Neuroscience Letters, 119 (1990) 97-100 Elsevier Scientific Publishers Ireland Ltd.
97
NSL 07262
Age-related changes in the subcortical afferents to the medial frontal cortex in mice: a W G A - H R P study Julio Villalobos, Daniel Beracochea and R o b e r t Jaffard Laboratoire de Psychophysiologie, URA CNRS 339, Universit#de Bordeaux L Talence (France) (Received 8 November 1989; Revised version received 5 July 1990; Accepted 11 July 1990)
Key words: Aging; Frontal cortex; Mouse; Retrograde transport HRP; Subeortieal afferent The subeortical afferent projections to the mediodorsal part of the frontal cortex were studied both qualitatively and quantitatively in young (3 months), adult (12 months) and aged (22 months) Balb/c mice by means of the retrograde transport of wheat germ agglutinine-horseradish peroxidase (WGA-HRP). A progressive decrease in the number of afferents was observed during aging with a differential pattern of reduction as a function of the subcortical structures. In adult mice a large reduction of afferents occurs in the diagonal band of Broea, the posterior thalamic nucleus, the zona incerta, the lateral hypothalamic area, the nuclei of amygdaloid complex, the posterior hypothalamic nucleus, the reticular pontine nucleus, the dorsal and medial raphe nuclei and the dorsal tegmental nucleus. In aged animals, only the anteromedial and mediodorsal thalamic nuclei, as well as the locus coeruleus appear to be clearly affected.
The prefrontal lobes are essential for higher mental functions and in particular those involved in the organisation of memory. Although lesions restricted to the frontal lobe do not ordinarily produce an amnesic syndrome in humans (in the sense that they do not interfere with information storage), memory performance may be affected on tasks that require access to various types of spatio-temporal information. Normal age-related changes in memory are usually termed 'benign', however older humans may not be as effective when they are submitted to effortful encoding/retrieval operations [3]. This observation is congruent with similar reports for patients with frontal lobe pathology [6]. In the aged brain, a large neuronal loss has been reported in humans and non human primates in various brain areas; however, there is no consensus on the existence of such neuroanatomical deficits in rodents. Animal experiments have provided behavioural evidence for age-related memory performance deficits in monkeys and rodents. Since it has been suggested that in rodents the medial prefrontal cortex is involved in memory processes [5] the present experiments were designed to investigate age-related changes of the subcortical afferents to the medial frontal cortex in mice. Thirty-six Balb/c mice of 3 (n = 10), 12 (n = 13) and 22 (n = 13) months of age were unilateraly injected durCorrespondence: J. Villalobos, Laboratoire de Psyehophysiologie, URA CNRS 339, Universit6 de Bordeaux I, Avenue des Facultrs, 33405 Talence Cedex, France. 0304-3940/90/$ 03.50 © 1990 Elsevier Scientific Publishers Ireland Ltd.
ing 9 min using stereotaxic procedures under Nesdonal anesthesia (100 mg/kg) in the mediodorsal parts of the frontal cortex with a 15 nl of 1.5 % wheat germ agglutinin horseradish peroxidase (WGA-HRP) complex dissolved in Tris buffer 0.1 M at pH 8.2 using a glass micropipette (20-25/an external tip diameter) connected to a Hamilton syringe (1/zl) coupled to a stepper motor. Twentyfour hours later, the animals were given an overdose of Nesdonal and were perfused intracardially with saline followed by a solution of 1% paraformaldehyde and 1.25% glutaraldehyde in 0.1 M phosphate buffer (PB) pH 7.4. The brains were dissected, postfixed (3 h) in the same fixative and stored overnight at 4°C in PB with 20 % sucrose. Thirty/an serials frozen sections were collected in PB and the peroxidase activity revealed by the sensitive TMB blue reaction [7]. The sections were counterstained with buffered Neutral red and stabilized [1]. Qualitative and quantitative analyses were carried out by observing and plotting by means of a camera lucida labelled neurons using a light microscope under 250 x magnification. The injection site was situated in all cases in the dorsomedial parts of the frontal cortex at different antero-posterior levels (Fig. la). The most posterior site was situated at the level of the beginning of the forceps minor of the corpus callosum. In no case, any major difference was observed in the localization of subcortical labelled neurons. In the anterior brain areas of young subjects, labelled neurons were situated in the ipsilateral part of the diagonal band of Broca and in the magnocellular preoptic nucleus. More caudally, labelled neurons
98
Fig. 1. Photomicrographs showing some of the subcortical afferents to the mediodorsal frontal cortex in mice. a: localization and extent of the WGA-HRP injection site. b: somatic labelling in the n. basalis magnoceUularisin the young animals, c and d: labelling in the dorsomedial thalamic n. in young (c) and aged (d) animals, e and f: retrograde labelled neurons in the locus coeruleus in young (e) and aged (f) mice. The density of labelled neurons was significantlylower in the aged animals. Bars: a = 100/zm;b,c,d,e,f = 100/am.
were f o u n d in the ventral p a l l i d u m a n d in the nucleus basalis magnocellularis. I n the thalamus, higher density of labelled n e u r o n s was observed in the a n t e r o m e d i a l a n d m e d i o d o r s a l nuclei. The n e u r o n s in the ventralis medialis, r h o m b o i d , reuniens a n d posterior thalamic nuclei were less labelled. A few labelled n e u r o n s were
f o u n d in the s u b t h a l a m i c z o n a incerta. I n the h y p o t h a l a mus, somatic labelling was observed in the posterior part of lateral h y p o t h a l a m i c area, the s u p r a m a m m i l l a r y nucleus a n d more weakly in the posterior h y p o t h a l a m i c nucleus. I n the a m y g d a l o i d complex, a high density of labelled n e u r o n s was located in the basolateral nucleus
99 a n d less intense labelling was f o u n d in the c o n t r a l a t e r a l p a r t o f this nucleus a n d in the a n t e r i o r a m y g d a l o i d nucleus. I n the b r a i n s t e m , the labelled n e u r o n s were situ a t e d in the d o r s a l a n d m e d i a l r a p h e nuclei, a n d o n l y a few labelled n e u r o n s were f o u n d in the p o n t o m e s e n c e p h a l i c reticular f o r m a t i o n . In the d o r s a l tegrnentum, a high d e n s i t y o f labelled n e u r o n s was o b s e r v e d in the locus coeruleus while fewer labelled n e u r o n s were f o u n d in the d o r s a l t e g m e n t a l g r a y a n d the ipsilateral p a r a b r a chial nucleus. These o b s e r v a t i o n s , p a r t i c u l a r l y those c o n c e r n i n g the t h a l a m o - c o r t i c a l p r o j e c t i o n s a r e in agreem e n t with the k n o w n o r g a n i z a t i o n o f the f r o n t a l c o r t e x in mice [4]. T h e q u a n t i t a t i v e analysis o n l y i n c l u d e d cases for which the l o c a l i z a t i o n a n d extent ( a p p r o x i m a t e l y 65 m m 2) o f the injection site were c o m p a r a b l e (Fig. 1). This led us to retain for analysis o n l y 3 subjects o f each age g r o u p . F o r each structure, the t o t a l n u m b e r o f labelled n e u r o n s was c o u n t e d , b y a d d i n g the n u m b e r o f n e u r o n s c o u n t e d in each slice. T h e results s h o w e d t h a t within the s a m e structure, there was little difference in the n u m b e r o f labelled n e u r o n s between a n i m a l s o f the s a m e age g r o u p . This o b s e r v a t i o n a l l o w e d us to a v e r a g e the t o t a l n u m b e r o f n e u r o n s for the s a m e structure. These m e a n s a r e s u m m a r i z e d in T a b l e I. The l o c a l i z a t i o n o f s u b c o r t i c a l afferents in a d u l t s (12 m o n t h s ) a n d a g e d (22 m o n t h s ) a n i m a l s was n o t q u a l i t a tively different f r o m t h a t o f y o u n g a n i m a l s (3 m o n t h s ) . T h e i n t e r - g r o u p s c o m p a r i s o n indicates t h a t the n u m b e r o f n e u r o n s r e t r o g r a d e l y labelled d e c r e a s e d progressively with age (see t o t a l n u m b e r o f labelled n e u r o n s , T a b l e I). H o w e v e r , the p a t t e r n o f r e d u c t i o n o f the afferent connections to the f r o n t a l c o r t e x as a f u n c t i o n o f age was n o t the s a m e for all s u b c o r t i c a l structures. I n adult, a large r e d u c t i o n o f s u b c o r t i c a l afferents o c c u r r e d in the d i a g o n a l b a n d o f Broca, the p o s t e r i o r t h a l a m i c nucleus, the z o n a incerta, the lateral h y p o t h a l a m i c nucleus, the reticular p o n t i n e nucleus, the d o r s a l a n d m e d i a l r a p h e a n d the d o r s a l t e g m e n t a l nuclei. In a g e d a n i m a l s the s a m e p a t t e r n o f afferents reduction was o b s e r v e d b u t was m u c h m o r e p r o n o u n c e d t h a n in 1 2 - m o n t h - o l d mice. H o w e v e r , very little o r n o c h a n g e s were o b s e r v e d in o t h e r structures such as the m a g n o c e l l u l a r p r e o p t i c nucleus, the ventral p a l l i d u m , the nucleus basalis m a g n o c e l l u l a r i s , the r h o m b o i d , the ventralis medialis a n d p a r a f a s c i c u l a r t h a l a m i c nuclei. In c o n t r a s t , we o b s e r v e d in aged mice o n l y a d r a m a t i c r e d u c t i o n o f subcortical afferents c o m i n g f r o m the a n t e r i o r t h a l a m i c a n d m e d i o d o r s a l t h a l a m i c nuclei (Fig. lc,d) as well as the afferents c o m i n g f r o m the p o s t e r i o r h y p o t h a l a m i c nuclei, the r e t i c u l o - p o n t i n e a n d locus coeruleus nuclei (Fig. le,f). H o w e v e r , as stated b y one referee o f this review, it m i g h t be n o t e d t h a t these r e d u c t i o n s are b a s e d
TABLE I A QUANTITATIVE ANALYSIS OF THE SUBCORTICAL AFFERENT PROJECTIONS TO THE MEDIODORSAL FRONTAL CORTEX IN YOUNG, ADULT, AND AGED MICE The results represent the mean number of neurons + standard error of mean (S.E.M.) plotted on the subcortical structures for the cases of a comparable injection site and extent observed for young adult and aged animals. For each structure of each case the same number of observations was conducted. Young (3 Adult (12 Aged (22 months) n = 3 months) n = 3 months) n = 3 Diagonal band of Broca Magn. preopticn. Ventral pallidum N. basalismagnocel.. Rhomboid Anteromedial thal. n. Ventralismedialis Mediodorsalth. n. Posterior thai. n. Zonaincerta Parafasicular thal. n. Lat. hypothal, area Anterior amygdaloid n. Basolat. amygd. n. ipsi Basolat. contro Basolat. supramammillaris Posteriorhypoth.n. Reticular pontine n. Dorsalraphe Medialraphe Dorsal tegmental n. Locus coeruleus Parabrachial n. Total
76.5 _ 72 + 56.3 _+ 243 + 283.2 __+
9.7 11.2 9.4 21.5 46.2
54.5 ± 8.2 36.1 + 61.8 + 10.3 73.1 _ 65.5 + 10.2 45 ± 248 ± 18.6 259 + 216.5 ± 28.6 205 ±
1564 _ 586 ± 577 _ 317.5 _ 88 +
234.6 63.4 42.6 64.8 24.3
1128 ± 584.6 ± 518.5 ± 132.7 +__ 30.5 +
216.8 58.6 51.2 48.4 14.6
9.4 12.24 8.6 23.4 31.4
512 + 86.9 609 ± 41.4 257.4__+ 35.3 126.2 _+ 95.6 27 ± 13.4
173.6 ± 21.4 171 ± 28.6 181.4 __+27.6 167.4_+ 36.4 57.1 ___ 13.3 38.2 ± 15.6 25.5 ±
6.3
14.5 ±
6.4
18 ± 7.5
1102 ±264.6492.5 ± 86.3 535 _91.9 249.4 ± 49.2 75.6 ___ 26.4 83.6 + 31.2 57 ___ 17.2 + 48.5 __+ 158.4 ± 81.1 ± 30.2 _ 208.5 ___ 22.1 ± 6201
11.4 5.8 12.6 28.2 9.7 9.6 32.4 9.3
36.1 __. 3.1 _+ 22.6 ± 69.4 ± 35.2 __. 16.4 ± 219.3 ± 19.3 + 4267
8.4 0.9 9.3 11.2 10.4 7.2 29.8 6.5
40
± 15.2 0 4.6 _ 2.3 76 + 15.1 41.3 + 11.6 18 __+ 6.1 110.2 ± 22.7 16 _ 7.6 3312
on an i n t e r g r o u p c o m p a r i s o n ; however, if the r e d u c t i o n o f labelled n e u r o n s is e v a l u a t e d b y the p e r c e n t a g e o f total n u m b e r o f labelled n e u r o n s p e r age g r o u p , then o n l y the r e d u c t i o n o f labelled n e u r o n s o b s e r v e d in the a n t e r o m e d i a n t h a l a m i c nucleus a p p e a r s to be i m p o r t a n t ( 11%) whereas o n l y w e a k m o d i f i c a t i o n s are o b s e r v e d in the o t h e r m e n t i o n e d structures. A s a whole, o u r results show a decrease o f subcortical afferents to the m e d i o - f r o n t a l c o r t e x in mice d u r i n g aging. This decrease m a y be a t t r i b u t e d to several factors. It m a y reflect either a differential n e u r o n a l d e a t h in the studied subcortical structures o r a d i m i n u t i o n o f terminal a r b o r i z a t i o n o f n e u r o n s p r o j e c t i n g to this cortical area. It m a y be d u e as well to a r e d u c t i o n o f p r e s y n a p t i c
100
uptake activities or to an impairment of retrograde axonal transport for macromolecules. The extent of the agerelated cellular loss process in rodents remains unclear [2]. However, our findings agree well with some studies showing on the one hand that the number of cholinergic neurons in the nucleus basalis magnocellularis in aged mice was not different from that of young animals [8] and on the other that dramatic neuronal losses occur during aging in the noradrenergic locus coeruleus [9]. This research was supported by the CNRS. We are grateful to Dr. Claude Messier for his help in writing this manuscript. 1 Adams, J.C., Stabilizing and rapid thionin staining of TMB-based HRP reaction product, Neurosci. Lett., 17 (1980) 7-9. 2 Coleman, P.D. and Flood, D.G., Neuron numbers and dendritic extent in normal aging and Alzbeimer's disease, NeurobioL of Aging, 8 (1987) 521-545.
3 Craik, F.I.M., Age differences in remembering. In L.R. Squire and N. Butters (Eds.), Neuropsychology of Memory, Guilford, New York, 1984, pp. 3-12. 4 Guldin, W.O., Pritzel, M. and Markowitsch, M., Prefrontal cortex of the mouse defined as cortical projection area of the thalamic mediodorsal nucleus, Brain Behav. Evol., 19 (1981) 93-107. 5 Kesner, R.P., Retroaspective and prospective coding of information: role of the medial prefrontal cortex, Exp. Brain Res., 74 (1989) 163167. 6 Luria, A.R., The frontal lobes and the regulation of behavior. In K.H. Pribram and A.R. Luria (Eds.), Psychophysiology of the Frontal Lobes, Academic Press, New York, 1973, pp. 3-26. 7 Mesulam, M., Tetramethyl benzidine for horseradish peroxidase neurohistochemistry: a non-carcinogenicblue reaction product with superior sensitivity for visualizing neural afferents and efferents, J. Histochem. Cytochem., 26 (1976) 106-117. 8 Mesulam, M.M., Mufson, E.J. and Rogers, J., Age-related shrinkage of cortically projecting cholinergic neurons: a selective effect, Ann. Neurol., 22 (1987) 31-36. 9 Sturrock, R.R. and Rao, K.A., A quantitative histological study of neuronal loss from the locus coeruleus of ageing mice, Neuropathol. Appl. Neurobiol., 11 (1985) 55-60.
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NSL 07262
Age-related changes in the subcortical afferents to the medial frontal cortex in mice: a W G A - H R P study Julio Villalobos, Daniel Beracochea and R o b e r t Jaffard Laboratoire de Psychophysiologie, URA CNRS 339, Universit#de Bordeaux L Talence (France) (Received 8 November 1989; Revised version received 5 July 1990; Accepted 11 July 1990)
Key words: Aging; Frontal cortex; Mouse; Retrograde transport HRP; Subeortieal afferent The subeortical afferent projections to the mediodorsal part of the frontal cortex were studied both qualitatively and quantitatively in young (3 months), adult (12 months) and aged (22 months) Balb/c mice by means of the retrograde transport of wheat germ agglutinine-horseradish peroxidase (WGA-HRP). A progressive decrease in the number of afferents was observed during aging with a differential pattern of reduction as a function of the subcortical structures. In adult mice a large reduction of afferents occurs in the diagonal band of Broea, the posterior thalamic nucleus, the zona incerta, the lateral hypothalamic area, the nuclei of amygdaloid complex, the posterior hypothalamic nucleus, the reticular pontine nucleus, the dorsal and medial raphe nuclei and the dorsal tegmental nucleus. In aged animals, only the anteromedial and mediodorsal thalamic nuclei, as well as the locus coeruleus appear to be clearly affected.
The prefrontal lobes are essential for higher mental functions and in particular those involved in the organisation of memory. Although lesions restricted to the frontal lobe do not ordinarily produce an amnesic syndrome in humans (in the sense that they do not interfere with information storage), memory performance may be affected on tasks that require access to various types of spatio-temporal information. Normal age-related changes in memory are usually termed 'benign', however older humans may not be as effective when they are submitted to effortful encoding/retrieval operations [3]. This observation is congruent with similar reports for patients with frontal lobe pathology [6]. In the aged brain, a large neuronal loss has been reported in humans and non human primates in various brain areas; however, there is no consensus on the existence of such neuroanatomical deficits in rodents. Animal experiments have provided behavioural evidence for age-related memory performance deficits in monkeys and rodents. Since it has been suggested that in rodents the medial prefrontal cortex is involved in memory processes [5] the present experiments were designed to investigate age-related changes of the subcortical afferents to the medial frontal cortex in mice. Thirty-six Balb/c mice of 3 (n = 10), 12 (n = 13) and 22 (n = 13) months of age were unilateraly injected durCorrespondence: J. Villalobos, Laboratoire de Psyehophysiologie, URA CNRS 339, Universit6 de Bordeaux I, Avenue des Facultrs, 33405 Talence Cedex, France. 0304-3940/90/$ 03.50 © 1990 Elsevier Scientific Publishers Ireland Ltd.
ing 9 min using stereotaxic procedures under Nesdonal anesthesia (100 mg/kg) in the mediodorsal parts of the frontal cortex with a 15 nl of 1.5 % wheat germ agglutinin horseradish peroxidase (WGA-HRP) complex dissolved in Tris buffer 0.1 M at pH 8.2 using a glass micropipette (20-25/an external tip diameter) connected to a Hamilton syringe (1/zl) coupled to a stepper motor. Twentyfour hours later, the animals were given an overdose of Nesdonal and were perfused intracardially with saline followed by a solution of 1% paraformaldehyde and 1.25% glutaraldehyde in 0.1 M phosphate buffer (PB) pH 7.4. The brains were dissected, postfixed (3 h) in the same fixative and stored overnight at 4°C in PB with 20 % sucrose. Thirty/an serials frozen sections were collected in PB and the peroxidase activity revealed by the sensitive TMB blue reaction [7]. The sections were counterstained with buffered Neutral red and stabilized [1]. Qualitative and quantitative analyses were carried out by observing and plotting by means of a camera lucida labelled neurons using a light microscope under 250 x magnification. The injection site was situated in all cases in the dorsomedial parts of the frontal cortex at different antero-posterior levels (Fig. la). The most posterior site was situated at the level of the beginning of the forceps minor of the corpus callosum. In no case, any major difference was observed in the localization of subcortical labelled neurons. In the anterior brain areas of young subjects, labelled neurons were situated in the ipsilateral part of the diagonal band of Broca and in the magnocellular preoptic nucleus. More caudally, labelled neurons
98
Fig. 1. Photomicrographs showing some of the subcortical afferents to the mediodorsal frontal cortex in mice. a: localization and extent of the WGA-HRP injection site. b: somatic labelling in the n. basalis magnoceUularisin the young animals, c and d: labelling in the dorsomedial thalamic n. in young (c) and aged (d) animals, e and f: retrograde labelled neurons in the locus coeruleus in young (e) and aged (f) mice. The density of labelled neurons was significantlylower in the aged animals. Bars: a = 100/zm;b,c,d,e,f = 100/am.
were f o u n d in the ventral p a l l i d u m a n d in the nucleus basalis magnocellularis. I n the thalamus, higher density of labelled n e u r o n s was observed in the a n t e r o m e d i a l a n d m e d i o d o r s a l nuclei. The n e u r o n s in the ventralis medialis, r h o m b o i d , reuniens a n d posterior thalamic nuclei were less labelled. A few labelled n e u r o n s were
f o u n d in the s u b t h a l a m i c z o n a incerta. I n the h y p o t h a l a mus, somatic labelling was observed in the posterior part of lateral h y p o t h a l a m i c area, the s u p r a m a m m i l l a r y nucleus a n d more weakly in the posterior h y p o t h a l a m i c nucleus. I n the a m y g d a l o i d complex, a high density of labelled n e u r o n s was located in the basolateral nucleus
99 a n d less intense labelling was f o u n d in the c o n t r a l a t e r a l p a r t o f this nucleus a n d in the a n t e r i o r a m y g d a l o i d nucleus. I n the b r a i n s t e m , the labelled n e u r o n s were situ a t e d in the d o r s a l a n d m e d i a l r a p h e nuclei, a n d o n l y a few labelled n e u r o n s were f o u n d in the p o n t o m e s e n c e p h a l i c reticular f o r m a t i o n . In the d o r s a l tegrnentum, a high d e n s i t y o f labelled n e u r o n s was o b s e r v e d in the locus coeruleus while fewer labelled n e u r o n s were f o u n d in the d o r s a l t e g m e n t a l g r a y a n d the ipsilateral p a r a b r a chial nucleus. These o b s e r v a t i o n s , p a r t i c u l a r l y those c o n c e r n i n g the t h a l a m o - c o r t i c a l p r o j e c t i o n s a r e in agreem e n t with the k n o w n o r g a n i z a t i o n o f the f r o n t a l c o r t e x in mice [4]. T h e q u a n t i t a t i v e analysis o n l y i n c l u d e d cases for which the l o c a l i z a t i o n a n d extent ( a p p r o x i m a t e l y 65 m m 2) o f the injection site were c o m p a r a b l e (Fig. 1). This led us to retain for analysis o n l y 3 subjects o f each age g r o u p . F o r each structure, the t o t a l n u m b e r o f labelled n e u r o n s was c o u n t e d , b y a d d i n g the n u m b e r o f n e u r o n s c o u n t e d in each slice. T h e results s h o w e d t h a t within the s a m e structure, there was little difference in the n u m b e r o f labelled n e u r o n s between a n i m a l s o f the s a m e age g r o u p . This o b s e r v a t i o n a l l o w e d us to a v e r a g e the t o t a l n u m b e r o f n e u r o n s for the s a m e structure. These m e a n s a r e s u m m a r i z e d in T a b l e I. The l o c a l i z a t i o n o f s u b c o r t i c a l afferents in a d u l t s (12 m o n t h s ) a n d a g e d (22 m o n t h s ) a n i m a l s was n o t q u a l i t a tively different f r o m t h a t o f y o u n g a n i m a l s (3 m o n t h s ) . T h e i n t e r - g r o u p s c o m p a r i s o n indicates t h a t the n u m b e r o f n e u r o n s r e t r o g r a d e l y labelled d e c r e a s e d progressively with age (see t o t a l n u m b e r o f labelled n e u r o n s , T a b l e I). H o w e v e r , the p a t t e r n o f r e d u c t i o n o f the afferent connections to the f r o n t a l c o r t e x as a f u n c t i o n o f age was n o t the s a m e for all s u b c o r t i c a l structures. I n adult, a large r e d u c t i o n o f s u b c o r t i c a l afferents o c c u r r e d in the d i a g o n a l b a n d o f Broca, the p o s t e r i o r t h a l a m i c nucleus, the z o n a incerta, the lateral h y p o t h a l a m i c nucleus, the reticular p o n t i n e nucleus, the d o r s a l a n d m e d i a l r a p h e a n d the d o r s a l t e g m e n t a l nuclei. In a g e d a n i m a l s the s a m e p a t t e r n o f afferents reduction was o b s e r v e d b u t was m u c h m o r e p r o n o u n c e d t h a n in 1 2 - m o n t h - o l d mice. H o w e v e r , very little o r n o c h a n g e s were o b s e r v e d in o t h e r structures such as the m a g n o c e l l u l a r p r e o p t i c nucleus, the ventral p a l l i d u m , the nucleus basalis m a g n o c e l l u l a r i s , the r h o m b o i d , the ventralis medialis a n d p a r a f a s c i c u l a r t h a l a m i c nuclei. In c o n t r a s t , we o b s e r v e d in aged mice o n l y a d r a m a t i c r e d u c t i o n o f subcortical afferents c o m i n g f r o m the a n t e r i o r t h a l a m i c a n d m e d i o d o r s a l t h a l a m i c nuclei (Fig. lc,d) as well as the afferents c o m i n g f r o m the p o s t e r i o r h y p o t h a l a m i c nuclei, the r e t i c u l o - p o n t i n e a n d locus coeruleus nuclei (Fig. le,f). H o w e v e r , as stated b y one referee o f this review, it m i g h t be n o t e d t h a t these r e d u c t i o n s are b a s e d
TABLE I A QUANTITATIVE ANALYSIS OF THE SUBCORTICAL AFFERENT PROJECTIONS TO THE MEDIODORSAL FRONTAL CORTEX IN YOUNG, ADULT, AND AGED MICE The results represent the mean number of neurons + standard error of mean (S.E.M.) plotted on the subcortical structures for the cases of a comparable injection site and extent observed for young adult and aged animals. For each structure of each case the same number of observations was conducted. Young (3 Adult (12 Aged (22 months) n = 3 months) n = 3 months) n = 3 Diagonal band of Broca Magn. preopticn. Ventral pallidum N. basalismagnocel.. Rhomboid Anteromedial thal. n. Ventralismedialis Mediodorsalth. n. Posterior thai. n. Zonaincerta Parafasicular thal. n. Lat. hypothal, area Anterior amygdaloid n. Basolat. amygd. n. ipsi Basolat. contro Basolat. supramammillaris Posteriorhypoth.n. Reticular pontine n. Dorsalraphe Medialraphe Dorsal tegmental n. Locus coeruleus Parabrachial n. Total
76.5 _ 72 + 56.3 _+ 243 + 283.2 __+
9.7 11.2 9.4 21.5 46.2
54.5 ± 8.2 36.1 + 61.8 + 10.3 73.1 _ 65.5 + 10.2 45 ± 248 ± 18.6 259 + 216.5 ± 28.6 205 ±
1564 _ 586 ± 577 _ 317.5 _ 88 +
234.6 63.4 42.6 64.8 24.3
1128 ± 584.6 ± 518.5 ± 132.7 +__ 30.5 +
216.8 58.6 51.2 48.4 14.6
9.4 12.24 8.6 23.4 31.4
512 + 86.9 609 ± 41.4 257.4__+ 35.3 126.2 _+ 95.6 27 ± 13.4
173.6 ± 21.4 171 ± 28.6 181.4 __+27.6 167.4_+ 36.4 57.1 ___ 13.3 38.2 ± 15.6 25.5 ±
6.3
14.5 ±
6.4
18 ± 7.5
1102 ±264.6492.5 ± 86.3 535 _91.9 249.4 ± 49.2 75.6 ___ 26.4 83.6 + 31.2 57 ___ 17.2 + 48.5 __+ 158.4 ± 81.1 ± 30.2 _ 208.5 ___ 22.1 ± 6201
11.4 5.8 12.6 28.2 9.7 9.6 32.4 9.3
36.1 __. 3.1 _+ 22.6 ± 69.4 ± 35.2 __. 16.4 ± 219.3 ± 19.3 + 4267
8.4 0.9 9.3 11.2 10.4 7.2 29.8 6.5
40
± 15.2 0 4.6 _ 2.3 76 + 15.1 41.3 + 11.6 18 __+ 6.1 110.2 ± 22.7 16 _ 7.6 3312
on an i n t e r g r o u p c o m p a r i s o n ; however, if the r e d u c t i o n o f labelled n e u r o n s is e v a l u a t e d b y the p e r c e n t a g e o f total n u m b e r o f labelled n e u r o n s p e r age g r o u p , then o n l y the r e d u c t i o n o f labelled n e u r o n s o b s e r v e d in the a n t e r o m e d i a n t h a l a m i c nucleus a p p e a r s to be i m p o r t a n t ( 11%) whereas o n l y w e a k m o d i f i c a t i o n s are o b s e r v e d in the o t h e r m e n t i o n e d structures. A s a whole, o u r results show a decrease o f subcortical afferents to the m e d i o - f r o n t a l c o r t e x in mice d u r i n g aging. This decrease m a y be a t t r i b u t e d to several factors. It m a y reflect either a differential n e u r o n a l d e a t h in the studied subcortical structures o r a d i m i n u t i o n o f terminal a r b o r i z a t i o n o f n e u r o n s p r o j e c t i n g to this cortical area. It m a y be d u e as well to a r e d u c t i o n o f p r e s y n a p t i c
100
uptake activities or to an impairment of retrograde axonal transport for macromolecules. The extent of the agerelated cellular loss process in rodents remains unclear [2]. However, our findings agree well with some studies showing on the one hand that the number of cholinergic neurons in the nucleus basalis magnocellularis in aged mice was not different from that of young animals [8] and on the other that dramatic neuronal losses occur during aging in the noradrenergic locus coeruleus [9]. This research was supported by the CNRS. We are grateful to Dr. Claude Messier for his help in writing this manuscript. 1 Adams, J.C., Stabilizing and rapid thionin staining of TMB-based HRP reaction product, Neurosci. Lett., 17 (1980) 7-9. 2 Coleman, P.D. and Flood, D.G., Neuron numbers and dendritic extent in normal aging and Alzbeimer's disease, NeurobioL of Aging, 8 (1987) 521-545.
3 Craik, F.I.M., Age differences in remembering. In L.R. Squire and N. Butters (Eds.), Neuropsychology of Memory, Guilford, New York, 1984, pp. 3-12. 4 Guldin, W.O., Pritzel, M. and Markowitsch, M., Prefrontal cortex of the mouse defined as cortical projection area of the thalamic mediodorsal nucleus, Brain Behav. Evol., 19 (1981) 93-107. 5 Kesner, R.P., Retroaspective and prospective coding of information: role of the medial prefrontal cortex, Exp. Brain Res., 74 (1989) 163167. 6 Luria, A.R., The frontal lobes and the regulation of behavior. In K.H. Pribram and A.R. Luria (Eds.), Psychophysiology of the Frontal Lobes, Academic Press, New York, 1973, pp. 3-26. 7 Mesulam, M., Tetramethyl benzidine for horseradish peroxidase neurohistochemistry: a non-carcinogenicblue reaction product with superior sensitivity for visualizing neural afferents and efferents, J. Histochem. Cytochem., 26 (1976) 106-117. 8 Mesulam, M.M., Mufson, E.J. and Rogers, J., Age-related shrinkage of cortically projecting cholinergic neurons: a selective effect, Ann. Neurol., 22 (1987) 31-36. 9 Sturrock, R.R. and Rao, K.A., A quantitative histological study of neuronal loss from the locus coeruleus of ageing mice, Neuropathol. Appl. Neurobiol., 11 (1985) 55-60.
