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Facilitation of a Distributed Shuttle-Box Conditioning with Posttraining Intracranial Self-Stimulation in Old Rats
NEUROBIOLOGY OF LEARNING AND MEMORY ARTICLE NO.
67, 254–258 (1997)
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BRIEF REPORT Facilitation of a Distributed Shuttle-Box Conditioning with Posttraining Intracranial Self-Stimulation in Old Rats LAURA ALDAVERT-VERA, DAVID COSTA-MISERACHS, ESTER MASSANE´S-ROTGER, CARLES SORIANO-MAS, PILAR SEGURA-TORRES, AND IGNACIO MORGADO-BERNAL1 Area de Psicobiologia, Departament de Psicobiologia i de Metodologia de les Cie`ncies de la Salut, Universitat Auto`noma de Barcelona, Ap. 46, 08193 Bellaterra Barcelona, Spain
warding intracranial electrical stimulation seems to be a consistent way to facilitate learning and memory processes in young adult animals, but its efficacy in old subjects has not been studied previously. Lateral hypothalamic (LH) intracranial self-stimulation (ICSS) has been shown to improve a variety of learned responses in adult rats (3–5 months old) (Aldavert-Vera, SeguraTorres, Costa-Miserachs, & Morgado-Bernal, 1996; Coulombe & White, 1980, 1982a, 1982b; Major & White, 1978; Segura-Torres, Capdevila-Ortı´s, Martı´Nicolovius, & Morgado-Bernal, 1988; Segura-Torres, Portell-Corte´s, & Morgado-Bernal, 1991). This modulatory effect upon memory processes does not seem to affect all subjects equally, at least under certain conditions. In an experiment performed in our laboratory using shuttle-box conditioning, we have shown that facilitation of 24-h retention by posttraining ICSS was stronger in those subjects that had shown a poor initial level of conditioning (Aldavert-Vera et al., 1996). Similarly, since old animals may show some mnesic difficulties, they may be especially able to take advantage of the facilitatory effect of ICSS treatment observed in young rats. Although we do not know of any experiment conducted to investigate the effects of ICSS on learning and memory processes in old animals, subseizure stimulation of the hippocampal formation (a structure from which ICSS can be obtained; Ursin, Ursin, & Olds, 1966) has been shown to enhance retention of inhibitory and active avoidance in old rats (Soumireu-Mourat, Martinez, Jensen, & McGaugh, 1980). In this context, the present experiment attempts to study the effects of posttraining ICSS on acquisition and long-term retention of two-way active avoidance conditioning in old rats.
Old Wistar rats (16 – 17 months) were trained in a twoway active avoidance task for 5 consecutive days (10 trials/day). Immediately after each training session a lateral hypothalamic intracranial self-stimulation session (ICSS group) or a sham-treatment session (Control group) was given to the animals. Long-term retention was tested 7 days after the last acquisition session. ICSS treatment led to a significant improvement in acquisition. In the long-term retention session the level of avoidance in both groups was similar to that achieved in the last acquisition session, although differences among groups failed to reach statistical significance. These results are compared with those obtained in previous experiments with young adult rats. While ICSS facilitated the process of acquisition in both young and old rats (however, it was much more powerful in young animals), further experiments are needed to elucidate whether this effect is long-lasting in old rats, as occurs in young adult subjects. q 1997 Academic Press
Since aged animals and humans are often afflicted with a decrease in the ability to retain some kinds of newly acquired information (Barnes, 1991; Light, 1991; McEntee & Crook, 1990; Petersen et al., 1992; Powell et al., 1991), submitting elderly subjects to procedures that have been shown to facilitate learning and memory processes is of great interest. In this sense, re1 This work was supported by a DGICYT grant (PM95-0128C03) and a DGU grant (1992). Address correspondence and reprint requests to Ignacio Morgado-Bernal, Area de Psicobiologia, Departament de Psicobiologia i de Metodologia de les Cie`ncies de la Salut, Universitat Auto`noma de Barcelona, Ap. 46, 08193 Bellaterra Barcelona, Spain. Fax: (34/3) 581 23 24. E-mail: [email protected].
254 1074-7427/97 $25.00 Copyright q 1997 by Academic Press All rights of reproduction in any form reserved.
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Subjects. We used 30 naive male Wistar rats, obtained from our laboratory breeding stock, with a mean age of 491.3 days (SD Å 4.7) at the beginning of the experiment and a mean weight of 762.2 g (SD Å 84.03) at the time of surgery. Procedure. Under sodium pentobarbital anesthesia (50 mg/kg, ip), each rat was implanted with a monopolar stainless steel electrode (130 mm) aimed at the lateral hypothalamus (LH) that was placed into the fibers of the medial forebrain bundle (MFB), according to the following stereotaxic coordinates: AP Å 01.8 mm from bregma; L Å 2.0 mm (right hemisphere), and P Å 08.5 mm, with the cranium surface as dorsal reference (Paxinos & Watson, 1986). Once the rats had recovered from surgery (7 days), they were taught to self-stimulate by pressing a lever in a conventional Skinner box (25 1 20 1 20 cm). Electrical brain stimulation consisted of a 0.2s train of 50-Hz sinusoidal waves at intensities between 10 and 200 mA. First, the range of current intensities that would support responding on a continuous reinforcement schedule was established for each rat (shaping session). Only rats in which stimulation produced no convulsive or other abnormal behaviors were included in the experiment. After this ICSS shaping session, rats were randomly distributed into two groups according to the independent variable TREATMENT (ICSS or Control). On 3 consecutive days, rats in the ICSS group were trained in the Skinner box to establish the optimum current intensity of ICSS (EOI sessions). In these sessions the rates of lever-pressing were recorded for successive increases in current intensities. Starting with 10 mA (rms), the current was increased by 10 mA every 2 min. Sessions ended when the rate of leverpressing did not increase ({5 responses) during three consecutive increases in the current or when it decreased by 20% with respect to the response rate for the preceding current intensity. The mean of the two current intensities that resulted in the highest response rate in each of the last two sessions was considered the optimum intensity (OI) of ICSS for each rat. Two days later, all rats were trained in a two-way active avoidance task (Campdem Instruments, Ltd.). The rats were submitted to one daily acquisition session (10 trials) of two-way active avoidance for 5 consecutive days (in a distributed paradigm). The conditioned stimulus was an 80-dB and 1-kHz tone of 3-s duration. The unconditioned stimulus was a 1-mA electrical footshock, presented for 30 s at maximum. The trials followed a variable interval sched-
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ule of 1 min. Immediately before the first conditioning acquisition session, one adaptation session (10 min) consisting of free ambulation in the conditioning box was given to familiarize the rats with the learning conditions (in this 10-min period, the number of crossings was also scored and considered the locomotor activity level in the adaptation session). In addition to the number of avoidance responses made in each conditioning session (considered the level of performance), intertrial crossings were scored (considered the level of locomotor activity in each conditioning session). Immediately following each of the five acquisition sessions, subjects in the ICSS group (n Å 14) received an ICSS session while subjects in the Control group (n Å 16) were placed in the ICSS box for 15 min, with the electrode clip connected but with the lever retracted. Optimum ICSS parameters to obtain facilitation of two-way active avoidance conditioning in 3-month-old rats (Segura-Torres et al., 1991) often produce convulsive or abnormal behavior in old rats (our observations). Therefore, for this experiment we decided to use ICSS parameters (1500 trains at 100% of OI) that were half the minimum (500 trains at 100% of OI) and optimum (2500 trains at 100% of OI) ICSS parameters that facilitate the acquisition of this kind of conditioning in young adult rats (Segura-Torres et al., 1991). Thus, the treatment consisted of one session of ICSS, administered immediately after every acquisition session, in which rats were allowed to bar-press for 1500 trains of ICSS at the subject’s OI. To test the long-term retention of the learned response, the rats received one additional avoidance session (10 trials) 7 days after the last acquisition session. Results. As shown in Fig. 1, the rats treated with ICSS clearly improved in shuttle-box learning, compared with the Control group. The effect of ICSS treatment seems to be progressive, being more evident in the last sessions. Even though the course of learning, especially from the second acquisition session, showed a general significant linear progressive upward tendency [MANOVA polynomial contrast (first degree); F(1, 28) Å 51.81, p õ .001], some differences were observed between groups [MANOVA polynomial contrast (first degree), Group 1 Session factor; F(1, 28) Å 6.41, p Å .017]. Thus, the ICSS group showed a more accented slope than the Control group, as we can deduce from the F and the p values [ICSS group: F(1, 28) Å 44.38; p õ .001; Control group: F(1, 28) Å 11.66; p Å .002]. In this sense, it can be observed that the differences be-
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The mean numbers of crossings made by the rats during the shuttle-box adaptation session and during the acquisition and LTR sessions (intertrial crossings) are shown in Fig. 2. As can be observed, the ICSS group made significantly more crossings than the Control group in both the adaptation session [t test: F(1, 28) Å 10.59, p Å .003] and the conditioning sessions [MANOVA: F(1, 28) Å 5.69, p Å .024]. Nevertheless, these differences could not explain the observed variations in learning, since correlational analyses showed that the locomotor activity in the shuttle-box was never related to the level of conditioning of the subjects. There were no statistical differences between groups in changes in body weight during the experiment (MANOVA).
FIG. 1. Effects of ICSS treatment on the evolution of avoidance conditioning throughout the acquisition and long-term retention (7 days) sessions. Results are expressed as means ({SEM).
tween groups increased progressively over the consecutive sessions, reaching their peak at the fifth acquisition session [MANOVA, F(1, 28) Å 5.32, p Å .029]. With reference to long-term retention, a MANOVA analysis showed no significant changes in the number of avoidances from the last acquisition session to the LTR session in ICSS and Control groups. Thus, the treatment effects seemed to be maintained after a LTR period. However, as can be seen in Fig. 1, the mean number of avoidances in the LTR session tended to decrease slightly in the ICSS group and to increase in the Control group with respect to the last acquisition session. These slight tendencies caused differences among groups in the LTR session that did not reach statistical significance. The mean OI of ICSS in the ICSS group was 108.21 mA (SD Å 67.19; min, 50 mA; max, 122 mA), and the mean time taken by the rats to complete 1500 bar-presses was 45 min 18 s (SD Å 22 min 37 s; min, 22 min; max, 99 min). The mean rate of ICSS lever response in all treatment sessions was 161.64 responses/min (SD Å 57.74). Correlational analyses showed that the ICSS parameters (response rates, total number of reinforcements, OI, or duration of both EOI and treatment sessions) of each rat were not related to the level of conditioning achieved in any acquisition or LTR session.
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Discussion. Our results indicate that posttraining ICSS of the LH improved the acquisition of a distributed two-way active avoidance conditioning in old rats. These results are consistent with those obtained in our previous experiments using the same conditioning but with young adult rats (Segura-Torres et al., 1988, 1991). In the present experiment, although both groups maintained the level of avoidance shown in the last acquisition session, differences among groups in the LTR session did not reach statistical significance. Instead, in the previously
FIG. 2. Locomotor activity in shuttle-box (adaptation and conditioning sessions) of the ICSS and Control groups. Results are expressed as means ({SEM).
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mentioned works ICSS facilitated not only acquisition of conditioning, but also long-term retention (10 and 31 days). The lack of facilitation in LTR when the treatment was administered to old rats may be due to several factors. On one hand, since the number of ICSS trains is a critical parameter in the facilitatory effect (Segura-Torres et al., 1991) and in the present experiment we used a number of trains lower than the optimum to obtain facilitation of acquisition and retention (see Procedure), it is possible that independent of the age of the subjects, the treatment was powerful enough to facilitate acquisition but not LTR. In agreement with this explanation, a study comparing the effects of treatments with different quantities of trains of ICSS in young adult rats pointed out that treatments of both 500 and 2500 trains of ICSS facilitated the acquisition of two-way active avoidance, but that only the higher treatment (2500 trains) had facilitatory effects on LTR. In any case, the effect of 500 trains of ICSS on acquisition was less facilitatory than that of 2500 trains (Segura-Torres et al., 1991). Thus we cannot discern whether the lack of effect on LTR is due to the lower facilitatory efficiency of this treatment on acquisition or on its consolidation. On the other hand, it could be that the treatment affects young and old rats differentially; that is, in both cases ICSS may enhance acquisition, but in young rats the changes produced by treatment may be long-lasting, while in old rats the effects may vanish as time goes on. To help determine this, we have compared the results for old rats in the present experiment with the results for a group of young rats from another experiment performed in our laboratory at the same time as the present experiment. Rats in that experiment received a treatment of 2500 trains of ICSS, with a conditioning paradigm and parameters identical to those in the present experiment. Results are shown in Fig. 3. First, we observe that aging does not seem to damage two-way active avoidance, since both control groups (the 3- and the 18-month-olds) showed identical acquisition and retention levels of conditioning. This result is contrary to our initial assumption and to previous studies showing that two-way active avoidance can present age-related deficits (Black, Nadel, & O’Keefe, 1977; Blokland & Jolles, 1994). Thus, it is not clear if active avoidance could be sensitive to hippocampal dysfunction or other dysfunctions related to normal aging (Barnes, 1991). Second, although ICSS treatment caused similar levels of performance in the last acquisition session in both young and old rats, treatment seems to be more powerful in young rats
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FIG. 3. Comparative effects of ICSS treatment on shuttle-box conditioning, on both old and young adult groups of rats. Results are expressed as means ({SEM).
than in old rats. Thus, while in old rats the facilitation was observed only in the fifth acquisition session, in young rats the ICSS effect was statistically significant throughout the entire acquisition and retention sessions [MANOVA, F(1, 57) Å 3.70; p Å .05]. Therefore, the slightest efficacy of ICSS treatment in old rats could also explain the nonmaintenance of its effect on the LTR. Apart from this, the fact that the ICSS group showed higher levels of locomotor activity in the shuttle-box can be explained by the generally heightened sympathetic arousal elicited by ICSS (Burgess, Davis, Wilson, Borg, Burgess, & Buggy, 1993). Thus, this result agrees with other previous results showing that ICSS causes an increase in voluntary locomotor activity (Schwarzberg & Roth, 1989). Even so, the different levels of locomotor activity cannot explain the differences observed in avoidance in the last acquisition session because no relation was found between these two variables in any experimental group. Thus, the facilitation of the acquisition process seems attributable to the ICSS treatment, but not to changes in locomotor activity produced by the ICSS treatment or to other variables, such as weight of ICSS parameters. Moreover, since ICSS treatment is always administered immediately after the training session, performance during the next acquisition session (24 h later) can hardly be influenced directly by the treat-
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ment. Therefore, our results are consistent with the view that the facilitatory effect of posttraining ICSS on acquisition is related to modulation of memory consolidation processes. In this way, the present results seem to support the idea that the generally accepted relationship between activation of the brain reward system and memory facilitation (Destrade & Jaffard, 1978; Segura et al., 1991) could also exist in aged animals. Some authors have found decreases in some neurotransmitters related to learning and memory with age, as dopamine released by brain stem neurons (Wickelgren, 1996). Since ICSS clearly improves dopamine release especially in the mesolimbic system (Nakahara, Ozaki, Miura, Miura, & Nagatsu, 1989), treatment with ICSS in old animals could have positive effects by compensating for dopamine deficiencies. This also implies that the aged brains of these animals still have plastic mechanisms that allow modulation of memory systems. Since in the present experiment conditioning did not show age-related deficits, it would be interesting in future experiments to use a memory task that is clearly impaired in aged animals. Furthermore, more experiments are needed to study the conditions necessary to obtain, if possible, long-lasting facilitatory effects in old rats, such as those seen in young adult subjects. REFERENCES Aldavert-Vera, L., Segura-Torres, P., Costa-Miserachs, D., & Morgado-Bernal, I. (1996). Shuttle-box memory facilitation by posttraining intracranial self-stimulation: Differential effects in rats with high and low basic conditioning levels. Behavioral Neuroscience, 110, 346–352. Barnes, C. A. (1991). Memory changes with age: Neurobiological correlates. In J. L. Martinez, Jr., & R. P. Kesner (Eds.), Learning and memory: A biological view. (pp. 259–296). San Diego: Academic Press. Black, A. H., Nadel, L., & O’Keefe, J. (1977). Hippocampal function in avoidance learning. Psychological Bulletin, 84, 1107– 1129. Blokland, A., & Jolles, J. (1994). Behavioral and biochemical effects of an ICV injection of streptozotocin in old Lewis rats. Pharmacology, Biochemistry and Behavior, 47, 883–887. Burgess, M. L., Davis, J. M., Wilson, S. P., Borg, T. K., Burgess, W. A., & Buggy, J. (1993). Effects of intracranial self-stimulation on selected physiological variables in rats. American Journal of Physiology, 264, R149–R155. Coulombe, D., & White, N. (1980). The effect of post-training
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lateral hypothalamic self-stimulation on aversive and appetitive classical conditioning. Physiology and Behavior, 25, 267– 272. Coulombe, D., & White, N. (1982a). Posttraining self-stimulation and memory: A study of some parameters. Physiological Psychology, 10, 343–349. Coulombe, D., & White, N. (1982b). The effect of post-training hypothalamic self-stimulation on sensory preconditioning in rats. Canadian Journal of Psychology, 36, 57–67. Destrade, C., & Jaffard, R. (1978). Post-trial hippocampal and lateral hypothalamic electrical stimulation: Facilitation on long-term memory of appetitive and avoidance learning tasks. Behavioral Biology, 22, 345–374. Light, L. L. (1991). Memory and aging: Four hypotheses in search of data. Annual Review of Psychology, 42, 333–376. Major, R., & White, N. (1978). Memory facilitation by self-stimulation reinforcement mediated by nigro-neostriatal bundle. Physiology and Behavior, 20, 723–733. McEntee, W. J., & Crook, T. H. (1990). Age-associated memory impairment: A role for catecholamines. Neurology, 40, 526– 530. Nakahara, K., Ozaki, N., Miura, Y., Miura, H., & Nagatsu, T. (1989). Increased dopamine and serotonin metabolism in rat nucleus accumbens produced by intracranial self-stimulation of medial forebrain bundle as measured by in vivo microdialysis. Brain Research, 495, 178–181. Paxinos, G., & Watson, C. (1986). The rat brain in stereotaxic coordinates (2nd ed). Sydney: Academic Press. Petersen, R. C., Smith, G., Kokmen, E., Ivnik, R. J., & Tangalos, E. G. (1992). Memory function in normal aging. Neurology, 42, 396–401. Powell, D. A., Buchanan, S. L., & Herna´ndez, L. L. (1991). Classical (Pavlovian) conditioning models of age-related changes in associative learning and their neurobiological substrates. Progress in Neurobiology, 36, 201–228. Schwarzberg, H., & Roth, N. (1989). Increased locomotor activity of rats by self-stimulation in a running wheel. Physiology and Behavior, 46, 767–769. Segura-Torres, P., Capdevila-OrtıB s, L., MartıB -Nicolovius, M., & Morgado-Bernal, I. (1988). Improvement of shuttle-box learning with pre- and post-trial intracranial self-stimulation in rats. Behavioural Brain Research, 29, 111–117. Segura-Torres, P., Portell-Corte´s, I., & Morgado-Bernal, I. (1991). Improvement of shuttle-box avoidance with post-training intracranial self-stimulation, in rats: A parametric study. Behavioural Brain Research, 42, 161–167. Soumireu-Mourat, B., Martinez, J. L., Jr., Jensen, R. A., & McGaugh, J. L. (1980). Facilitation of memory processes in aged rats by subseizure hippocampal stimulation. Physiology and Behavior, 25, 263–265. Ursin, R., Ursin, H., & Olds, L. (1966). Self-stimulation of hippocampus in rats. Journal of Comparative and Physiological Psychology, 61, 353–359. Wickelgren, I. (1996). For the cortex, neuron loss may be less than thought. Science, 273, 48–50.
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BRIEF REPORT Facilitation of a Distributed Shuttle-Box Conditioning with Posttraining Intracranial Self-Stimulation in Old Rats LAURA ALDAVERT-VERA, DAVID COSTA-MISERACHS, ESTER MASSANE´S-ROTGER, CARLES SORIANO-MAS, PILAR SEGURA-TORRES, AND IGNACIO MORGADO-BERNAL1 Area de Psicobiologia, Departament de Psicobiologia i de Metodologia de les Cie`ncies de la Salut, Universitat Auto`noma de Barcelona, Ap. 46, 08193 Bellaterra Barcelona, Spain
warding intracranial electrical stimulation seems to be a consistent way to facilitate learning and memory processes in young adult animals, but its efficacy in old subjects has not been studied previously. Lateral hypothalamic (LH) intracranial self-stimulation (ICSS) has been shown to improve a variety of learned responses in adult rats (3–5 months old) (Aldavert-Vera, SeguraTorres, Costa-Miserachs, & Morgado-Bernal, 1996; Coulombe & White, 1980, 1982a, 1982b; Major & White, 1978; Segura-Torres, Capdevila-Ortı´s, Martı´Nicolovius, & Morgado-Bernal, 1988; Segura-Torres, Portell-Corte´s, & Morgado-Bernal, 1991). This modulatory effect upon memory processes does not seem to affect all subjects equally, at least under certain conditions. In an experiment performed in our laboratory using shuttle-box conditioning, we have shown that facilitation of 24-h retention by posttraining ICSS was stronger in those subjects that had shown a poor initial level of conditioning (Aldavert-Vera et al., 1996). Similarly, since old animals may show some mnesic difficulties, they may be especially able to take advantage of the facilitatory effect of ICSS treatment observed in young rats. Although we do not know of any experiment conducted to investigate the effects of ICSS on learning and memory processes in old animals, subseizure stimulation of the hippocampal formation (a structure from which ICSS can be obtained; Ursin, Ursin, & Olds, 1966) has been shown to enhance retention of inhibitory and active avoidance in old rats (Soumireu-Mourat, Martinez, Jensen, & McGaugh, 1980). In this context, the present experiment attempts to study the effects of posttraining ICSS on acquisition and long-term retention of two-way active avoidance conditioning in old rats.
Old Wistar rats (16 – 17 months) were trained in a twoway active avoidance task for 5 consecutive days (10 trials/day). Immediately after each training session a lateral hypothalamic intracranial self-stimulation session (ICSS group) or a sham-treatment session (Control group) was given to the animals. Long-term retention was tested 7 days after the last acquisition session. ICSS treatment led to a significant improvement in acquisition. In the long-term retention session the level of avoidance in both groups was similar to that achieved in the last acquisition session, although differences among groups failed to reach statistical significance. These results are compared with those obtained in previous experiments with young adult rats. While ICSS facilitated the process of acquisition in both young and old rats (however, it was much more powerful in young animals), further experiments are needed to elucidate whether this effect is long-lasting in old rats, as occurs in young adult subjects. q 1997 Academic Press
Since aged animals and humans are often afflicted with a decrease in the ability to retain some kinds of newly acquired information (Barnes, 1991; Light, 1991; McEntee & Crook, 1990; Petersen et al., 1992; Powell et al., 1991), submitting elderly subjects to procedures that have been shown to facilitate learning and memory processes is of great interest. In this sense, re1 This work was supported by a DGICYT grant (PM95-0128C03) and a DGU grant (1992). Address correspondence and reprint requests to Ignacio Morgado-Bernal, Area de Psicobiologia, Departament de Psicobiologia i de Metodologia de les Cie`ncies de la Salut, Universitat Auto`noma de Barcelona, Ap. 46, 08193 Bellaterra Barcelona, Spain. Fax: (34/3) 581 23 24. E-mail: [email protected].
254 1074-7427/97 $25.00 Copyright q 1997 by Academic Press All rights of reproduction in any form reserved.
AID
NLM 3760
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6v0b$$$221
04-15-97 10:41:30
nlma
AP: NLM
ICSS AND SHUTTLE-BOX CONDITIONING IN OLD RATS
Subjects. We used 30 naive male Wistar rats, obtained from our laboratory breeding stock, with a mean age of 491.3 days (SD Å 4.7) at the beginning of the experiment and a mean weight of 762.2 g (SD Å 84.03) at the time of surgery. Procedure. Under sodium pentobarbital anesthesia (50 mg/kg, ip), each rat was implanted with a monopolar stainless steel electrode (130 mm) aimed at the lateral hypothalamus (LH) that was placed into the fibers of the medial forebrain bundle (MFB), according to the following stereotaxic coordinates: AP Å 01.8 mm from bregma; L Å 2.0 mm (right hemisphere), and P Å 08.5 mm, with the cranium surface as dorsal reference (Paxinos & Watson, 1986). Once the rats had recovered from surgery (7 days), they were taught to self-stimulate by pressing a lever in a conventional Skinner box (25 1 20 1 20 cm). Electrical brain stimulation consisted of a 0.2s train of 50-Hz sinusoidal waves at intensities between 10 and 200 mA. First, the range of current intensities that would support responding on a continuous reinforcement schedule was established for each rat (shaping session). Only rats in which stimulation produced no convulsive or other abnormal behaviors were included in the experiment. After this ICSS shaping session, rats were randomly distributed into two groups according to the independent variable TREATMENT (ICSS or Control). On 3 consecutive days, rats in the ICSS group were trained in the Skinner box to establish the optimum current intensity of ICSS (EOI sessions). In these sessions the rates of lever-pressing were recorded for successive increases in current intensities. Starting with 10 mA (rms), the current was increased by 10 mA every 2 min. Sessions ended when the rate of leverpressing did not increase ({5 responses) during three consecutive increases in the current or when it decreased by 20% with respect to the response rate for the preceding current intensity. The mean of the two current intensities that resulted in the highest response rate in each of the last two sessions was considered the optimum intensity (OI) of ICSS for each rat. Two days later, all rats were trained in a two-way active avoidance task (Campdem Instruments, Ltd.). The rats were submitted to one daily acquisition session (10 trials) of two-way active avoidance for 5 consecutive days (in a distributed paradigm). The conditioned stimulus was an 80-dB and 1-kHz tone of 3-s duration. The unconditioned stimulus was a 1-mA electrical footshock, presented for 30 s at maximum. The trials followed a variable interval sched-
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ule of 1 min. Immediately before the first conditioning acquisition session, one adaptation session (10 min) consisting of free ambulation in the conditioning box was given to familiarize the rats with the learning conditions (in this 10-min period, the number of crossings was also scored and considered the locomotor activity level in the adaptation session). In addition to the number of avoidance responses made in each conditioning session (considered the level of performance), intertrial crossings were scored (considered the level of locomotor activity in each conditioning session). Immediately following each of the five acquisition sessions, subjects in the ICSS group (n Å 14) received an ICSS session while subjects in the Control group (n Å 16) were placed in the ICSS box for 15 min, with the electrode clip connected but with the lever retracted. Optimum ICSS parameters to obtain facilitation of two-way active avoidance conditioning in 3-month-old rats (Segura-Torres et al., 1991) often produce convulsive or abnormal behavior in old rats (our observations). Therefore, for this experiment we decided to use ICSS parameters (1500 trains at 100% of OI) that were half the minimum (500 trains at 100% of OI) and optimum (2500 trains at 100% of OI) ICSS parameters that facilitate the acquisition of this kind of conditioning in young adult rats (Segura-Torres et al., 1991). Thus, the treatment consisted of one session of ICSS, administered immediately after every acquisition session, in which rats were allowed to bar-press for 1500 trains of ICSS at the subject’s OI. To test the long-term retention of the learned response, the rats received one additional avoidance session (10 trials) 7 days after the last acquisition session. Results. As shown in Fig. 1, the rats treated with ICSS clearly improved in shuttle-box learning, compared with the Control group. The effect of ICSS treatment seems to be progressive, being more evident in the last sessions. Even though the course of learning, especially from the second acquisition session, showed a general significant linear progressive upward tendency [MANOVA polynomial contrast (first degree); F(1, 28) Å 51.81, p õ .001], some differences were observed between groups [MANOVA polynomial contrast (first degree), Group 1 Session factor; F(1, 28) Å 6.41, p Å .017]. Thus, the ICSS group showed a more accented slope than the Control group, as we can deduce from the F and the p values [ICSS group: F(1, 28) Å 44.38; p õ .001; Control group: F(1, 28) Å 11.66; p Å .002]. In this sense, it can be observed that the differences be-
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The mean numbers of crossings made by the rats during the shuttle-box adaptation session and during the acquisition and LTR sessions (intertrial crossings) are shown in Fig. 2. As can be observed, the ICSS group made significantly more crossings than the Control group in both the adaptation session [t test: F(1, 28) Å 10.59, p Å .003] and the conditioning sessions [MANOVA: F(1, 28) Å 5.69, p Å .024]. Nevertheless, these differences could not explain the observed variations in learning, since correlational analyses showed that the locomotor activity in the shuttle-box was never related to the level of conditioning of the subjects. There were no statistical differences between groups in changes in body weight during the experiment (MANOVA).
FIG. 1. Effects of ICSS treatment on the evolution of avoidance conditioning throughout the acquisition and long-term retention (7 days) sessions. Results are expressed as means ({SEM).
tween groups increased progressively over the consecutive sessions, reaching their peak at the fifth acquisition session [MANOVA, F(1, 28) Å 5.32, p Å .029]. With reference to long-term retention, a MANOVA analysis showed no significant changes in the number of avoidances from the last acquisition session to the LTR session in ICSS and Control groups. Thus, the treatment effects seemed to be maintained after a LTR period. However, as can be seen in Fig. 1, the mean number of avoidances in the LTR session tended to decrease slightly in the ICSS group and to increase in the Control group with respect to the last acquisition session. These slight tendencies caused differences among groups in the LTR session that did not reach statistical significance. The mean OI of ICSS in the ICSS group was 108.21 mA (SD Å 67.19; min, 50 mA; max, 122 mA), and the mean time taken by the rats to complete 1500 bar-presses was 45 min 18 s (SD Å 22 min 37 s; min, 22 min; max, 99 min). The mean rate of ICSS lever response in all treatment sessions was 161.64 responses/min (SD Å 57.74). Correlational analyses showed that the ICSS parameters (response rates, total number of reinforcements, OI, or duration of both EOI and treatment sessions) of each rat were not related to the level of conditioning achieved in any acquisition or LTR session.
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Discussion. Our results indicate that posttraining ICSS of the LH improved the acquisition of a distributed two-way active avoidance conditioning in old rats. These results are consistent with those obtained in our previous experiments using the same conditioning but with young adult rats (Segura-Torres et al., 1988, 1991). In the present experiment, although both groups maintained the level of avoidance shown in the last acquisition session, differences among groups in the LTR session did not reach statistical significance. Instead, in the previously
FIG. 2. Locomotor activity in shuttle-box (adaptation and conditioning sessions) of the ICSS and Control groups. Results are expressed as means ({SEM).
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ICSS AND SHUTTLE-BOX CONDITIONING IN OLD RATS
mentioned works ICSS facilitated not only acquisition of conditioning, but also long-term retention (10 and 31 days). The lack of facilitation in LTR when the treatment was administered to old rats may be due to several factors. On one hand, since the number of ICSS trains is a critical parameter in the facilitatory effect (Segura-Torres et al., 1991) and in the present experiment we used a number of trains lower than the optimum to obtain facilitation of acquisition and retention (see Procedure), it is possible that independent of the age of the subjects, the treatment was powerful enough to facilitate acquisition but not LTR. In agreement with this explanation, a study comparing the effects of treatments with different quantities of trains of ICSS in young adult rats pointed out that treatments of both 500 and 2500 trains of ICSS facilitated the acquisition of two-way active avoidance, but that only the higher treatment (2500 trains) had facilitatory effects on LTR. In any case, the effect of 500 trains of ICSS on acquisition was less facilitatory than that of 2500 trains (Segura-Torres et al., 1991). Thus we cannot discern whether the lack of effect on LTR is due to the lower facilitatory efficiency of this treatment on acquisition or on its consolidation. On the other hand, it could be that the treatment affects young and old rats differentially; that is, in both cases ICSS may enhance acquisition, but in young rats the changes produced by treatment may be long-lasting, while in old rats the effects may vanish as time goes on. To help determine this, we have compared the results for old rats in the present experiment with the results for a group of young rats from another experiment performed in our laboratory at the same time as the present experiment. Rats in that experiment received a treatment of 2500 trains of ICSS, with a conditioning paradigm and parameters identical to those in the present experiment. Results are shown in Fig. 3. First, we observe that aging does not seem to damage two-way active avoidance, since both control groups (the 3- and the 18-month-olds) showed identical acquisition and retention levels of conditioning. This result is contrary to our initial assumption and to previous studies showing that two-way active avoidance can present age-related deficits (Black, Nadel, & O’Keefe, 1977; Blokland & Jolles, 1994). Thus, it is not clear if active avoidance could be sensitive to hippocampal dysfunction or other dysfunctions related to normal aging (Barnes, 1991). Second, although ICSS treatment caused similar levels of performance in the last acquisition session in both young and old rats, treatment seems to be more powerful in young rats
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FIG. 3. Comparative effects of ICSS treatment on shuttle-box conditioning, on both old and young adult groups of rats. Results are expressed as means ({SEM).
than in old rats. Thus, while in old rats the facilitation was observed only in the fifth acquisition session, in young rats the ICSS effect was statistically significant throughout the entire acquisition and retention sessions [MANOVA, F(1, 57) Å 3.70; p Å .05]. Therefore, the slightest efficacy of ICSS treatment in old rats could also explain the nonmaintenance of its effect on the LTR. Apart from this, the fact that the ICSS group showed higher levels of locomotor activity in the shuttle-box can be explained by the generally heightened sympathetic arousal elicited by ICSS (Burgess, Davis, Wilson, Borg, Burgess, & Buggy, 1993). Thus, this result agrees with other previous results showing that ICSS causes an increase in voluntary locomotor activity (Schwarzberg & Roth, 1989). Even so, the different levels of locomotor activity cannot explain the differences observed in avoidance in the last acquisition session because no relation was found between these two variables in any experimental group. Thus, the facilitation of the acquisition process seems attributable to the ICSS treatment, but not to changes in locomotor activity produced by the ICSS treatment or to other variables, such as weight of ICSS parameters. Moreover, since ICSS treatment is always administered immediately after the training session, performance during the next acquisition session (24 h later) can hardly be influenced directly by the treat-
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ment. Therefore, our results are consistent with the view that the facilitatory effect of posttraining ICSS on acquisition is related to modulation of memory consolidation processes. In this way, the present results seem to support the idea that the generally accepted relationship between activation of the brain reward system and memory facilitation (Destrade & Jaffard, 1978; Segura et al., 1991) could also exist in aged animals. Some authors have found decreases in some neurotransmitters related to learning and memory with age, as dopamine released by brain stem neurons (Wickelgren, 1996). Since ICSS clearly improves dopamine release especially in the mesolimbic system (Nakahara, Ozaki, Miura, Miura, & Nagatsu, 1989), treatment with ICSS in old animals could have positive effects by compensating for dopamine deficiencies. This also implies that the aged brains of these animals still have plastic mechanisms that allow modulation of memory systems. Since in the present experiment conditioning did not show age-related deficits, it would be interesting in future experiments to use a memory task that is clearly impaired in aged animals. Furthermore, more experiments are needed to study the conditions necessary to obtain, if possible, long-lasting facilitatory effects in old rats, such as those seen in young adult subjects. REFERENCES Aldavert-Vera, L., Segura-Torres, P., Costa-Miserachs, D., & Morgado-Bernal, I. (1996). Shuttle-box memory facilitation by posttraining intracranial self-stimulation: Differential effects in rats with high and low basic conditioning levels. Behavioral Neuroscience, 110, 346–352. Barnes, C. A. (1991). Memory changes with age: Neurobiological correlates. In J. L. Martinez, Jr., & R. P. Kesner (Eds.), Learning and memory: A biological view. (pp. 259–296). San Diego: Academic Press. Black, A. H., Nadel, L., & O’Keefe, J. (1977). Hippocampal function in avoidance learning. Psychological Bulletin, 84, 1107– 1129. Blokland, A., & Jolles, J. (1994). Behavioral and biochemical effects of an ICV injection of streptozotocin in old Lewis rats. Pharmacology, Biochemistry and Behavior, 47, 883–887. Burgess, M. L., Davis, J. M., Wilson, S. P., Borg, T. K., Burgess, W. A., & Buggy, J. (1993). Effects of intracranial self-stimulation on selected physiological variables in rats. American Journal of Physiology, 264, R149–R155. Coulombe, D., & White, N. (1980). The effect of post-training
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