detectable after rats were trained on an operantly reinforced stimulus discrimination task. The e experiments showed the averaged evoked potentials (AEPs) to be (i) restricted to the region of perforant path synaptic terminals in the outer molecular layer (OM) and (ii) correlated with the occurrence of granule cell unit discharges during simple and differential discrimination learning. Various behavioral manipulations (extinction and reconditioning) resulted in the elimination Abbreviations: AEPs-averaged evoked potentials; OM-outer molecular layer. 1 Supported by National Science Foundation grant BNS to S.A.D /81/ $02.00/0 Copyright 1981 by Academic Press, Inc. All rights of reproduction in any form reserved.

2 616 DEADWYLER, WEST, AND ROBINSON and reappearance of the OM AEPs. In more recent reports we showed that the OM AEP reflects a two-component neural process which is controlled differentially by the perforant path and septal inputs to the dentate gyrus (5, 20). The following report describes further characteristics of the OM AEP in terms of the changes in these two components which accompany tests of stimulus generalization. METHODS Four male Sprague-Dawley rats 90 to 120 days of age were implanted with stimulating electrodes and a chronic microdrive system which has been described in detail elsewhere (4). Two animals (rats 201 and 203) received only electrode implant surgery; the other two animals were implanted with electrodes but in addition, one received destruction of both the medial and lateral septal nuclei (rat 208) and the other, bilateral destruction of the entorhinal cortex (rat 210). The electrode track penetrated the dentate gyrus through an orientation that was perpendicular to the perforant path fibers in the outer molecular layer (13, 15). Stimulating electrodes were permanently implanted in the angular bundle and contralateral CA3 field of the hippocampus and were utilized to activate perforant path and hippocampal commissural fibers, respectively (1, 2). At the beginning of each recording session the microdrive containing the recording electrode was mounted on the animal and the electrode positioned by physiological criteria in the outer molecular layer of the dentate gyrus (2, 3). Animals food-deprived to 80% normal body weight were placed in a sound-shielded rectangular Plexiglas chamber in which auditory (tone) stimuli, behavioral responses (nosepokes into a photocell beam), and reinforcement (a small drop of water) were automatically programmed such that only behavioral responses emitted in the presence of a designated (training) tone frequency (3.5 khz, 60 db) were reinforced (2). A stimulus generalization procedure (12) in which separate test (ones of different frequencies [4.6 (A), 4.1 (B), 2.9 (C), and 2.4 (D) khz] bracketing the training stimulus frequency was utilized. The four test tones were substituted randomly into the stimulus presentation sequence a total of 50 times (on the average once per three presentations) during each of two daily recording sessions (300 total trials per day). Responding during presentation of the test stimuli was never reinforced. The number of nosepoke responses in the presence of each stimulus (i.e., percentage behavioral response) was utilized as the measure of stimulus generalization. RESULTS Averaged sensory-evoked potentials (AEPs) recorded from the outer molecular (OM) layer of the dentate gyrus during simple operant stimulus

3 STIMULUS GENERALIZATION IN DENTATE GYRUS 617 FIG. 1. Examples of OM AEPs recorded during stimulus generalization test for animal 203. Cond-the averaged evoked potential (AEP) of the outer molecular layer (OM) to the training tone (3.5 khz) during performance of the auditory discrimination task. The four OM AEPs obtained for each test stimulus are shown immediately below the training tone AEP (Test Tones). A-D refer to tones of 4.6,4.1,2.9, and 2.4 khz, respectively. Each OM AEP is the average of 50 tone presentations. Tone onset at dotted line (160 ms). Open and solid circles denote N 1 and N 2 respectively. Calibration, 300 V. discrimination performance were identical in waveform to those reported recently from our laboratory (5, 20). The OM AEP waveform consisted of two distinct negative-going peaks: the N 1 component with onset latency of 20 ms and duration of 15 ms, and the N 2 component with onset latency of 45 to 55 ms and duration of 50 to 60 ms. The N l and N 2 components were shown to vary independently in amplitude with respect to the behavioral significance of the stimulus. N l was shown to be larger in unconditioned animals and during extinction in conditioned animals, whereas the amplitude of N 2 was shown to be large after the establishment of criterion stimulus discrimination behavior and nonexistent in naive or in behaviorally extinguished animals (2, 5). Tests of stimulus generalization were conducted only after at least 10 days of criterion stimulus discrimination behavior (average ratio of unreinforced to reinforced responses was less than two to one) in which OM AEPs to the training stimulus showed a well developed N 2 component with N 1 minimal in amplitude. AEPs to each of the test stimuli and to the training stimulus consisted of averages of 50 trials summed over two daily

4 618 DEADWYLER, WEST, AND ROBINSON FIG. 2. Behavioral response gradient (open triangles) and amplitude profile of N 1 and N 2 components of the averaged evoked potential (AEP) of the outer molecular layer for training tone and each test tone (open and solid dots, respectively) from the two normal animals (203 and 201). Each point represents a percentage of the training tone value. Note that the amplitude of N 1 is designated by the ordinate on the right. Amplitudes were measured from the AEP to each tone. Note symmetry of behavioral response gradient generated by each animal, indicating generalized responding along the frequency dimension to test tones A-D. sessions. Figure 1 shows examples of the OM AEPs to the training stimulus (COND.) and the four test stimuli (A-D) in a normal animal (rat 203). It is clear that unlike the AEP evoked by the training stimulus, which was predominantly N 2, the four test stimuli AEPs contained both negative components (N 1 and N 2 ). In this animal, N 1 appeared at significantly increased amplitudes in the different test OM AEPs, whereas N 2 was reduced in amplitude in response to three of four of these same stimuli. The test stimuli nearest in frequency to the training stimulus (B and C) tended to have smaller N 1 components than the two more dissimilar frequencies (A and D). Figure 2 shows the relationship between the behavioral response gradient and the peak amplitude profiles of both components of the OM

5 STIMULUS GENERALIZATION IN DENTATE GYRUS 619 AEP as a function of the training and test stimulus frequencies for the two normal animals (201 and 203). In general, the amplitude of N l was inversely related to the degree of behavioral responding to the test stimuli for both animals and the amplitude of N 2 (which was maximal to the training stimulus) was reduced in seven of the eight test stimulus presentations. OM AEPs to the test stimuli exhibited waveforms which ranged between those obtained after acquisition and extinction of the behavioral response (5). Test stimulus D elicited the largest N 1 component and smallest N 2 in the two normal animals and resembled an extinction type OM AEP more closely than any of the other test stimuli (Figs. 1D, 2). Test stimuli B and C, on the other hand, produced smaller reductions in N 2 and smaller increases in N 1 in these two normal animals (Figs. 1B, C and 2). This pattern of change in N 1 and N 2 with respect to the behavioral significance of the stimulus was verified in several different stimulus presentation paradigms and is consistent with the pattern generated by differential conditioning procedures (5,20). There were occasional disparities, however, between the changes evoked in OM AEPs by test stimuli and the corresponding behavioral response pattern. In particular, test stimulus A evoked a large N 2 component and larger N 1 in animal 203, but the response pattern was the same as that for tone D, which evoked an N 2 of reduced amplitude and a very large N 1 component (see Fig. 2). There were, accordingly, two major factors which influenced the OM AEP during stimulus generalization testing. First, the large percentage increase in N 1 in test AEPs was in part a direct consequence of the almost complete absence of this component in training AEPs. Second, the smaller percentage decrease in N 2 amplitude evoked by the four test stimuli most likely reflected our previous observations that N 2 was only significantly reduced or eliminated in conditioned animals through comprehensive behavioral extinction procedures which lasted for at least 50 trials (2). The intermittent reinforcement (on the average 30% of the total trials) of behavioral responses to the training stimulus during generalization testing would have counteracted this extinction effect; hence N 2 was not markedly attenuated during presentation of any of the four test stimuli. Effects of Entorhinal and Septal Lesions. Figure 3 shows the behavioral response gradient and OM AEP profile of two animals tested for generalization after either bilateral destruction of the entorhinal cortex (210) or a lesion of the medial and lateral septal nuclei (208). As shown in previous reports from this laboratory (5, 20), destruction of the entorhinal cortex severely reduced or eliminated the N 1 component of the OM AEP. OM AEPs obtained from the animal with the entorhinal lesion (210) during generalization testing also contained only the N 2 component. However, the percentage change in amplitude of the N 2 component to the four test

6 620 DEADWYLER, WEST, AND ROBINSON FIG. 3. Behavioral and amplitude profiles of N 1 and N 2 components from two animals with lesions. Same measures as in Fig. 2. The rat with an entorhinal lesion (210) showed no N 1 component in the averaged evoked potential. N 2 follows the steep asymmetric behavioral response gradient to test tones suggesting a lack of generalization for all frequency dimensions of the test stimuli. Septal lesion animal (208): note behavioral response gradient and reduced N 1 component to test stimuli A and B. The symmetrical relationship of N 1 and N 2 components to the test stimuli was not as pronounced as in normal animals. stimuli was much more drastic and followed closely the behavioral response gradient (Fig. 3). In addition, the behavioral response gradient was much narrower and more asymmetric than that obtained from normal animals, indicating that all four test stimuli were effectively discriminated from the training stimulus. Responses to test stimuli A and B occurred slightly more frequently than those to C and D; however, the number of responses to A was not different from B, and the number of responses to C was also not different from D. Thus, the animal with an entorhinal lesion demonstrated equal stimulus discrimination ability to the two normal animals, but correspondingly showed less stimulus generalization. The N 2 amplitude profile in this animal varied closely with the behavioral response

7 STIMULUS GENERALIZATION IN DENTATE GYRUS 621 gradient and showed the largest percentage change of the four animals tested. Thus, the absence of an entorhinal cortex did not impair the animal's ability to discriminate the training stimulus from any of the four test stimuli. In contrast to the effects of bilateral entorhinal destruction, a lesion that destroyed both the medial and lateral septal nuclei in another animal (208) produced (i) decreased stimulus discrimination between the four test and training stimuli, as indicated by the behavioral response gradient; and (ii) OM AEPs which exhibited markedly disparate N 1 and N 2 components compared with normal animals. Test stimuli to which responses occurred on 90% of the presentations (i.e., which were not discriminated from the training stimulus) elicited N 1 components which were as large as those obtained in normal animals, even though responding was reduced by 80% to the same stimulus (208, tones C and D). Conversely, test stimuli that elicited a significant (30%) reduction in behavioral responding were not associated with increased N 1 components as was observed under similar conditions in normal animals. The N 2 component also showed more variability than in normal animals and did not conform to the behavioral response gradient. The fact that both the N 1 and N 2 components were present in the animal with a septal lesion during stimulus generalization testing confirms our previous finding which indicated that septal lesions do not eliminate either of these components during simple discrimination tasks (5). However, even though both components were present, the size of the N 1 component was largest to the two test stimuli which were least discriminated (behaviorally) from the training stimulus. From these preliminary results it can be stated that the animal with a septal lesion was affected differently from that with an entorhinal lesion in tests of stimulus generalization, and that both lesions produced behavioral and evoked potential correlates of stimulus discrimination which were different from those obtained from normal animals. DISCUSSION The above results provide further evidence for our hypothesis that N 1 and N 2 of the OM AEP reflect the differential activity of two separate anatomic pathways which converge on the cellular elements within the dentate gyrus, and that each of these pathways becomes active under different behavioral circumstances. The generalization tests indicate that both N 1 and N 2 were sensitive to the similarity of each of the test stimuli to the training stimulus. In the two normal animals, test stimuli to which responses occurred less frequently than to the training stimulus (especially test stimuli C and D) elicited larger amplitude N 1 components and reduced

8 622 DEADWYLER, WEST, AND ROBINSON N 2 components. We previously suggested that N 1 is a reflection of the input from the perforant path fibers and is eliminated in animals sustaining bilateral removal of the entorhinal cortex (5, 20). This was confirmed in this study which showed that N 1 was not present in OM AEPs from an animal with an entorhinal lesion during generalization testing. In contrast to normal animals, N 2 amplitude varied directly with behavioral responding and showed significantly more reduction in the animal with an entorhinal lesion. Finally, behavioral differentiation between test stimuli Band C and the training stimulus was equivalent compared with normal animals, suggesting that stimulus discrimination was intact in the animal with the entorhinal lesion. A lesion of the medial and lateral septal nuclei produced alterations in generalization behavior which were different from those produced by entorhinal lesions. Test stimuli were responded to more frequently by the rat with a septal lesion, thus showing reduced stimulus discrimination. Although N 1 and N 2 were present in the animal with a septal lesion and exhibited changes to the test stimuli which were similar in some respects to those obtained in normal animals, these changes were not as closely related to those in the behavioral response pattern. In addition, the flat behavioral generalization gradient in the animal with a septal lesion suggests that the test stimuli were less effectively discriminated from the training stimulus than in normal animals. There are four studies in the literature dealing with the effects of hippocampal destruction on stimulus generalization behavior in rats. Three of those studies showed that damage to the hippocampus resulted in steeper generalization gradients after the animals were retrained on the original stimulus discrimination (7, 8, 18). In contrast, generalization gradients for hippocampal-damaged animals which were not retrained were not as steep as in normal animals (21). The behavioral effects of entorhinal destruction on stimulus generalization in the present study were therefore similar to those reported for retrained animals with hippocampal lesions. The animal with a septal lesion showed a flattened generalization gradient which was similar to those reportedly produced by lesions of the amygdala (8, 18). In the past there have been many suggestions as to the basis for behavioral differences between normal animals and those sustaining damage to the hippocampus and/or its input pathways (6,10,11,14,16,17, 19). In the results mentioned above, the behavioral response profile obtained for stimulus generalization in normal rats and the associated N 1 profile of the OM AEP elicited by the different test stimuli are consistent with the interpretation that the entorhinal cortex routes sensory information into the hippocampus if such sensory input is unfamiliar or

9 STIMULUS GENERALIZATION IN DENTATE GYRUS 623 unexpected (20). Furthermore, lesions of the entorhinal cortex produced a sharpening of the behavioral generalization gradient which reflected less tendency to respond to "unfamiliar" stimuli. As no N 1 component was present in this animal it appears that the entorhinal input was partially responsible for maintaining responding to unfamiliar stimuli in the normal animals. The animal with an entorhinal lesion gave no indication of differential responding to any of the test stimuli, although maintaining good discriminative responding to the training stimulus. It is precisely this steepening of the generalization gradient which results from damage to the hippocampus itself (see above). These factors make it likely that the role of the hippocampus in stimulus generalization behavior requires input from the entorhinal cortex. The amplitude of the N 2 component of the OM AEP varied directly though minimally with the degree of generalization from the training stimulus to the four test stimuli. The slight reduction in amplitude of N 2 corresponded to reduced behavioral responding to the test stimuli in seven of eight tests in two normal animals. We previously showed that N 2 reflects the associative features of the sensory stimulus and its appearance and amplitude is modulated by medial septal connections (2, 5, 20). In the present experiment, a lesion of the medial and lateral septum disrupted the behavioral response gradient such that test stimuli were less discriminable from the training stimulus compared with normal animals. Gaffan (9, 10) showed similar behavioral deficits in stimulus generalization tests of rats and monkeys with lesions of the fornix. Thus, the two functions-recognition and association of sensory stimuliseem to be controlled in the dentate gyrus by two separate major input structures; i.e., the entorhinal cortex and medial septum, respectively. These functions can be distinguished in the normal animal by the N 1 and N 2 components in the OM AEP during tests of stimulus generalization as well as during other manipulations of the saliency of the sensory stimulus. The complex electrophysiologic interactions associated with these two components suggest that much information processing in the hippocampus requires highly refined synaptic inputs which signify features of the sensory stimulus in terms of (i) familiarity or "novelty" and (ii) associative history. An important question remains as to how these features are further represented by the remainder of hippocampal circuitry and subsequently how that representation is translated into a meaningful output message to other brain structures. REFERENCES 1. DEADWYLER, S. A., J. A. WEST, C. W. COTMAN, AND G. S. LYNCH Neurophysiological analysis of the commissural projections to the dentate gyrus of the rat. J. Neurophysiol. 38:

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