BEHAVIORAL AND NEURAL BIOLOGY 25, 268--270
(1979)
BRIEF REPORT Environmental Enrichment and Brain Chromatin LYNDA
UPHOUSE
AND
BRUCE
TEDESCHI
1
Department of Psychology, Yale University, New Haven, Connecticut 06520
Twenty-eight-day-old Fischer rats were assigned to enriched, isolated, or standard environmental rearing for 60 days. Chromatin from cortex, cerebellum, and rest of brain was assayed for capacity to support in vitro RNA synthesis. No significant differences were observed across rearing groups. These results are compared to previous findings after shorter rearing intervals and are discussed in terms of a transient influence of the rearing environment on brain chromatin. Previous experiments have d e m o n s t r a t e d an impact of postweaning rearing environments on brain chromatin (Uphouse, 1978; U p h o u s e & Moore, 1978). When differential rearing continued from 28 to 60 days of age, enriched rats exhibited reduced whole brain chromatin-directed R N A synthesis. H o w e v e r , cortical chromatin from enriched animals supported m o r e R N A synthesis than did chromatin from isolated or standard colony-housed rats, but rest of brain chromatin was elevated in isolated animals. These data suggested that both enrichment and isolation had influenced brain chromatin. When differential rearing was extended from 28 to 88 days of age, whole brain chromatin from the three rearing groups was no longer different; but it remained possible that differences in parts of brain continued to exist. In the present study, parts of brain chromatin was examined from rats differentially reared from 28 to 88 days of age. Sixty male, Fischer inbred rats (Charles Rivers Laboratories, North Wilmington, Mass.) were used in the experiment. After arrival in the laboratory at 28 days of age the rats were assigned in two groups of 30 animals to enriched (EC), isolated (IC), or standard colony (SC) conditions as previously described (Uphouse & Moore, 1978). After 60 days in their respective environments, the rats were sacrificed by decapitation; brains were quickly removed, weighed, and dissected into cortex, rest of brain, and cerebellum as described by U p h o u s e (1978). Tissue was stored Research supported by NSF Grant BNS-74-03883. The authors wish to express appreciation to Ms. Ella FutreU for typing the manuscript. 268 0163- 1047/79/020268-03$02.00/0 Copyright © 1979 by Academic Press, Inc. All 1ights of reproduction in any form reserved.
269
ENVIRONMENT AND BRAIN CHROMATIN
at -85°C until chromatin extraction. Tissue from three to four animals of the same experimental condition was pooled and chromatin was extracted as previously described (Uphouse & Moore, 1978). Chromatin was always prepared in triplets so that matched tissues from enriched, isolated, and standard colony animals were simultaneously analyzed. The capacity for chromatin to support RNA synthesis was measured in the presence of exogenous Escherichia coli RNA polymerase (R-0501, Type II; Sigma Chemical Company), unlabeled ATP, CTP, and GTP (Sigma Chemical Company), and [3H]-UTP [5-3H]uridine 5'triphosphate, 22.7 Ci/mmole, New England Nuclear). Assay conditions and computation of relative RNA synthesis were as previously described (Uphouse & Moore, 1978). The data from individual brain regions were analyzed by one-way ANOVA (Winer, 1971). The relative capacity for chromatin to support RNA synthesis in vitro is shown in Table 1. Although the trend for cortical chromatin differences was in the same direction as that noted in rats differentially reared for only 32 days (Uphouse, 1978), the differences were smaller in magnitude and not as consistent as those observed in previous studies. Analysis of variance did not reveal significant differences among the treatment groups [F(2, 15) = 1.33]. Similarly, rest of brain and cerebellum did not differ across treatment conditions [respectively, F(2, 15) = 1.07; F (2, 12) = 2.12]. Since the chromatin changes previously found in animals differentially reared from 28 to 60 days were no longer present with the longer rearing interval, it is possible that differential rearing may exert only a transient influence on the capacity for chromatin to support in vitro RNA synthesis. However, a conclusive demonstration that the response is not permanent would require a more extensive investigation than employed in the present experiment. Ideally, several durations of differential rearing should be simultaneously investigated. The apparent transience of the part of brain response is, however, in agreement with previous studies of whole brain chromatin (Uphouse & Moore, 1978) and is compatible with the suggestion that the rearing condiTABLE 1 Relative R N A Synthesis by C h r o m a t i n from 88-Day-Old Differentially Reared Rats Rearing condition Part of brain
Chromatin preparations"
EC
IC
SC
Cortex Rest of brain Cerebellum
6 6 5
11.13 ± 1.65 12.47 ~ 0.948 11.84 _ 1.03
10.62 _+ 1.62 12.43 _+ 2.85 10.57 ___ 1.82
12.62 _ 2.85 14.2 - 2.83 12.43 ___ 1.43
T i s s u e from three to four brains was pooled for each chromatin preparation.
270
UPHOUSE AND TEDESCHI
tions influenced a developmental change in chromatin template activity, and that with increased duration in the rearing environment, all animals would eventually reach a similar level. It is equally possible that the animals habituated to their rearing environments. It has been suggested that isolated animals initially exhibit responses characteristic of stress but that, with increasing time in isolation, this response is diminished (Geller, 1971). Alternatively, enriched animals may habituate to the novel or arousal-producing aspects of the enriched environment. The apparent transience of the chromatin response observed both in whole brain and in parts of brain is in sharp contrast to the relatively permanent morphological changes seen in cortex after environmental enrichment (Rosenzweig, Bennett, & Diamond, 1972). H o w e v e r , it is difficult to ascertain what proportion of the morphological differences actually occur during the longer rearing durations. The continuance of morphological differences, even with much longer durations than those utilized in the present experiment, may reflect the maintenance of morphological changes which occurred earlier during differential rearing rather than their continued induction. Chromatin-directed R N A synthesis may be more responsive to variations in the environment (e.g., arousal, novelty, or stress) which diminish with increased time in the rearing environments. While such a response may be important for the facilitation of morphological c h a n g e s , it may not be essential for their maintenance. F u r t h e r m o r e , it is important to note that the in vitro m e a s u r e m e n t of R N A synthesis is a quantitative m e a s u r e m e n t and the lack of a difference does not preclude the presence of more subtle macromolecular changes. Such changes with longer rearing durations have been previously suggested ( U p h o u s e & Bonnet, 1975; Grouse, Schrier, Bennett, Rosenzweig, & Nelson, 1978).
REFERENCES Oeller, E. (1971). Some observations on the effects of environmental complexity and isolation on biochemical ontogeny. In M. B. Sterman, D. J. McGinty, & A. M. Adinolfi (Eds.), Brain Development and Behavior, pp. 277-296. New York: Academic Press.
Grouse, L. D., B. K., Bennett, E. L., Rosenzweig, M. R., & Nelson, P. G. (1978). Sequence diversity studies of rat brain RNA: effects of environmental complexity, on rat brain RNA diversity. Journal of Neurochemistry 30, 191-203. Rosenzweig, M. R., Bennett, E. L., & Diamond, M. C. (1972). Chemical and anatomical plasticity of brain: Replications and extensions. In J. Gaito (Ed.), Macromolecules and Behavior, pp. 205-277. New York: Appleton-Century-Crofts. Winer, B. J. (1971). Statistical Principles in Experimental Design, 2nd ed. New York: McGraw-Hill. Uphouse, L. (1978). In vitro RNA synthesis by chromatin from three brain regions of differentially reared rats. Behavioral Biology 22, 39-49. Uphouse, L., & Bonner, J. (1975). Preliminary evidence for the effects of environmental complexity on hybridization of rat brain RNA to rat unique DNA. Developmental Psychology 8, 171-178. Uphouse, L., & Moore, R. (1978). Effect of rearing condition on in vitro RNA synthesis by brain chromatin. Behavioral Biology 22, 28-38.
(1979)
BRIEF REPORT Environmental Enrichment and Brain Chromatin LYNDA
UPHOUSE
AND
BRUCE
TEDESCHI
1
Department of Psychology, Yale University, New Haven, Connecticut 06520
Twenty-eight-day-old Fischer rats were assigned to enriched, isolated, or standard environmental rearing for 60 days. Chromatin from cortex, cerebellum, and rest of brain was assayed for capacity to support in vitro RNA synthesis. No significant differences were observed across rearing groups. These results are compared to previous findings after shorter rearing intervals and are discussed in terms of a transient influence of the rearing environment on brain chromatin. Previous experiments have d e m o n s t r a t e d an impact of postweaning rearing environments on brain chromatin (Uphouse, 1978; U p h o u s e & Moore, 1978). When differential rearing continued from 28 to 60 days of age, enriched rats exhibited reduced whole brain chromatin-directed R N A synthesis. H o w e v e r , cortical chromatin from enriched animals supported m o r e R N A synthesis than did chromatin from isolated or standard colony-housed rats, but rest of brain chromatin was elevated in isolated animals. These data suggested that both enrichment and isolation had influenced brain chromatin. When differential rearing was extended from 28 to 88 days of age, whole brain chromatin from the three rearing groups was no longer different; but it remained possible that differences in parts of brain continued to exist. In the present study, parts of brain chromatin was examined from rats differentially reared from 28 to 88 days of age. Sixty male, Fischer inbred rats (Charles Rivers Laboratories, North Wilmington, Mass.) were used in the experiment. After arrival in the laboratory at 28 days of age the rats were assigned in two groups of 30 animals to enriched (EC), isolated (IC), or standard colony (SC) conditions as previously described (Uphouse & Moore, 1978). After 60 days in their respective environments, the rats were sacrificed by decapitation; brains were quickly removed, weighed, and dissected into cortex, rest of brain, and cerebellum as described by U p h o u s e (1978). Tissue was stored Research supported by NSF Grant BNS-74-03883. The authors wish to express appreciation to Ms. Ella FutreU for typing the manuscript. 268 0163- 1047/79/020268-03$02.00/0 Copyright © 1979 by Academic Press, Inc. All 1ights of reproduction in any form reserved.
269
ENVIRONMENT AND BRAIN CHROMATIN
at -85°C until chromatin extraction. Tissue from three to four animals of the same experimental condition was pooled and chromatin was extracted as previously described (Uphouse & Moore, 1978). Chromatin was always prepared in triplets so that matched tissues from enriched, isolated, and standard colony animals were simultaneously analyzed. The capacity for chromatin to support RNA synthesis was measured in the presence of exogenous Escherichia coli RNA polymerase (R-0501, Type II; Sigma Chemical Company), unlabeled ATP, CTP, and GTP (Sigma Chemical Company), and [3H]-UTP [5-3H]uridine 5'triphosphate, 22.7 Ci/mmole, New England Nuclear). Assay conditions and computation of relative RNA synthesis were as previously described (Uphouse & Moore, 1978). The data from individual brain regions were analyzed by one-way ANOVA (Winer, 1971). The relative capacity for chromatin to support RNA synthesis in vitro is shown in Table 1. Although the trend for cortical chromatin differences was in the same direction as that noted in rats differentially reared for only 32 days (Uphouse, 1978), the differences were smaller in magnitude and not as consistent as those observed in previous studies. Analysis of variance did not reveal significant differences among the treatment groups [F(2, 15) = 1.33]. Similarly, rest of brain and cerebellum did not differ across treatment conditions [respectively, F(2, 15) = 1.07; F (2, 12) = 2.12]. Since the chromatin changes previously found in animals differentially reared from 28 to 60 days were no longer present with the longer rearing interval, it is possible that differential rearing may exert only a transient influence on the capacity for chromatin to support in vitro RNA synthesis. However, a conclusive demonstration that the response is not permanent would require a more extensive investigation than employed in the present experiment. Ideally, several durations of differential rearing should be simultaneously investigated. The apparent transience of the part of brain response is, however, in agreement with previous studies of whole brain chromatin (Uphouse & Moore, 1978) and is compatible with the suggestion that the rearing condiTABLE 1 Relative R N A Synthesis by C h r o m a t i n from 88-Day-Old Differentially Reared Rats Rearing condition Part of brain
Chromatin preparations"
EC
IC
SC
Cortex Rest of brain Cerebellum
6 6 5
11.13 ± 1.65 12.47 ~ 0.948 11.84 _ 1.03
10.62 _+ 1.62 12.43 _+ 2.85 10.57 ___ 1.82
12.62 _ 2.85 14.2 - 2.83 12.43 ___ 1.43
T i s s u e from three to four brains was pooled for each chromatin preparation.
270
UPHOUSE AND TEDESCHI
tions influenced a developmental change in chromatin template activity, and that with increased duration in the rearing environment, all animals would eventually reach a similar level. It is equally possible that the animals habituated to their rearing environments. It has been suggested that isolated animals initially exhibit responses characteristic of stress but that, with increasing time in isolation, this response is diminished (Geller, 1971). Alternatively, enriched animals may habituate to the novel or arousal-producing aspects of the enriched environment. The apparent transience of the chromatin response observed both in whole brain and in parts of brain is in sharp contrast to the relatively permanent morphological changes seen in cortex after environmental enrichment (Rosenzweig, Bennett, & Diamond, 1972). H o w e v e r , it is difficult to ascertain what proportion of the morphological differences actually occur during the longer rearing durations. The continuance of morphological differences, even with much longer durations than those utilized in the present experiment, may reflect the maintenance of morphological changes which occurred earlier during differential rearing rather than their continued induction. Chromatin-directed R N A synthesis may be more responsive to variations in the environment (e.g., arousal, novelty, or stress) which diminish with increased time in the rearing environments. While such a response may be important for the facilitation of morphological c h a n g e s , it may not be essential for their maintenance. F u r t h e r m o r e , it is important to note that the in vitro m e a s u r e m e n t of R N A synthesis is a quantitative m e a s u r e m e n t and the lack of a difference does not preclude the presence of more subtle macromolecular changes. Such changes with longer rearing durations have been previously suggested ( U p h o u s e & Bonnet, 1975; Grouse, Schrier, Bennett, Rosenzweig, & Nelson, 1978).
REFERENCES Oeller, E. (1971). Some observations on the effects of environmental complexity and isolation on biochemical ontogeny. In M. B. Sterman, D. J. McGinty, & A. M. Adinolfi (Eds.), Brain Development and Behavior, pp. 277-296. New York: Academic Press.
Grouse, L. D., B. K., Bennett, E. L., Rosenzweig, M. R., & Nelson, P. G. (1978). Sequence diversity studies of rat brain RNA: effects of environmental complexity, on rat brain RNA diversity. Journal of Neurochemistry 30, 191-203. Rosenzweig, M. R., Bennett, E. L., & Diamond, M. C. (1972). Chemical and anatomical plasticity of brain: Replications and extensions. In J. Gaito (Ed.), Macromolecules and Behavior, pp. 205-277. New York: Appleton-Century-Crofts. Winer, B. J. (1971). Statistical Principles in Experimental Design, 2nd ed. New York: McGraw-Hill. Uphouse, L. (1978). In vitro RNA synthesis by chromatin from three brain regions of differentially reared rats. Behavioral Biology 22, 39-49. Uphouse, L., & Bonner, J. (1975). Preliminary evidence for the effects of environmental complexity on hybridization of rat brain RNA to rat unique DNA. Developmental Psychology 8, 171-178. Uphouse, L., & Moore, R. (1978). Effect of rearing condition on in vitro RNA synthesis by brain chromatin. Behavioral Biology 22, 28-38.
