Neonatal Handling and Environmental Enrichment ... - Semantic Scholar

3 downloads 0 Views 1MB Size Report
neonatal handling and/or environmental enrichment leads to enduring effects (their mag- nitude frequently depending upon the rat line) on those behaviors.
Behavior Genetics, Vol. 27, No. 6, 1997

Neonatal Handling and Environmental Enrichment Effects on Emotionality, Novelty/Reward Seeking, and Age-Related Cognitive and Hippocampal Impairments: Focus on the Roman Rat Lines A. Fernandez-Teruel,1,3 R. M. Escorihuela,1 B. Castellano,2 B. Gonzalez,2 and A. Tobena1

Roman high- and low-avoidance (RHA/Verh and RLA/Verh) rats are selected and bred for extreme divergence in two-way active avoidance acquisition. In addition, compared to RLA/Verh rats, RHA/Verh rats are (behaviorally and physiologically) less anxious or reactive to stressors, show increased novelty (sensation)-seeking behavior as well as a higher preference for rewarding substances, and are usually less efficient in learning tasks not involving shock administration. The present article reviews evidence showing that neonatal handling and/or environmental enrichment leads to enduring effects (their magnitude frequently depending upon the rat line) on those behaviors. For example, it has been found that neonatal handling reduces most of the (behavioral and physiological) signs of emotionality/anxiety in RLA/Verh rats, while environmental enrichment increases their novelty seeking (also the case with RHA/Verh rats), saccharin and ethanol intake, and sensitivity to amphetamine. Finally, initial results (currently being further elaborated upon) support a preventive action of both environmental treatments on agerelated impairments in learning a spatial, water maze task as well as on hippocampal neuronal atrophy. KEY WORDS: Roman high- and low-avoidance rats; reactivity to stress; anxiety; novelty/reward seeking; neonatal handling; enriched environment; age-related deficit; spatial learning; hippocampus.

perform about 60% avoidances after a single 50trial session in the shuttle box, while RLA/Verh animals achieve 10% (or less) avoidance performance (Escorihuela et al., 1995a). In successive 50trial training sessions (at 24-h intervals), RHA/Verh rats accomplish 90% avoidances by the second session (i.e., after 100 trials), whereas RLA/Verh rats need approximately 8 sessions (400 trials) to approach 50% avoidance performance (Escorihuela et al., 1995a). These studies suggest that the well-known "warm-up" effect in shuttlebox avoidance is extremely marked in RLA/Verh rats during the initial trials of each consecutive session, probably as a consequence of their strong, genetically based predisposition toward conditioned

THE ROMAN/Verh RAT LINES

The Swiss sublines of Roman high- and lowavoidance (RHA/Verh and RLA/Verh, respectively) rats have been selectively bred for rapid (RHA/Verh) versus extremely poor (RLA/Verh) acquisition of two-way, active (shuttle-box) avoidance learning (Driscoll and Battig, 1982). Recent studies have shown that RHA/Verh rats usually 1 1Medical

Psychology Unit, School of Medicine, Autonomous University of Barcelona, 08193 Bellaterra, Barcelona, Spain. 2 Histology Unit, School of Medicine, Autonomous University of Barcelona, 08193 Bellaterra, Barcelona, Spain. 3 To whom correspondence should be addressed. Telephone: 34-3-581 23 78/34-3-581 23 77. Fax: 34-3-581 14 35. e-mail: [email protected].

513 0001-8244/97/1100-0513$12.50/0 © 1997 Plenum Publishing Corporation

514

Fernandez-Teruel, Escorihuela, Castellano, Gonzalez, and Tobena

fear responses. Accordingly, little between-session progress (in terms of avoidance responses) has usually been observed during those initial trials, even in sessions in which RLA/Verh rats already show more than 50% avoidances in the last 20 trials (e.g., after the fourth or fifth 50-trial session) (see Escorihuela et al., 1995a). One of the most prominent features resulting from psychogenetic selection of the Roman/Verh rat lines has been their divergent emotionality profiles, with RLA/Verh rats being more reactive to stressors than RHA/Verh rats (Aubry et al., 1995; Castanon and Mormede, 1994; Driscoll and Battig, 1982; Fernandez-Teruel et al., 199la; Ferre et al., 1995a; Gentsch et al., 1981, 1982; Roozendaal et al., 1992; Steimer et al., 1997a,b; Tobena et al., 1996; Walker et al., 1989; for further references see Escorihuela et al., 1995a). Such findings are also in agreement with similar results observed in other rat strains selected for extreme differences in shuttle-box avoidance acquisition (see Brush, 1991). In addition, compared to the RLA/Verh line, RHA/Verh rats may be considered to be higher novelty (sensation) seekers (Fernandez-Teruel et al., 1992a; Lipp, 1979; Siegel, 1997; Siegel and Driscoll, 1996), and accordingly, they show a higher preference for rewarding substances (Razafimanalina et al., 1996) and display higher behavioral sensitivity to psychostimulants as well as stronger mesolimbic dopaminergic responses to these drugs (Corda et al., 1997; Driscoll et al., 1986, 1990; Giorgi et al., 1997; Haney et al., 1994). The present article reviews the main findings of a series of studies dealing with the effects of two environmental treatments (i.e., neonatal handling and environmental enrichment) on the divergent emotional and novelty-seeking profiles, as well as on some age-related impairments, observed in the Roman/Verh rat lines.

SUMMARY OF EFFECTS OF EARLY (NEONATAL) HANDLING AND ENRICHMENT Emotionality (fearfulness), reactivity to stressors, and exploratory behavior can be enduringly altered by neonatal handling of rats (NH; consisting of removing the pups from the nest and placing

them individually in cages for a few minutes, every day, during the first 21 days of life) and by environmental enrichment (EE; housing the rats, for several weeks/months, in large cages—8—12 animals/cage—containing a variety of objects and "playthings") (for details of our NH and EE procedures see Escorihuela et al., 1994a, 1995b). Generally speaking, NH or EE treatments produce in adult rats, depending upon the treatment and the specific measurements, a decrease in emotionality-related measures [such as defecation, freezing, and/or endocrine responses to novelty/stressors (Daly, 1973; Denenberg, 1975; Escorihuela et al., 1994a; Ferre et al., 1995b; Garbanati et al., 1983; Klein et al., 1994; Levine, 1957; Levine et al., 1967; Meaney et al., 1988; Nunez et al., 1996)], increased activity/exploration in novel situations (Fernandez-Teruel et al., 1990; Escorihuela et al., 1994a,b; Levine et al., 1967; Nunez et al., 1995; Renner, 1987; Renner and Rosenzweig, 1987; Widman and Rosellini, 1990), and increased learning efficiency or improved ability to cope with aversively motivated tasks (e.g., Cooper and Zubek, 1958; Diamond, 1988; Escorihuela et al., 1994a,b; Greenough et al., 1990; Meaney et al., 1988; Mohammed et al., 1993; Nunez et al., 1995; Renner and Rosenzweig, 1987; Wong, 1972). It is worth mentioning, however, that NH positive effects have been more consistently observed in behavioral and endocrine measures related to emotional reactivity, while its effects on tasks with a low involvement of emotional factors have been much more controversial (Daly, 1973; Denenberg, 1975; Escorihuela et al., 1994b). Conversely, EE positive effects have usually appeared more clearly, although not exclusively, in the latter type of tasks, especially in learning procedures (Escorihuela et al., 1994a,b; Garbanati et al., 1983; Klein et al., 1994; Renner and Rosenzweig, 1987). Neonatal handling has also been found to reduce hypothalamus-pituitary-adrenal (HPA) axis responses to stressors (Levine et al., 1967; Meaney et al., 1988, 1991; Nunez et al., 1996), and possibly as a consequence of that, it also increases hippocampal glucocorticoid receptors and prevents the cognitive impairments and hippocampal neuronal loss associated with aging (Meaney et al., 1988, 1991; Sapolsky, 1992). On the other hand, exposure of animals to an enriched environment leads to an enhancement of brain growth and increases in the density of neu-

Neonatal Handling and Enrichment Effects

rons and synaptic contacts, as well as in the size of neuronal nuclei and the number of dendritic branches (Diamond, 1988; Greenough et al., 1990; Kempermann et al., 1997). A possible relationship between the mechanisms of those two treatments (i.e., NH and EE) has been suggested by the findings that both NH and EE increase hippocampal NGF (nerve growth factor) and glucocorticoid receptor expression (Meaney et al., 1988; Mohammed et al., 1993; Pham et al., 1997; Sapolsky, 1992). Thus, the effects of experience in infancy and/or adolescence (and even through adulthood, as may be the case with EE treatment) have been consistently found in a number of behavioral and biological variables, using several stocks of rats and mice. It has also been known for many years that the extent and type of consequences of early stimulation treatments are at least partly dependent upon genetic constitution (e.g., King, 1958; Levine and Broadhurst, 1963; Levine and Wetzel, 1963). However, the use of bidirectionally selected and bred animals in studies of early stimulation (i.e., NH or EE) has been much less frequent, even though the use of such animals can provide a control of the genetic bases and stability of contrasting behaviors.

EFFECTS OF NEONATAL HANDLING ON EMOTIONAL REACTIVITY AND NOVELTY-SEEKING BEHAVIOR IN THE ROMAN/Verh RAT LINES In collaboration with Dr. P. Driscoll (ETHZentrum, Zurich, Switzerland), a series of studies on the effects of neonatal handling (NH) and/or enrichment (EE) on the behavior of (hypoemotional) RHA/Verh rats and (hyperemotional) PvLA/Verh rats was initiated several years ago, the results of which are summarized in Tables I and II. In a number of "spontaneous activity/exploration" tests (involving novelty) it was consistently found that NH treatment induced both short-term and long-lasting significant decreases of emotional behavior, as indicated by a reduced defecation frequency, as well as an increase in both exploratory activity and frequency of entry into illuminated compartments [both in the "hexagonal tunnel labyrinth" and in the "black/white box" tests (e.g., Fernandez-Teruel et al., 199la, 1992b;

515 Tobena et al., 1996)] and an increased latency to start (and shorter time spent) self-grooming (Fernandez-Teruel et al., 199la, 1992a,b, 1993; Steimer et al., 1997b; Tobena et al., 1996). These NH effects were usually more marked in the more fearful RLA/Verh line, thus frequently leading to significant "rat line X treatment" interactions (e.g., Fernandez-Teruel et al., 199la, 1992a,b). Neonatal handling effects were also measured on behavioral tests measuring anxiety or fear, such as a fear-conditioning situation and the hyponeophagia test, which have shown that RLA/Verh rats are more anxious/fearful than are RHA/Verh animals (Ferre et al., 1995a). NH reduced defecations in the fear-conditioning situation, especially in RLA/Verh rats (Fernandez-Teruel, 1995; Fernandez-Teruel et al., 1997), and it also reduced selfgrooming behavior and the "latency to start eating" in the hyponeophagia test (Tobena et al., 1996). Again, NH effects were more prominent in RLA/Verh rats than in their RHA/Verh counterparts, so that neonatally handled RLA/Verh rats achieved scores similar to those of control (nonhandled) RHA/Verh animals (Fernandez-Teruel et al., 1997; Tobena et al., 1996). Of special interest was the finding that neonatal handling improved shuttle-box avoidance acquisition in 18-month-old RLA/Verh rats, while not affecting the performance of RHA/Verh animals (Escorihuela et al., 1995a). Considering the wellknown emotionality-reducing effects of neonatal handling, and that shuttle-box avoidance acquisition is improved by anxiolytics and impaired by anxiogenic drugs (Fernandez-Teruel et al., 1991b), those NH effects appear to be consistent with the previously mentioned reduction of emotionality/anxiety observed in other studies after NH treatment. Some support for this contention is provided by the fact that NH effects in RLA/Verh rats were observed mainly during the initial trials of some training sessions, i.e., during the training phases presumably involving the highest levels of anxiety or conditioned fear (Escorihuela et al., 1995a; Fernandez-Teruel et al., 1991b). Another recent finding has been the observation that NH reduces HPA-axis (corticosterone) and prolactin responses to stressors only in the RLA/Verh line, thus abolishing the differences between both rat lines, especially regarding the stressor-induced prolactin response (see Steimer et al., 1997a,b; Tobena et al., 1996). This is a particularly

516

Fernandez-Teruel, Escorihuela, Castellano, Gonzalez, and Tobena

Table I. Summary of Effects of Neonatal Handling (NH) on Behaviors Related to Emotionality/ Anxiety, Reactivity to Stress, and Novelty Seeking in RHA/Verh and RLA/Verh Ratsa Effects of NHb Differences between the lines

RHA

RLA

(A) Emotionality and anxiety measures New cage Defecation Hexagonal tunnel labyrinth Activity Entries into the lit center Defecation Open field Activity Defecation Black/white (BAV)boxc Initial B/W crossing latency Self-grooming latency Defecation Hyponeophagia test Latency to start eating Time spent self-grooming Defecation Fear conditioning Defecation Shuttle-box avoidance acquisition Hormonal responses to stress ACTH Corticosteroned Prolactin Preference for new objects or spaces Hole-board exploration (new objects) a

RHA < RLA

(16,17)

|

||

(10,11)

RHA > RLA RHA > RLA RHA < RLA

(2,12,14,18,19)

| | |

ft ft ||

(12,14,15)

RHA > RLA RHA < RLA

(1,3,4,12,13,16, 17,24)

= =|

= |

RHA < RLA RHA > RLA RHA = RLA

(24,26)

= ft |

=| | ||

(25,26)

RHA < RLA RHA < RLA RHA < RLA

=

(9,16,23,26)

=

|| || =|

(9,26)

RHA < RLA RHA > RLA

(10,21,22,29) (5,7,9)

| |

(10,11) (9)

RHA RHA RHA (B) RHA

=

< RLA < RLA (1,3,4,5,17,24, < RLA 26,27,28) Measures of novelty (sensation) seeking > RLA (10,11)

RHA > RLA

(10,11,13)

= =

= =

=

(13)

=

| |

(25,26)

=

|

||

(10,11)

=

|

(10,13)

Symbols in the table indicate the magnitude and direction of the effects found in the different studies, referenced as numbers in parentheses. =, no differences; < or >, differences (in the direction indicated) that are not reliable among studies; =| or =|, slight but nonsignificant decrease or increase; | or |, significant decrease or increase; || or ||, marked and significant decrease or increase. Numbers in parentheses refer to References: (1) Aubry et al., 1995; (2) Battig, 1983; (3) Castanon and Mormede, 1994; (4) Castanon et al., 1994; (5) Castanon et al., 1995; (6) Chaouloff et al., 1994; (7) Driscoll and Battig, 1982; (8) Driscoll et al., 1986; (9) Escorihuela et al., 1995a; (10) Fernandez-Teruel, 1995; (11) Fernandez-Teruel et al., 1997; (12) Fernandez-Teruel et al., 199 la; (13) Fernandez-Teruel et al., 1992a; (14) Fernandez-Teruel et al., 1992b; (15) Fernandez-Teruel et al., 1993; (16) Ferre et al., 1995a; (17) Gentsch et al., 1982; (18) Martin et al., 1982; (19) Nil and Battig, 1981; (20) Razafimanalina et al., 1996; (21) Roozendaal et al., 1992; (22) Roozendaal et al. 1993; (23) Shephard and Broadhurst, 1983; (24) Steimer et al., 1997a; (25) Steimer et al., 1997b; (26) Tobena et al., 1996; (27) Walker et al, 1989; (28) Walker et al., 1992; (29) Wilcock and Bush 1972; (30) results currently in preparation. b Animals receiving neonatal handling were placed individually in plastic cages twice daily (morning and evening) for 10 min; after the first 5 min and at the end of the session, each pup was gently handled for 3 s. Handling treatment started on postnatal day 1 and finished on postnatal day 21. Nonhandled animals were left undisturbed until weaning (postnatal day 22). c Chauloff et al. (1994) have reported opposite results in the B/W box by using different experimental procedures (i.e., testing started by placing each rat in the center of the white compartment). d Differences between studies in corticosterone measures are reviewed in the present issue by Steimer et al. (1997b).

Neonatal Handling and Enrichment Effects

517

Table II. Summary of Effects of Environmental Enrichment (EE) on Behaviors Related to Emotionality, Anxiety, and Novelty/ Reward Seeking in RHA/Verh and RLA/Verh Ratsa Effects of EEb Differences between the lines

RHA

RLA

| |

| =

(13)

| = |

= | = |

(10,13,30)

| =

| |

(A) Emotionality and anxiety measures Open field Activity Defecation Hole board Defecation Self-grooming Elevated plus-maze (dim light; dark phase)c Total arm entries Open arm entries Social interaction test Active social interaction Preference for new objects and spatial configurations Hole board Exploration of new objects Saccharin intake (free choice) Ethanol intake (free choice) Stereotyped behavior after amphetamine administration

RHA > RLA RHA < RLA

(1,3,4,12,13,16, 17,24)

RHA < RLA RHA < RLA

(10-13,30)

RHA > RLA RHA > RLA

(6,30; see Table

III)

RHA > RLA (6,30) = (B) Measures related to novelty (sensation) seeking RHA > RLA

(10,30)

RHA > RLA RHA > RLA RHA > RLA

(10,11,13) (11,20) (11,20; see Fig. 1)

RHA > RLA

(8,11,30; see Fig. 2)

(30; see Table

=

(30)

ft

(10,30)

|

ft | |

(10,11,13) (11) (11)

|

|

(11,30)

|| ft =

III)

a

See Table I, footnote a, for symbols and reference numbers (in parentheses). Animals receiving environmental enrichment were placed in groups of 10-12 or 7-9 (depending on the experiment) in cages (100X43X50 cm) containing several objects and "playthings" which were changed every 2 days. The internal configuration of the cages was also changed every 2 days, creating different spaces with several types of stairs, ropes, tunnels, and so forth. Enrichment treatment was administered for a period of 6 months and started when the animals were 6 weeks old. Nonenriched animals were housed in pairs, in standard macrolon cages. c This experiment was performed under dim red light during the dark phase of the light-dark cycle. b

notable result, as prolactin reactivity to stressors is related to the divergent emotionality shown by both rat lines (Castanon and Mormede, 1994; Gentsch et al., 1982; Steimer et al., 1997a; Tobena et al., 1996), and it also appears to be genetically linked to the shuttle-box avoidance differences characterizing the Roman/Verh rats (Castanon et al., 1995). As described in the first section above, RHA/Verh rats typically show a greater amount of novelty (sensation)-seeking behavior than do RLA/Verh rats, as reflected by increased exploration of (and preference for) novel objects and spaces (or new spatial arrangements) and, more importantly, by enhanced exploration of new (unknown) objects in the hole-board test. The latter has been widely recommended as a test for specific

exploratory behavior (independent of general locomotor activity) in rats (Abel, 1995; FernandezTeruel, 1995; Fernandez-Teruel et al., 1992a; File and Wardill, 1975; Sara et al., 1995). Neonatal handling has also been found to enhance both short- and long-term novelty-seeking-related behaviors, these effects being relatively more marked in RLA/Verh rats (Fernandez-Teruel et al., 1992a; unpublished results). To summarize the effects described in the present section, it can be concluded that NH treatment generally has short- and long-term emotionality or anxiety-reducing effects in both Roman/Verh rat lines, while enhancing novelty seeking behavior. Such effects are frequently (and relatively) stronger in RLA/Verh than in RHA/Verh rats (see Table I).

518

Fernandez-Teruel, Escorihuela, Castellano, Gonzalez, and Tobena

Table III. Mean ± SE of Behavioral Measures in the Elevated Plus-Maze Shown by Control (C) and Enriched (EE) RHA/Verh and RLA/Verh Ratsa RLA

RHA C

Total entries Open-arm entries Time in open arms Rearings (enclosed arms)

18.6 5.4 54.2 20.4

± ± ± ±

0.8* 0.7 8.0 2.8

13.2 4.7 55.3 30.4

± ± ± ±

EE

C

EE 0.8 0.6 8.0 1.5**

6.4 2.5 114.0 9.8

± ± ± ±

1.2 0.5** 35.6 2.3**

10.0 4.4 62.4 18.8

± ± ± ±

2.0 0.9 14.0 5.5

a

Each rat was placed facing an enclosed arm and tested for 5 min in the elevated plus-maze during the dark phase of the lightdark cycle and under very dim (red) light conditions. Animals were approximately 8 months old. Data analysis indicated overall significant line effects in total entries [ANOVA; F(l,33)=42.91, p < .01; distance, F(1,59) = 14.93, p < .001]. N = 5-10 animals/group. Young (Y) animals were 5 months old, whereas all the remaining groups were 24 months old. (*) p < .05 vs. the corresponding C group (same line); (+)P< .05 vs. all groups; (C) p < .05 between the groups indicated (Duncan's test).

provements induced by neonatal handling, as well as the prevention of age-related cognitive impairments, have also been related to the ability of that treatment to prevent the hippocampal damage (i.e., neuronal atrophy) frequently associated with aging (Meaney et al., 1988, 1991; Pham et al., 1997; Sapolsky, 1992). Accordingly, 1 month after being tested in the MWM, the rats used here were sac-

CA-3

CA-4

RHA/Verh rats ± — (caudal) +++ +++ ± + + RLA/Verh ± ++++ + + +

_ + rats — +++ + ± -

FD

Hylus

+ (inner)

+ ++ +

(inner) + +

± (caudal) ++++ + + +

± + + ++

+ ± -

Rats (3-6/group) were anesthetized with pentobarbital (50 mg/kg i.p.) and fixed by perfusion transcardially with 4% paraformaldehyde in 0.1 M phosphate buffer (pH 7.4). The brains were removed and postfixed in the same solution for 2 h at 4°C. Brains were embedded in paraffin and cut into 6um coronal sections, collected in parallel series, and stained with hematoxylin-eosin for the demonstration of hippocampal cytoarchitecture and atrophic neurons (for the procedure see Bancroft and Stevens, 1996). The table shows a representative summary of the qualitative evaluation of several sections (from medial to caudal hippocampus) for each experimental group and for regions CA-1, CA-2, and CA-4, the FD (fascia dentata), and the hylus. (— ) No rat from this group showed signs of neuronal atrophy in any section corresponding to this particular region. (±) Some few neurons with signs of atrophy appeared occasionally in some rats or sections. (+, + + , + + + , + + + +) All rats showed atrophic neurons. The number of + signs indicates the relative amount of atrophic neurons observed. Caudal indicates that the effect observed was restricted to the caudal part of those areas. Inner refers to an effect restricted to the inner part of the granular layer.

rificed and hippocampal sections produced for observation of cytoarchitecture and neuronal atrophy. An initial, qualitative histological evaluation of regions CA1, CA3, and CA4, the fascia dentata, and the hilus (Table IV and Fig. 4) revealed that, while young untreated rats showed generally normal neurons, aged untreated rats from both lines showed a considerable degree of cytoarchitectural disorganization with neuronal atrophy and glial reactivity (Gonzalez et al., 1994). Conversely, a marked reduction of such neuronal changes, across all the regions studied, was observed in aged ani-

522

Fernandez-Teruel, Escorihuela, Castellano, Gonzalez, and Tobena

Fig. 4. Representative microphotographs (original magnification, X290), from RLA/Verh rats, of the effects observed in both rat lines across different hippocampal regions. These examples show the neuronal morphology in the CA-1 region [bregma, —3.60 (Paxinos, 1986)] from the different experimental groups: Y, young, untreated rats; C, aged, untreated rats; NH, aged, neonatally handled rats; EE, aged, environmentally enriched rats; HE, aged, both treatments. The arrowhead shows a neuron with normal morphological features, in contrast with atrophic neurons (arrow) displaying a dystrophic morphology with discernible retraction of the cell body and increased eccentricity. The greater amplitude of the HE microphotograph (original magnification, XI16), is to clearly show more the wider CA-1 area and the preventive effects of the treatments.

Neonatal Handling and Enrichment Effects

mals that received any of the environmental treatments (i.e., NH, EE, or HE; Table IV) regardless of rat line. NH-, EE-, and HE-treated rats actually displayed a hippocampal cytoarchitecture and neuronal morphology very similar to that seen in young, untreated rats. Even this cursory histological evaluation appears to indicate changes in hippocampal morphology that are consistent across both rat lines and across the three environmental treatments. It is therefore tempting to speculate that treatment-induced changes in the hippocampal function of aged rats could play a role in differences observed in spatial learning performance (Fiala et al., 1978; Hargreaves et al., 1992; Meaney et al., 1988, 1991; Mohammed et al., 1993; Paylor et al., 1992; Pham et al., 1997; Sapolsky, 1992; Sharp et al., 1987). CONCLUSIONS The evidence reviewed here indicates that RHA/Verh rats are less emotional/fearful, more novelty/reward seeking, and less efficient in some learning tasks (not involving shock) than are their RLA/Verh counterparts. Importantly, most of the genetically based behavioral and physiological criteria examined have been found to be enduringly affected by ontogenetic factors, such as neonatal handling and/or environmental enrichment, frequently leading to the abolishment of between-line differences and to "treatment X rat line" interactions. Pending a more detailed quantitative analysis, the histological observations of the hippocampus of aged treated and untreated rats from both lines show at least two clear-cut patterns: (1) aged rats from either line show a pronounced degree of neuronal atrophy (across the different hippocampal areas), compared to young, untreated rats; and (2) aged rats from both lines receiving any of the treatments (NH, EE, or HE) display a permanent hippocampal cytoarchitecture and neuronal morphology similar to that observed in young, untreated rats. This appears to coincide with the prevention of age-related spatial learning impairments observed in the treated groups. Current and future research is focusing on the possible changes that NH and EE could induce in neuroregulatory systems presumably involved in the behavioral and physiological effects of those treatments, e.g., cholinergic transmission, gluco-

523

corticoid receptors, mesolimbic dopaminergic function, and GABAa/benzodiazepine neurotransmission. ACKNOWLEDGMENTS This work was supported by the FISss (95/1779) and CIRIT (GRQ 94-2005). The continuous and invaluable collaboration of Dr. Peter Driscoll is gratefully acknowledged. We are also grateful to M. A. Martil for his important collaboration in the histological part of the work.

REFERENCES Abel, E. (1995). Further evidence for the dissociation of locomotor activity and head-dipping in rats. Physiol. Behav. 57:529-532. Aubry, J. M., Bartanusz, V., Driscoll, P., Schulz, P., Steimer, T., and Kiss, J. Z. (1995). Corticotropin-releasing factor (CRF) and vasopressin (VP) mRNA levels in Roman high- and low-avoidance (RHA-RLA) rats; Response to open field exposure. Neuroendocrinology 61:89-97. Bancroft, I. D., and Stevens, A. (1996). Theory and Practice of Histological Techniques, Curchill-Livingstone, Hong Kong. Battig, K. (1983). Spontaneous tunnel maze locomotion in rats. In Zbinden, G., et al. (eds.), Application of Behavioral Pharmacology in Toxicology, Raven Press, New York, pp. 15-26. Bliss, T. V. P., and Collingridge, G. L. (1993). A synaptic model of memory: long-term potentiation in the hippocampus. Nature 361:31-39. Brush, F. R. (1991). Genetic determinants of individual differences in avoidance learning: Behavioral and endocrine characteristics. Experientia 47:1008-1019. Castanon, N., and Mormede, P. (1994). Psychobiogenetics: Adapted tools for the study of the coupling between behavioral and neuroendocrine traits of emotional reactivity. Psychoneuroendocrinology 19:257-282. Castanon, N., Dulluc, J., Le Moal, M., and Mormede, P. (1994). Maturation of the behavioral and neuroendocrine differences between the Roman rat lines. Physiol. Behav. 55:775-782. Castanon, N., Perez-Diaz, F., and Mormede, P. (1995). Genetic analysis of the relationships between behavioral and neuroendocrine traits in Roman High and Low avoidance rat lines. Behav. Genet. 25:371-384. Chaouloff, F., Castanon, N., and Mormede, P. (1994). Paradoxical differences in animal models of anxiety among the Roman rat lines. Neurosci. Lett. 182:217-221. Cooper, R. M., and Zubek, J. P. (1958). Effects of enriched and restricted early environments on the learning ability of bright and dull rats. Can J. Psychol. 12:159-164. Corda, M. G., Lecca, D., Piras, G., Di Chiara, G., and Giorgi, O. (1997). Biochemical parameters of dopaminergic and GABAergic neurotransmission in the CNS of Roman high-avoidance and Roman low-avoidance rats. Behav. Genet. 27:527-536. Daly, M. (1973). Early stimulation of rodents: A critical review of present interpretations. Br. J. Psychol. 64:435—460. D'Angio, M., Serrano, A., Driscoll, P., and Scatton, B. (1988). Stressful environmental stimuli increase extracellular DOPAC levels in the prefrontal cortex of hypoemotional (Ro-

524

Fernandez-Teruel, Escorihuela, Castellano, Gonzalez, and Tobena

man high-avoidance) but not hyperemotional (Roman low-avoidance) rats. An in vivo voltametric study. Brain Res. 451:237-247. Dawson, G. R., and Tricklebank, M. D. (1995). Use of the elevated plus maze in the search for novel anxiolytic agents. Trends Pharmacol. Sci. 16:33—36. Delhi, F., Piazza, P. V., Mayo, W., Le Moal, M., and Simon, H. (1996). Novelty-seeking in rats. Biobehavioral characteristics and possible relationship with the sensationseeking trait in man. Neuropsychobiology 34:136-145. Denenberg, V. H. (1975). Effects of exposure to stressors in early life upon later behavioural and biological processes. In Levi, L. (ed.), Society, Stress, and Disease: Childhood and Adolescence, Vol. 2, Oxford University Press, New York, pp. 269-281. Denenberg, V. H., Woodcock, J. M., and Rosenberg, K. M. (1968). Long-term effects of preweaning and postweaning free-environment experience on rats problem-solving behavior. J. Comp. Physiol. Psychol. 66:533-535. Diamond, M. C. (1988). Enriching Heredity, Free Press, New York. Driscoll, P., and Battig, K. (1982). Behavioral, emotional and neurochemical profiles of rats selected for extreme differences in active, two-way avoidance performance. In Lieblich, I. (ed.), Genetics of the Brain, Elsevier Biomedical Press, Amsterdam, pp. 95-123. Driscoll, P., Lieblich, I., and Cohen, E. (1986). Amphetamineinduced stereotypic responses in Roman high- and lowavoidance rats. Pharmacol. Biochem. Behav. 24:13291332. Driscoll, P., Dedek, J., D'Angio, M., Claustre, Y., and Scatton, B. (1990). A genetically-based model for divergent stress responses: behavioral, neurochemical and hormonal aspects. Adv. Anim. Breed. Genet. 5:97-107. Driscoll, P., Ferre, P., Fernandez-Teruel, A., Levi de Stein, M., Wolfman, C., Medina, J., Tobena, A., and Escorihuela, R. M. (1995). Effects of prenatal diazepam on two-way avoidance behavior, swimming navigation and brain levels of benzodiazepine-like molecules in male Roman high- and low-avoidance rats. Psychopharmacology 122: 51-57. Escorihuela, R. M., Tobena, A., and Fernandez-Teruel, A. (1994a). Environmental enrichment reverses the detrimental action of early inconsistent stimulation and increases the beneficial effects of postnatal handling on shuttlebox learning in adult rats. Behav. Brain Res. 61:169-173. Escorihuela, R. M., Tobena, A., and Fernandez-Teruel, A. (1994b). L'estimulacio infantil. Efectes de I 'ambient primerenc i I'herencia sobre I'emotivitat i I'aprenentatge, Servei de Publicacions de la Universitat Autonoma de Barcelona, Barcelona. Escorihuela, R. M., Tobena, A., Driscoll, P., and FernandezTeruel, A. (1995a). Effects of training, early handling and perinatal flumazenil on shuttle box acquisition in Roman low-avoidance rats: Toward overcoming a genetic deficit. Neurosci. Biobehav. Rev. 19:353-367. Escorihuela, R. M., Tobena, A., and Fernandez-Teruel, A. (1995b). Environmental enrichment and postnatal handling prevent spatial learning deficits in aged hypoemotional (Roman high-avoidance) and hyperemotional (Roman low-avoidance) rats. Learn. Memory 2:40-48. Escorihuela, R. M., Fernandez-Teruel, A., Tobena, A., Vivas, N. M., Marmol, F., Badia, A., and Dierssen, M. (1995c). Early environmental stimulation produces long-lasting changes on beta-adrenoceptor transduction system. Neurobiol. Learn. Memory 64:49-57. Fernandez-Teruel, A. (1995). Influencing genetic differences in emotional reactivity through environmental manipula-

tions. Paper presented at the Conference Jacques Monod "Genetics, Neurogenetics and Behavior II," La Londeles-Maures, France. Fernandez-Teruel, A., Escorihuela, R. M., Jimenez, P., and Tobena, A. (1990). Infantile stimulation and perinatal administration of Ro 15-1788: Additive anxiety-reducing effects in rats. Eur. J. Pharmacol. 191:11-114. Fernandez-Teruel, A., Escorihuela, R. M., Driscoll, P., Tobena, A., and Battig, K. (1991a). Infantile (handling) stimulation and behavior in young Roman high- and low-avoidance rats. Physiol. Behav. 50:563-565. Fernandez-Teruel, A., Escorihuela, R. M., Nunez, J. F., Zapata, A., Boix, F., Salazar, W., and Tobena, A. (1991b). The early acquisition of two-way (shuttle-box) avoidance as an anxiety-mediated behavior: Psychopharmacological validation. Brain Res. Bull. 26:173-176. Fernandez-Teruel, A. Escorihuela, R. M., Nunez, J. F., Goma, M., Driscoll, P., and Tobena, A. (1992a). Early stimulation effects on novelty-induced behavior in two psychogenetically-selected rat lines with divergent emotionality profiles. Neurosci. Lett. 137:185-188. Fernandez-Teruel, A., Escorihuela, R. M., Driscoll, P., Tobena, A., and Battig, K. (1992b). Differential effects of early stimulation and/or perinatal flumazenil treatment in young low- and high-avoidance rats. Psychopharmacology 108: 170-176. Fernandez-Teruel, A., Driscoll, P., Escorihuela, R. M., Tobena, A., and Battig, K. (1993). Postnatal handling, perinatal flumazenil, and adult behavior of the Roman rat lines. Pharmacol. Biochem. Behav. 44:783-789. Fernandez-Teruel, A., Escorihuela, R. M., Driscoll, P., Gil, L., Aguilar, R., and Tobena, A. (1997). Effects of early experience and pharmacological treatments on stress reactions and novelty seeking in the Roman rat lines. Paper presented at the meeting Stress of Life. Stress and Adaptation from Molecules to Man, Budapest, Hungary. Ferre, P., Fernandez-Teruel, A., Escorihuela, R. M., Driscoll, P., Corda, M. G., Giorgi, O., and Tobena, A. (1995a). Behavior of the Roman/Verh high- and low-avoidance rat lines in anxiety tests: Relationship with defecation and self-grooming. Physiol. Behav. 58:1209-1213. Ferre, P., Nunez, J. F., Garcia, E., Tobena, A., Escorihuela, R. M., and Fernandez-Teruel, A. (1995b). Postnatal handling reduces anxiety as measured by emotionality rating and hyponeophagia tests in female rats. Pharmacol. Biochem. Behav. 51:199-203. Fiala, B. A., Joyce, J. N., and Greenough, W. T. (1978). Environmental complexity modulates growth of grannule cells dendrites in developing but not adult hippocampus of rats. Exp. Neurol. 59:372-383. File, S., and Wardill, A. G. (1975). Validity of head-dipping as a measure of exploration in a modified hole-board. Psychopharmacologia 44:53-59. Garbanati, J. A., Sherman, G. F., Rosen, G. D., Hofmann, M., Yutzey, D. A., and Denenberg, V. H. (1983). Handling in infancy, brain laterality and muricide in rats. Behav. Brain Res. 7:351-359. Gentsch, C., Lichtsteiner, M., and Feer, H. (1981). Locomotor activity, defecation score and corticosterone levels during an openfield exposure: A comparison among individually and group-housed rats, and genetically selected rat lines. Physiol. Behav. 27:183-186. Gentsch, C., Lichtsteiner, M., Driscoll, P., and Feer, H. (1982). Differential hormonal and physiological responses to stress in Roman high- and low-avoidance rats. Physiol. Behav. 28:259-263. Giorgi, O., Orlandi, M., Escorihuela, R. M., Driscoll, P., Lecca, D., and Corda, M. G. (1994). GABAergic and do-

Neonatal Handling and Enrichment Effects paminergic transmission in the brain of Roman highavoidance and Roman low-avoidance rats. Brain Res. 638:133-138. Giorgi, O., Corda, M. G., Carboni, G., Frau, V., and Di Chiara, G. (1997). Effects of cocaine and morphine in rats from two psychogenetically selected lines: A behavioral and brain dialysis study. Behav. Genet. 27:537-546. Gonzalez, B., Castellano, B., Vela, J., Fabregas, P., Dalmau, I., Escorihuela, R. M., Tobena, A., and Fernandez-Teruel, A. (1994). Enrichment and early handling protect against age-related deficits. A behavioral and histological study in RHA/Verh and RLA/Verh rats. In Coleman, P. D., Mora, F., and Nieto-Sampedro, M. (eds.), Workshop on Deterioration, Stability and Regeneration of the Brain During Normal Aging, Edicion Peninsular, Madrid, pp. 61-63. Greenough, W. T., Winters, G. S., and Wallace, C. S. (1990). Morphological changes in the nervous system arising from behavioral experience: What is the evidence that they are involved in learning and memory? In Squire, L. R., and Lindenlaub, E. (eds.), The Biology of Memory, Symposia Medica Hoechst 23, Schattauder Verlag, Stuttgart/New York, pp. 159-185. Griebel, G., Moreau, J.-L., Jenck, F., Martin, J. R., and Misslin, R. (1993). Some critical determinants of the behaviour of rats in the elevated plus-maze. Behav. Proc. 29: 37-48. Haney, M., Castanon, N., Cador, M., Le Moal, M., and Mormede, P. (1994). Cocaine sensitivity in Roman High and Low Avoidance rats is modulated by sex and gonadal hormone status. Brain Res. 645:179-185. Hargreaves, E. L., Boon, F., and Cain, D. P. (1992). Rats housed in a complex environment exhibit greater hilar LTP than individually housed littermates. Soc. Neurosci. Abstr. 18:344. Kempermann, G., Kuhn, H. G., and Gage, F. H. (1997). More hippocampal neurons in adult mice living in an enriched environment. Nature 386:493-495. King, J. A. (1958). Parameters relevant to determining the effects of early experience upon adult behavior of animals. Psychol. Bull. 55:46-58. Klein, S. L., Lambert, K. G., Durr, D., Schaefer, T., and Waring, R. E. (1994). Influence of environmental enrichment and sex on predator stress response in rats. Physiol. Behav. 56:291-297. Levine, S. (1957). Infantile experience and resistance to physiological stress. Science 126:405—406. Levine, S., and Broadhurst, P. L. (1963). Genetic and ontogenetic determinants of adult behavior in the rat. J. Comp. Physiol. Psychol. 56:423-428. Levine, S., and Wetzel, A. (1963). Infantile experiences, strain differences, and avoidance learning. J. Comp. Physiol. Psychol. 56:879-881. Levine, S., Haltmeyer, G. C., Karas, G. G., and Denenberg, V. H. (1967). Physiological and behavioral effects of infantile stimulation. Physiol. Behav. 2:55—59. Lipp, H. P. (1979). Differential hypothalamic self-stimulation behaviour in Roman high-avoidance and low-avoidance rats. Brain Res. Bull. 4:553-559. Martin, J. R., Oettinger, R., Driscoll, P., Buzzi, R., and Battig, K. (1982). Effects of chlordiazepoxide and imipramine on maze patrolling within two different maze configurations by psychogenetically selected lines of rats. Psychopharmacology 78:58-62. Meaney, M. J., Aitken, D. H., Berkel, C., Bhatnagar, S., and Sapolsky, R. (1988). Effects of neonatal handling on agerelated impairments associated with the hippocampus. Science 239:766-768.

525 Meaney, M. J., Mitchell, J. B., Aitken, D. H., Bhatnagar, S., Bodnoff, S. R., Iny, L. J., and Sarrieau, A. (1991). The effects of neonatal handling on the development of the adrenocortical response to stress: Implications for neuropathology and cognitive deficits in later life. Psychoneuroendocrinology 16:85-103. Mohammed, A. H., Henricksson, B. G., Soderstrom, S., Ebendal, T., Olsson, T., and Seckl, J. R. (1993). Environmental influences on the central nervous system and their implications for the aging rat. Behav. Brain Res. 57:183-191. Morris, R. G. M. (1984). Development of a water-maze procedure for studying spatial learning in the rat. J. Neurosci. Methods 11:47-60. Morris, R. G. M., Garrud, P., Rawlins, J. N. P., and O'Keefe, J. (1982). Place navigation impaired in rats with hippocampal lesions. Nature 24:681-683. Nil, R., and Battig, K. (1981). Spontaneous maze ambulation and Hebb-Williams learning in Roman high-avoidance and Roman low-avoidance rats. Behav. Neural Biol. 33: 465-475. Nunez, J. F., Ferre, P., Garcia, E., Escorihuela, R. M., Fernandez-Teruel, A., and Tobena, A. (1995). Postnatal handling reduces emotionality ratings and accelerates two-way active avoidance in female rats. Physiol. Behav. 57:831835. Nunez, J. F., Ferre, P., Escorihuela, R. M., Tobena, A., and Femandez-Teruel, A. (1996). Effects of postnatal handling of rats on emotional, HPA-axis, and prolactin reactivity to novelty and conflict. Physiol. Behav. 60: 1355-1359. Olds, J. L., Anderson, M. L., McPhie, D. L., Staten, L. D., and Alkon, D. L. (1989). Imaging of memory-specific changes in the distribution of protein kinase C in hippocampus. Science 245:866-869. Paxinos, G. (1986). The Rat Brain in Stereotaxic Coordinates, Academic Press, Sydney. Paylor, R., Morrison, S. K., Rudy, J. W., Waltrip, L. T., and Wehner, J. M. (1992). Brief exposure to an enriched environment improves performance on the Morris water task and increases hippocampal cytosolic protein kinase C activity in young rats. Behav. Brain Res. 52:49-59. Pham, T. M., Soderstrom, S., Henricksson, B. G., and Mohammed, A. H. (1997). Effects of neonatal stimulation on later cognitive function and hippocampal nerve growth factor. Behav. Brain Res. 86:113-120. Razafimanalina, R., Mormede, P., and Velley, L. (1996). Gustatory preference-aversion profiles for saccharin, quinine and alcohol in Roman high- and low-avoidance lines. Behav. Pharmacol. 7:78-84. Renner, M. J. (1987). Experience-dependent changes in exploratory behavior in the adult rat (Ratus Norvegicus): Overall activity level and interactions with objects. J. Comp. Psychol. 101:94-100. Renner, M. J., and Rosenzweig, M. R. (1987). Enriched and Impoverished Environments. Effects on Brain and Behavior, Springer-Verlag, New York. Roozendaal, B., Wiersma, A., Driscoll, P., Koolhaas, J. M., and Bohus, B. (1992). Vasopressinergic modulation of stress responses in the central amygdala of the Roman high-avoidance and low-avoidance rat. Brain Res. 596: 35-40. Roozendaal, B., Koolhaas, J. M., and Bohus, B. (1993). Posttraining norepinephrine infusion into the central amygdala differentially enhances later retention in Roman highavoidance and low-avoidance rats. Behav. Neurosci. 107:575-579. Sapolsky, R. M. (1992). Stress, the Aging Brain, and the Mechanisms of Neuron Death, MIT Press, London.

526

Fernandez-Teruel, Escorihuela, Castellano, Gonzalez, and Tobena

Sara, S. J., Dyon-Laurent, C., and Herve, A. (1995). Novelty seeking behavior in the rat is dependent upon the integrity of the noradrenergic system. Cogn. Brain Res. 2:181-187. Sharp, P. E., Barnes, C. A., and McNaughton, B. L. (1987). Effects of aging on environmental modulation of hippocampal evoked responses. Behav. Neurosci. 101:170-178. Shephard, R. A., and Broadhurst, P. L. (1983). Hyponeophagia in the Roman rat strains: Effects of 5-metoxy-N,N-dimethyltryptamine, diazepam, methysergide and the stereoisomers of propanolol. Eur. J. Pharmacol. 95:177-184. Siegel, J. (1997). Augmenting and reducing of visual evoked potentials in high- and low-sensation seeking humans, cats, and rats. Behav. Genet. 27:557-564. Siegel, J., and Driscoll, P. (1996). Recent developments in an animal model of visual evoked potential augmenting/reducing and sensation seeking behavior. Neuropsychobiology 34:130-135. Steimer, Th., Driscoll, P., and Schulz, P. E. (1997a). Brain metabolism of progesterone, coping behaviour and emotional reactivity in male rats from two psychogenetically selected lines. J. Neuroendocrinol. 9:169-175. Steimer, Th., La Fleur, S., and Schulz, P. E. (1997b). Neuroendocrine correlates of emotional reactivity and coping in male rats from the Roman high (RHA/Verh)- and low(RLA/Verh) avoidance lines. Behav. Genet. 27:503-512. Tobena, A., Steimer, Th., Escorihuela, R. M., Nunez, J. F., Ferre, P., Driscoll, P., and Femandez-Teruel, A. (1996). Effects of postnatal handling in Sprague-Dawley, Roman high avoidance/Verh and Roman low avoidance/Verh rats. Soc. Neurosci. Abstr. 22:462. Walker, C. D., Rivest, R. W., Meaney, M. J., and Aubert, M. L. (1989). Differential activation of the pituitary-adrenocortical axis after stress in the rat: Use of two genetically

selected lines (Roman low- and high-avoidance rats) as a model. J. Endocrinol. 123:477-485. Walker, C. D., Aubert, M. L., Meaney, M. J., and Driscoll, P. (1992). Individual differences in the activity of the hypothalamus-pituitary-adrenocortical system after stressors: Use of psychogenetically selected rat lines as a model. In Driscoll, P. (ed.), Genetically Defined Animal Models of Neurobehavioral Dysfunctions, Birkhauser, Boston, pp. 276-296. Walsh, R. N., Budtz-Olsen, O. E., Penny, J. E., and Cummings, R. A. (1969). The effects of environmental complexity on the histology of the rat hippocampus. J. Comp. Neurol. 137:361-366. Wehner, J. M., Sleight, S., and Upchurch, M. (1990). Hippocampal protein kinase C activity is reduced in poor spatial learners. Brain Res. 523:181-187. Widman, D. R., and Rosellini, R. A. (1990). Restricted daily exposure to environmental enrichment increases the diversity of exploration. Physiol. Behav. 47:57-62. Wilcock, J., and Bush, M. A. (1972). Heterosis for punishment-induced inhibition of drinking in laboratory rats. Life Sci. 11:403-412. Willig, F., M'Harzi, M., Bardelay, C., Viet, D., and Delacour, J. (1991). Roman strains as a psychogenetic model for the study of working memory: Behavioral and biochemical data. Pharmacol. Biochem. Behav. 40:7-16. Wong, R. (1972). Infantile handling and associative processes of rats. Br. J. Psychol. 63:101-108. Zeier, H., Battig, K., and Driscoll, P. (1978). Acquisition of DRL-20 behavior in male and female, Roman high- and low-avoidance rats. Physiol. Behav. 20:791-793. Zuckerman, M. (1996). The psychobiological model for impulsive unsocialized sensation seeking: A comparative approach. Neuropsychobiology 34:125-129.