Brain Research, 581 (1992) 1-9 © 1992 Elsevier Science Publishers B.V. All rights reserved. 0006-8993/92l$05.00

BRES 17692

1

Research Reports

Activation of mesocortical dopaminergic system in the rat in response to neonatal medial prefrontal cortex lesions. Concurrence with functional sparing J.M. de Brabander, C.G. van Eden, J.P.C. de Bruin and M.G.P. Feenstra The Netherlands Institute for Brain Research, Amsterdam (The Netherlands)

(Accepted 17 December 1991)) Key words: Key words:Frontal cortex; Recovery; Brain damage; Dopamine: Rat

Neonatal lesions of the medial part of the rat prefrontal cortex (mPFC) (performed at the age of 6 days) resulted in a sparing in the performance of spatial delayed alternation (SDA) and an increase in dopaminergic (DA) innervation. The increased DA innervation was primarily observed in the remaining part of the mPFC. The DA fibre density was considerably higher in the non-ablated part of the mPFC, and the fibres were thicker with more large varicosities compared with sham-operated controls. Biochemical measurements showed a 3.5-fold increase in DA concentration in the remaining part of the mPFC of the animals with neonatal lesions when compared with the mPFC of sham-operated animals. In addition the DA metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) were increased. The metabolite/transmitter ratios, indicating DA utilisation, did not significantly differ from controls. The increased DA innervation and the increased concentration of DA and its metabolites in the animals with neonatal lesions further support our hypothesis that the mesocortical DA system is involved in the neural mechanism of sparing of function observed after neonatal mPFC lesions. However, sparing of function in animals with no discernable mPFC forces us to conclude that this DA response cannot be the only factor involved in the mechanism of sparing of function. INTRODUCTION Lesions of the medial part of the prefrontal cortex ( m P F C ) in adult rats can result in a p e r m a n e n t l y impaired p e r f o r m a n c e of behavioural tasks, e.g. spatial delayed alternation ( S D A ) . H o w e v e r , m P F C lesions m a d e during the neonatal p e r i o d can show a sparing of these behavioural tasks 2'5"13'22'24'25'32. O u r aim is to investigate the n e u r o a n a t o m i c a l basis for this behavioural sparing in the rat. Several mechanisms have been p r o p o s e d to explain the occurrence of functional sparing and recovery of function in response to brain damage. O n e of these mechanisms implies an active reorganisation of neuronal circuitry, in which neural areas related to the d a m a g e d area take over its function 9'1°'28. A n u m b e r of investigators have r e p o r t e d p h e n o m e n a such as collateral sprouting and rerouting in response to brain d a m a g e 15'27'29'3°'

34. These observations resulted in the supposition that active morphological changes in neural circuitry m a y mediate sparing and recovery of function. In a previous study 6 we o b s e r v e d such changes in the dopaminergic ( D A ) projection from the ventral tegmental area (VTA) to the m P F C in response to partial m P F C lesions, perf o r m e d at the age of 6 days. C o n c o m i t a n t with a sparing

of function there was an increase in D A innervation which was most conspicuous in that part of the m P F C adjacent to the d a m a g e d part, i.e. the prelimbic and infralimbic areas (PL and IL). In these prefrontal subareas the density and thickness of D A fibres was increased and large fibre varicosities were m o r e numerous. Moreover, rats with adult m P F C lesions, exhibiting impaired performance of the S D A task, showed no such increase in D A innervation. These findings led us to postulate that d o p a m i n e might be involved in sparing of function. H o w e v e r , although findings o b t a i n e d by immunocytochemical staining techniques may reveal changes in the pattern and the degree of innervation of the d o p a m i n ergic system, they fail to reveal the functional status of the system. O n e way to d e t e r m i n e this would be to biochemically m e a s u r e the levels of d o p a m i n e and its metabolites 3,4-dihydroxyphenylacetic acid ( D O P A C ) and homovanillic acid ( H V A ) . The present study was conducted to (1) replicate the observations of behavioural sparing concomitant with the D A response, and (2) extend immunocytochemical results with biochemical data, which would reflect possible changes in the metabolic activity of the D A system.

Correspondence: J.M. de Brabander, The Netherlands Institute for Brain Research, Meibergdreef 33, 1105 AZ Amsterdam, The Netherlands. Fax: (31) (20) 6918466.

MATERIALS AND METHODS

Animals Male Wistar rats (n = 40) were born at our institute from pregnant females obtained from Harlan TNO. At birth (postnatal day 0, PNO) all animals were cross-fostered and weaned between PN20 and PN22. The animals were housed in groups of 4 in macrolon cages (75 × 55 × 23 cm) and kept on a 12-h light-dark cycle (lights on 15.30 h). The average temperature was between 22°C and 24°C, humidity between 50% and 60%. Just prior to and during the period of behavioural testing, food availability was reduced to achieve a body weight of approximately 85% of ad libitum fed rats (standard rodent pellets, Hope Farms, Woerden, The Netherlands). After completion of behavioural testing, the animals remained on this regimen until they were sacrificed.

Surgery The animals were operated on PN6. They were randomly assigned to either the experimental (E; n = 24) or sham-operated (S; n = 16) group. Every litter contained E and S rats. Under ether anaesthesia the skull of both E and S rats was opened and the anterior sinus sagittalis superior was tigated. In E animals the dorsal part of the medial prefrontal cortex was bilaterally ablated by aspiration (for a detailed description of the procedures, see De Brabander et al.5).

Behavioural testing At the age of 11 weeks, all rats were tested in a T-maze for their SDA behaviour. The T-maze consisted of a startbox plus main alley (65 cm) and two side arms of 40 cm (apparatus described in detail in Stare et al.38). In this test animals must alternate between left and right side arms to obtain a reward (Noyes food pellet, 45 mg). Testing was preceded by an adaptation and autoshaping period. The rats were first tested for 10 days with an intertrial interval of 0 s (ITI 0) between the 16 daily trials. Testing was continued for another 10 days with a 15-s intertrial interval (ITI 15; see De Brabander et al.S). The number of errors was recorded.

Histology and biochemistry After completion of behavioural testing, when the animals were 16-17 weeks old, E and S animals were randomly divided into either an immunocytochemical or biochemical group. TWenty animals (12 E and 8 S) were perfused for immunocytochemical DA staining and 20 animals (12 E and 8 S) were decapitated for biochemical high performance liquid chromatography (blPLC) measurement of DA and its metabolites DOPAC and HVA. Immunocytochemical staining. For the immunocytochemical DA staining (see also De Brabander et al.6), the animals were perfused with 5% glutaraldehyde in 0.05 M acetate buffer, pH = 4.0. Following perfusion the brains were removed and postfixed in the same fixative for an additional 30 min. The brains were then immersed in 0.05 M Tris-buffered saline (TBS) containing 1% Na2S205 (pH = 7.2). Using a vibroslice, alternate coronal sections (50 #m) were gathered for DA and Nissl staining, and collected in TBS containing 1% Na2S20 s (for the DA sections) or TBS (for the Nissl sections). The DA sections were stained using an antibody sensitive to DA, raised in our institute 11 and subsequently incubated via the peroxidase-antiperoxidase (PAP) method4°, with 0.5% nickel to enhance the 3,3-diamonobenzidine (DAB; Sigma) reaction. For the Nissl staining the sections were first mounted on glass slides and dried, then rinsed in a solution of ethanol and chloroform (1:1) to remove the lipids which brightens the stain (0.5% thionine). Photomicrographs of the D A sections were made of cortical layers V and VI of IL at a standard level of +2.5 mm of Bregma47 for both S and E animals. The photographs did not show whether the section came from an S or E rat. One sham-operated animal was not included in our analyses, due to suboptimal quality of the staining. The photomicrographs were blindly ranked by three separate investigators according to the degree of DA innervation, i.e. fibre density, thickness and number of large varicosities. Rank

number 1 was given to the photomicrograph of the section with the highest degree of innervation. The median rank numbers of the three investigators per photomicrograph (per animal) were statistically tested (see statistics). Biochemical HPLC measurements'. For the biochemical assays animals were decapitated and the brains were rapidly removed. The left and right mPFC (or what was left of it in the E group) were dissected out on an ice-cold plate (for description in detail, see Kalsbeek et al.lS). Immediately after dissection, the pieces were frozen on dry ice, weighed and stored at -80°C until analysed. DA and its metabolites, DOPAC and HVA, were determined by HPLC using electrochemical detection46,

Statistics Statistical analysis of the behavioural data was performed by means of the Statistical Package for the Social Sciences (SPSSX), using MANOVA with repeated measurements. The degree of the DA response was analysed by testing the median rank number of the photomicrographs (vide supra) with a Mann-Whitney U-test37. The biochemical data were also compared by means of the MannWhitney U-test. One animal of the E group with an extremely high DA concentration was discarded on the basis of the Grubbs's outliers test 8. The ratios DOPAC/DA and HVA/DA were calculated according to the 'Jackknife' method31 and analysed using the t-test. Paired comparisons of behavioural, immunocytochemical and biochemical data were made using the non-parametric Pearson correlation test 37.

RESULTS

Morphology of the lesion site After behavioural testing, the E and S animals were randomly divided into immunocytochemical and biochemical groups. In the E animals the lesions could be determined in two ways (1) in the immunocytochemical E group by studying the thionine-stained sections, and (2) in the biochemical E group by macroscopical inspection during dissection. In the immunocytochemical group, the S animals showed no indications of a disturbed cytoarchitectonic structure. In the E animals the lesion was visible as a cavity in the cerebrum, lined by glial tissue (Fig. 1). Along the border of the lesion site cortical cells formed a thin homogeneous band, in which the different layers were hard to distinguish. Lesion size and location was determined by cytoarchitectonic criteria 26'43'45'47, as described previously 6. In all animals the mPFC subareas frontal area 2 (Fr2) and the dorsal anterior cingulate area (ACd) had been removed completely, along with variable parts of the prelimbic (PL) and infralimbic areas (IL) (Fig. 2). One animal had an extremely large lesion, in which IL was completely removed and the caudate putamen severely damaged. The lesion size and location of the animals assigned for biochemical assays were investigated during dissection. There was no indication that these lesions differed from those of the immunocytochemical group. One animal had such an extensive lesion that the entire mPFC was removed and no sample could be taken.

Fig. 1. Photomicrograph of a coronal Nissl-stained section of the brain of a rat with a representative neonatal (PN6) mPFC lesion of the immunocytochemically processed group. The lesion is visible as a cavity lined by glial tissue: subareas Fr2, ACd and PL are removed. The boxed area indicates the location of the photomicrograph in Fig. 4.

Thus, both E groups contained one animal with an extremely large lesion. The d a t a o b t a i n e d from these two animals will be described separately.

Behavioural observation of sparing of function The animals were tested in a T-maze to assess their ability in learning to alternate b e t w e e n t h e arms, first with a 0-s intertrial interval (ITI 0; 10 sessions), then with a delay of 15 s b e t w e e n trials (ITI 15; 10 sessions). In the S group (n = 16) the percentage of correct responses during sessions with a delay of 0 s (ITI 0) increased rapidly (Fig. 3). A s early as testing day 4 most animals alternated with m o r e than 80% correct responses. A f t e r completion of 10 sessions, the task was m a d e m o r e difficult by lengthening the I T I delay to 15 s (ITI 15). O n testing day 1 the p e r f o r m a n c e of the S group was strongly decreased c o m p a r e d with testing day 10 of ITI 0, but this i m p r o v e d within a few days when the responses were again 80% correct or more. The animals with neonatal lesions (n = 22; cf. morphology of the lesion site) p e r f o r m e d as well as the S animals. T h e r e were no significant group differences in either the ITI 0 or I T I 15 tests, although Fig. 3 suggests that lesioned animals may t e m p o r a r i l y lag behind the p e r f o r m a n c e of the S animals. It is n o t e d that the two animals with e x t r e m e l y large lesions were not impaired in their S D A performance. A t the tenth ITI 0 session, they had 80% and 87% correct responses and at the last ITI 15 session 87% and 100%.

Dopamine in the mPFC Immunocytochemical staining. In 8 S and 12 E animals coronal sections of the frontal cortex were immunocytochemically stained for d o p a m i n e . The distribution and

Fig. 2. Reconstruction of the smallest (hatched) and largest (dotted) mPFC lesions made at PN6 (n = 11) of the immunocytochemically processed animals 33. The mPFC subareas Fr2 and ACd were completely removed along with variable parts of PL and IL.

m o r p h o l o g y of D A fibres in the P F C was essentially the same as described previously 6. In the frontal cortex of S animals the highest density of D A fibres was present in P F C areas. O t h e r frontal areas, outside the P F C , contained fewer fibres which were located primarily in layer VI (for details see Berger et al. 1 and Van E d e n et al.44). G e n e r a l l y speaking, within the m P F C the density of fibres was considerably higher in the basal cortical layers (V and VI) than in the m o r e

ITI 15

ITI 0

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5 6 7 Sessions

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Fig. 3. Performance of controls (filled circles; n = 16) and animals with neonatal lesions (open circles; n = 22) in the SDA task. The first 10 sessions consisted of 16 trials each with an intertrial interval (ITI) of 0 s, followed by another 10 sessions with a 15 s ITI.

superficial layers I, II and III. The DA fibres ran a specific course within different cortical layers: generally, the fibres in layer VI ran parallel and in layer II, III and V, perpendicular to the pial surface. The few fibres observed in layer I ran parallel to the medial cerebral surface in a ventrodorsal direction. In addition, regional differences in fibre density were seen between the cytoarchitectonically distinctive mPFC subareas IL, PL, ACd and Fr2. The density of DA fibres was highest in PL, in both basal and superficial cortical layers. IL and ACd contained fewer fibres in the superficial layers compared with PL, whereas the density in the basal layers was similar, except for that part of IL directly adjacent to the nucleus accumbens. Here the basal layers displayed a denser innervation. Of the four subareas, the lowest fibre density was found in Fr2, in all of its cortical layers. In this subarea, fibres were primarily limited to layer VI and the lower part of layer V. The dopaminergic innervation in animals with neona-

tal lesions (Figs. 4 and 5) was increased primarily in those medial prefrontal areas which were spared by the lesion. Here the density of DA fibres was higher, the fibres were thicker and there were more large fibre varicosities. The course of the DA fibres was similar to that seen in S animals. Directly adjacent to the lesion site,

TABLE I

Median rank numbers of the degree of the DA innervation of IL of experimental (E) and sham-operated (S) animals (see also Fig.

5) Photomicrographs were ranked by three investigators with the densest innervation rated as rank number 1. Note that the rank number of only two E animals are within the range of the S animals.

Median rank number of the three investigators E animals (n = 11) S animals (n = 7)

1

2

10

11

3

4

13 13

6

7

8 9

16 17 18

9

10 15

Fig. 4. Photomicrograph of the dopaminergic innervation of the basal layers of the PFC subarea IL at the level o f +2.5 mm of Bregma (Zilles atlas 47) in an animal with a neonatal (PN6) mPFC lesion. The boxed area indicates the location of the photomicrographs A,C, and E in Fig. 5. P, = pial surface; W, = white matter. Bar = 0.2 ram.

Fig. 5. Photomicrographs of the IL subarea (boxed area in Fig. 4) of (A) a sham-operated animal with the normal innervation pattern (rank number 18; Table I), with in (B) a detail of the DA fibres in basal layer V; (C) an experimental animal with a neonatal mPFC lesion with a slightly increased DA innervation (rank number 10), with in (D) a detail of the DA fibres in basal layer V; (E) an experimental animal with a large DA response (rank number 1), with in (F) a detail. W, = white matter. Bar in (E) = 0.08 mm (A, C, and E same magnification); bar in (F) = 0.02 mm (B, D, and F same magnification).

however, fibres were observed to r u n parallel to the border of the lesion, i.e. perpendicular to the medial cerebral surface. Photomicrographs of the basal layers (V and VI) from a standardised part of IL of both E (n = 11) and S animals (n = 7) were rated blindly on the basis of the degree of D A innervation (Table I). The obtained median rank n u m b e r s resulted in a clear distinction b e t w e e n E

and S group. There is only one E animal with a high m e d i a n rank n u m b e r , thus with virtually no increase in D A innervation compared with the controls. The average of the median rank n u m b e r s of the photographs of the E and S group was 6.8 4- 1.3 (-+ S.E.M.) and 14.0 + 1.2, respectively (P -< 0.01). We investigated whether changes in the dopaminergic innervation had occurred in other mesolimbocortical

TABLE II The effect of neonatal mPFC lesions on DA, DOPAC and HVA concentrations (in ng/g tissue + S.E.M. in the (remaining) mPFC and on the metabolite/transmitter ratios

Samples from both hemispheres were pooled.

DA DOPAC HVA DOPAC/DA HVA/DA

S animals (n = 8)

E animals (n = 10)

Percentageof S animals

146.5 + 42.1 + 46.7 + 0.285 + 0.288 +

525.2 + 127.5 + 110.7 + 0.243 + 0.203 +

358%* 303%** 237%** 85% 70%

29.8 6.7 6.7 0.01 0.01

143.3 27.5 16.9 0.02 0.02

*P < 0.01; **P < 0.001 (MWU-test).

brain areas innervated by D A fibres originating in the VTA. Only in the lateral part of the PFC (IPFC) a slight increase in D A fibre density was detected in 6 out of the 12 E animals. However, no relationship could be found between the increased innervation in the mPFC and the 1PFC. In the one animal with an extraordinarily large lesion, i.e. with almost nothing left of the mPFC, no D A response could be observed in the IPFC or any other mesolimbocortical brain area. Biochemical analysis o f dopamine and metabolites.

Concentrations of DA, D O P A C and HVA in the mPFC were measured in 8 S and 10 E animals (cf. morphology of the lesion site and statistics). In the S animals, the dissected part included the whole of mPFC, i.e. Fr2, ACd, PL and IL, whereas in the E animals only IL and the remaining part of PL were included in the sample (cf. morphology of the lesion site). Thus, tissue sample weights were different: 37.7 + 3.5 mg (+ S.E.M.) for the S animals and 21.9 + 2.3 mg for the E animals. The levels of D A and its metabolites D O P A C and HVA are presented in Table II. Animals with neonatal lesions contained significantly more D A in the remaining mPFC subareas, confirming the immunocytochemical results. The increase in D A content was 3.5-fold, and D O P A C and HVA content increased 3- and 2.5-fold, respectively, compared with concentrations in S animals. These differences were highly significant (MWU, P -< 0.01, P ~ 0.001 and P -< 0.001, respectively). The ratios D O P A C / D A and HVA/DA did not differ significantly between S and E animals. Relations between lesion-size, behaviour, tochemical staining or D A concentrations

The E animals differed sion size and dopaminergic ability in lesion size (Fig. SDA task (Fig. 3) was not

immunocy-

in their SDA behaviour, leresponse. Although the vari2) and performance in the very high, analyses of corre-

lation were performed to test whether a relationship could be detected between: (1) lesion size and performance in the SDA task, (2) lesion size and intensity of D A staining, (3) performance in the SDA task and intensity of D A staining, and (4) performance in the SDA task and D A concentration. None of the above 4 comparisons revealed a significant relationship. The Spearman rank-order correlation coefficients were: (1) r S = -0.14, (2) r~ = +0.18, (3) rs = -0.31 and (4) r~ = -0.47. Furthermore, the observation of behavioural sparing of the two animals in which the entire mPFC was removed, obviously without a D A response in the remaining mPFC, also contradicts the existence of a relationship. Thus, the size of the neonatal lesion is not critical for either the degree of sparing, or the concomitant D A response. In addition, the D A response, assessed with either immunocytochemical or biochemical tools, is no predictor for the performance of the SDA task. DISCUSSION The present study has shown that animals with partial mPFC lesions were spared in their performance of SDA behaviour. Neuroanatomically, an increased dopaminergic innervation was observed in the remaining part of mPFC, mainly in IL and PL, both immunocytochemically and biochemically. Behavioural sparing

The effects of neonatal and adult mPFC lesion have been examined in a previous study 5. Sparing of SDA behaviour was observed after neonatal lesions, whereas animals were impaired in the performance of this learning task when similar lesions were made in adulthood. This impairment was most pronounced during the testing days with a delay of 15 s between the trials; the animals failed to improve their performance. This difference in performance after neonatal versus adult lesions is in agreement with the observations of other investigators, using other mammalian species and/or behavioural tasks 2A3'24'32. The bilateral lesions of the animals in the present study included the mPFC subareas Fr2, ACd and variable parts of PL and IL, whereas in our previous study lesions involved only Fr2 and ACd 5. However, these somewhat larger neonatal mPFC lesions also resuited in a completely spared performance of SDA. In addition, the data of the present study show that the extent of the lesion was not of crucial importance for sparing of function to occur. The findings of Kolb and colleagues also support the observation that the occurrence of sparing of function is not limited by the extent of the mPFC lesion. Using other behavioural tasks they re-

ported sparing of function even after removal of the entire frontal cortex 22-25'32.

Dopamine, a neural substrate of sparing of function An enhanced dopaminergic innervation in the remaining part of the mPFC was observed in the animals with neonatal mPFC lesions: more branching, more and thicker D A fibres and more large varicosities. These data confirm our previous findings 6, and, although the lesions were somewhat larger in the present study, an enhancement of DA innervation in the remaining part of the mPFC was still present. In the immunocytochemically stained sections, the clearest dopaminergic reaction was found in the remaining mPFC, while in some animals an increased density of the D A fibre network was also observed in the 1PFC. A response of the 1PFC could be expected because of its close DA-ergic relationship with the mPFC. Both PFC parts are innervated by D A fibres and together with the VTA they represent the mesocortical D A system. However, the response in 1PFC did not occur in all animals and was certainly not as prominent as in the mPFC. Even in the one animal with complete mPFC removal no change of the mesocortical D A system, e.g. in the 1PFC, could be detected. Yet, this animal was spared in its SDA behaviour. Thus, the D A response in the remaining mPFC and 1PFC is probably not the only mechanism involved in sparing of function. Biochemical analysis also showed a large increase in the concentrations of D A and its metabolites D O P A C and HVA in the remaining mPFC of the animals with neonatal lesions, thus confirming the immunocytochemical findings. However, the dramatic increase (3.5-fold) in D A levels in animals with neonatal lesions compared with their controls should be interpreted with caution. The tissue sampled in lesioned and control animals differed and D A fibres and terminals are not homogeneously distributed over the dorsal (Fr2 and ACd) and ventral (PL and IL) mPFC areas 6'42'44. Such a difference in D A concentrations between dorsal and ventral mPFC has also been found in studies using biochemical methods. Data of Tassin et al. 42 showed that tissue samples of the ventral mPFC contained up to two times as much D A as samples of the dorsal mPFC. These findings have also been confirmed in our own laboratory using HPLC techniques (personal communication R.P.W. Heinsbroek and M.G.P. Feenstra). In the present study, the tissue sampled of lesioned animals contained only the ventral mPFC subareas, whereas the tissue of the controls contained the entire mPFC. Since the precise size of the lesions was unknown beforehand, it was impossible to dissect in controls only that part of the mPFC comparable to the non-ablated and dissected part of the mPFC in

experimentals. However, we can make an estimation of the D A concentration in the ventral mPFC of controls. Assuming that ventral and dorsal mPFC are of the same size and that the D A concentration of the ventral mPFC is twice as high as the dorsal mPFC, the D A concentration in the ventral mPFC is estimated to be one-third higher than in the entire mPFC. This is still much less than the 3.5-fold increase, which was found in the samples of the animals with neonatal lesions (containing only the ventral mPFC). Moreover, the absolute amount of D A in the much smaller samples of the animals with neonatal lesions was two times higher than in the samples of controls (11.5 and 5.4 ng, respectively). The metabolic rate in the S and E groups was assessed by D O P A C / D A and HVA/DA ratios. These ratios are used as indices of the rate of D A utilisation and they may also provide a useful index of DA release 17'21. In the present study it was established that DA, as well as its metabolites D O P A C and HVA, were increased in the animals with neonatal mPFC lesions. Furthermore, their ratios, D O P A C / D A and HVA/DA, were not significantly different although there was a trend towards a slight decrease when compared with controls. Thus, there are no significant changes in the metabolic activity of the dopaminergic system in response to the neonatal lesions. Taken together with the immunocytochemical observations, these biochemical data demonstrate an increased exension of the D A fibre network along with an increase in D A and its metabolites and therefore an increase in absolute utilisation of DA. This D A response is accompanied by behavioural sparing. However, although we expected to find a positive correlation between the degree of the dopaminergic response in the remainder of the mPFC and the degree of behavioural performance in the SDA task, correlation analysis failed to reveal such a relationship. The small variability in behavioural scores undoubtedly contributed to this negative finding: all animals were able to perform the SDA task at control level. It it also possible that the SDA task is not the most adequate one to examine the existence of a correlation with the D A innervation. Perhaps other behaviours, which also depend on the integrity of the mPFC, would have been more sensitive for the detection of such a positive correlation, e.g. water-maze performance, tongue extension, beam-walking etc. 22. In a previous study, we hypothesised that there was an involvement of dopamine in sparing of function because increased D A innervation was observed in animals with mPFC lesions produced neonatally 6. This was not seen in animals with lesions produced in the adult, which were also shown to be severely deficient in the SDA task 6. No data are available about possible changes in D A levels after adult mPFC ablations, but since no D A

immunocytochemical response was present, such changes are not expected. A n involvement of d o p a m i n e in behavioural sparing is also supported by the important role of D A in PFC-related tasks. Various studies, both in rhesus monkeys and rats, have reported that damaging

ing of function to occur. D A seems to be only one factor contributing to the observed sparing/recovery of function after brain lesions. Further studies will be required to determine whether other (monoaminergic) systems could

the D A pathway in adult animals results in an impaired performance of PFC-related tasks 3'4'7'19'38. Electrophys-

also be involved in the neural mechanism of sparing of function. Furthermore, the role of D A could be further investigated in the mechanism of behavioural recovery

iologically the importance of d o p a m i n e for an animal's performance of PFC-related tasks has recently been shown by Sawaguchi and his colleagues in monkeys 35'36.

after adult lesions. Several neurotrophic substances have been reported to induce behavioural and/or anatomical recovery after brain injury (e.g. Stein et al.39). Future

They reported that dopamine facilitated the activity of

studies will be carried out to determine whether such substances can induce recovery of function after adult

PFC neurons, which were directly involved in a PFC-related task. Also the ability of D A agonists to induce a recovery after frontal cortex lesions is an indication of an important role of dopamine in plasticity of the frontal c o r t e x 1 2 ' 1 4 ' 1 6 ' 2 0 ' 4 1 .

m P F C lesions, and whether recovery of function is accompanied by a similar response of the D A mesocortical system.

Thus, there are a n u m b e r of important features of the dopaminergic system to support the proposed involvem e n t of this transmitter system in sparing of function. However, although the mesocortical D A increase coincides with the observed sparing of function, this D A response does not seem to be an obligatory factor for spar-

Acknowledgements. We thank Alice Rinkens for her histological assistance, Ellis Jansen for her assistance in behavioural testing, and Margriet Botterblom for her biochemical assistance. We also want to thank Bob Baker for correcting the English, Henk Stoffels for drawing the figures and Gerben van der Meulen for preparing the photomicrographs.

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