Brain Research, 537 (1990) 271-278 Elsevier

271

BRES 16196

Spatial memory deficits in aged rats: contributions of monoaminergic systems Victoria Luine, Deborah Bowling and Michelle Hearns Department of Psychology, Hunter College of the City University of New York, New York, NY 10021 (U.S.A.) (Accepted 31 July 1990)

Key words: Aging; Spatial memory; Monoamine; Basal forebrain; Fischer 344 rat

Age-dependent changes in monoaminergic systems and their relationship to senescent memory decline were investigated in 4- and 25-26-month-old, female, Fischer 344 rats. Spatial memory performance was tested on an 8-arm radial maze, and levels of norepinephrine (NE), dopamine (DA) and metabolites 3,4-dihydroxyphenylacetic acid and homovanillic acid, serotonin (5-HT) and metabolite 5hydroxyindoleacetic acid were measured in brain areas which contribute to memory function - - basal forebrain cholinergic nuclei, subfields of the hippoeampus, frontal and entorhinal cortex - - and in monoaminergiccell body areas. The performance of aged subjects was sitmifieanfly impaired as compared to young subjects, and alterations of 20-60% in monoamine and metabolite levels were measured in specific brain areas of aged rats. Decreased NE levels were found in basal forebrain nuclei and cortical areas but not in hippocampal subfields of aged rats. Changes in the 5-HT system were present in hippocampal, cortical and basal forebrain sites. Changes in the DA system were the most pervasive with aged rats showing decreased DA and/or metabolites in several basal forebrain nuclei, cortical areas, and the hippoeampus. Aged rats showed 50% decreases of monoamines in locus coeruleus and substantia nigra and 30% decreases in the dorsal raphe nucleus. Some but not all of the changes correlated with memory performance. The present results in rats support evidence that age-dependent changes in monoaminergic function in discrete brain sites contribute to senescent memory decline and suggest that monoaminergic-cholinergicinteractions within basal forebrain nuclei may be important in this decline.

INTRODUCTION

memory have been shown 9'22'28'29'43.

The ability to perform tasks requiring memory declines with aging in rodents, non-human primates and humans3; however, the central mechanisms responsible for senescent memory loss are not well understood. Decreased cholinergic function has been shown in memory loss in Alzheimer's disease 38,49 and in some aged animal models 5'14'25. A number of other neural systems are also altered with aging (see ref. 33 for review), but whether such age-related changes contribute to senescent memory loss has not been documented. An older literature links some aspects of learning and memory to monoaminergic function 4"52. For example 6-hydroxydopamine (6-OHDA)-induced lesions in the prefrontal cortex of monkeys results in decreased performance of two spatial memory tasks 2. More recent studies using drug or lesion paradigms in young rats show that attenuations in norepinephrine (NE), dopamine (DA), or serotonin (5-HT) function impair performance in some but not all tasks requiring memory 1,1°'4s. In addition, interactions between monoaminergic and cholinergic systems in the performance of tasks requiring

In this study, we investigated whether alterations in monoaminergic function contribute to senescent memory decline. Monoamines and metabolites were measured in aged and young rats that were also evaluated for spatial memory performance on the 8-arm radial maze. Levels of NE, DA, 3,4-dihydroxyphenylacetic acid (DOPAC) and" homovanillic acid (HVA) (where detectable) and 5hydroxyindoleacetic acid (5-HIAA) were measured in the hippocampus and frontal and entorhinal cortices, brain areas which contribute to memory function 3. In addition, because of reported monoaminergic-cholinergic interactions in memory performance, nuclei in the basal forebrain, which contain cholinergic cell bodies 32,as and have a major role in memory performance 3,12,13'z3, were also sampled. Monoamines and metabolites were also measured in monoaminergic cell body areas in the hindbrain. Alterations of 20-60% in monoamine and metabolite levels were measured in specific brain areas of aged rats, and many of these changes correlated with spatial memory performance of the rats. Thus, this study provides preliminary evidence in rats that attenuated monoaminergic activity contributes to age-related de-

Correspondence: V. Luine, Department of Psychology,Hunter College, 695 Park Ave., New York, NY 10021, U.S.A. 0006-8993/90/$03.50 © 1990 Elsevier Science Publishers B.V. (Biomedical Division)

272 c l i n e s in m e m o r y

and

suggests

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monoaminergic-

c h o l i n e r g i c i n t e r a c t i o n s w i t h i n b a s a l f o r e b r a i n sites m a y b e critical in s e n e s c e n t m e m o r y loss.

MATERIALS AND METHODS

Animals Young (3 months upon arrival) and aged (24-25 months upon arrival) female, Fischer 344 rats (National Institute on Aging Colony at Harlan-Sprague-Dawley, Indianapolis, IN) were caged individually and maintained on a reversed light-dark cycle (lights off 07.00-19.00 h). One week prior to training on the maze and during testing, the feeding schedule was restricted so that a body weight of 80-85% of free-feeding levels could be reached and maintained. Behavioral testing Spatial memory was tested and evaluated on an 8-arm maze according to the general methods of Olton et. al. 34 with a pellet of Thrive dry cat food serving as food reinforcement. The maze was located in a dimly lit room containing black and white posters on the walls, asymmetrical lighting, and furniture, all of which provided a rich environment of extra-maze cues. Since all rats were naive to the maze task, they received two training sessions per day (morning and afternoon) for 5 consecutive days. By the last two training sessions, the task was identical to the test trials: one pellet was placed in each of the food receptacles and a maximum of 10 min was allowed to visit all 8 arms and eat the food reinforcement. Data from the l0 training sessions were not included in the behavioral analyses. Subjects were tested twice a day for 10 days for a total of 20 trials. To begin each trial, the rat was placed on the central platform in a random orientation and then allowed to enter any of the arms. A visit to an arm was scored if the subject transversed 3/4 s of the length of the arm, if it entered the end of the arm but did not eat, or if it entered the arm and ate the Thrive. The subject was allowed to choose arms in any order until either all 8 arms had been visited or 10 min had elapsed. Re-entries into an arm previously visited during that session were counted as mistakes or errors. Five aged rats did not perform on the maze: they remained primarily in the center hub throughout training and for up to 8 test trials. Four other aged rats died before completing the 20 trials. Data for these 9 aged rats were eliminated from the behavioral and neurochemical analyses, leaving 15 aged rats for which data were obtained and analyzed. All 10 young rats which began the study completed the 20 trials on the maze. Behavior was tested in 4 cohorts of subjects, each containing aged and young subjects. Choice accuracy was scored by two measures: number of correct choices in the first 8 visits and number of correct choices until the first mistake. For statistical analyses, choice accuracy data were averaged to give 4 blocks of 5 trials each. Differences in choice accuracy between the young and old subjects was analyzed by two-way (age vs trials) repeated measures ANOVA. Tissue sampling and biochemical measurements Within 1 h following the twentieth trial, subjects were sacrificed by rapid decapitation, and their brains removed and stored at -70 °C until all subjects had completed the behavioral trials. A portion of the brain containing the frontal cortex was obtained by excising approximately 2 mm of tissue from the rostral end. This piece was thaw mounted onto a microscope slide. Serial, 300-/zm-thick sections were made from the rest of the brain, beginning at approximately plate 15 and ending at plate 53 in the atlas of Palkovits and Brownstein 35, thaw mounted on microscope slides, and stored at -70 °C. Brain areas were identified with atlases35'36 and micropunched according to the methods of Palkovits and Brownsteinas as previously utilized and described by this laboratory T M . During sampling, one-half of each nucleus/area (alternate left and right sides of brain) was punched for monoamine measurements and the other half was punched for choline acetyltransferase measurements,

the results of which can be found in Luine and Hearns 25. The areas sampled, the plate numbers sampled from (ref. 35), and number of 500-/am punches taken (n) were as follows: vertical nucleus of the diagonal bands, 15-17 (n = 9); horizontal nucleus of the diagonal bands 19-21 (n = 3); nucleus basalis (also referred to as ventral globus pallidus or ventral pallidum, 34,35), 18-21 (n = 6); dentate gyrus (dorsal hippocampus), 28-33 (n = 12); CA1 (dorsal hippocampus), 28-33 (n = 12); entorhinal cortex 36-38 (n = 10); striatum, 18, (4 in the dorsal portion), and frontal cortex (n = 2); substantia nigra, 37 (n = 4); locus coeruleus, 51 (n = 2); dorsal raphe nucleus, 42,43 (n = 6). The medial septal area was excised from the septum in plate 17 using a small scapula. NE, DA and its metabolites DOPAC and HVA, and 5-HT and its metabolite 5-HIAA were measured using high-performance liquid chromatography (HPLC) with electrochemical detection as described previously 15"41. Briefly, punched samples were expelled into 60/tl of a sodium acetate buffer (pH 5) containing 1 x 10 -7 M a-methyldopamine as an internal standard. Following centrifugation, the supernatant was removed and 2/~l of a 1 mg/ml ascorbate oxidase solution (Boehringer Mannheim) was added to each sample. 40 /~1 of the supernatant was injected into a Waters Associates chromatographic system consisting of a WISP, 590 pump, and C-18 reverse-phase microbondapak column. A Bioanalytical Systems LC4B potentiostat was set at +0.6 V with respect to an Ag/AgCI reference electrode. Concentrations of neurotransmitters were calculated by reference to standards using peak integration with a computer assisted Waters Maxima 820 system. The pellet was dissolved in 100 kd of 0.2 N NaOH for protein determination by the Bradford method 6. Amine levels are expressed as pg//~g protein. Differences in levels between aged and young rats were tested by Student's t-test. Possible relationships between monoamine or metabolite levels and behavioral parameters were tested by correlational analyses. RESULTS Performance of aged rats on the 8-arm radial maze was s i g n i f i c a n t l y i m p a i r e d as c o m p a r e d t o t h e y o u n g rats. F o r t h e 20 trials, t h e a v e r a g e n u m b e r o f c o r r e c t c h o i c e s in t h e first 8 visits in t h e y o u n g g r o u p w a s 7 . 2 0 _+ 0.07 a n d in t h e a g e d g r o u p w a s 5.83 + 0.16 (F1,23 = 60.2, P < 0.001). Aged

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Spatial memory deficits in aged rats: contributions of monoaminergic systems.

Age-dependent changes in monoaminergic systems and their relationship to senescent memory decline were investigated in 4- and 25-26-month-old, female,...
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