- Email: [email protected]
Transmitter balances in the olfactory cortex: Adaptations to early methamphetamine trauma and rearing environment
BR A IN RE S E A RCH 1 1 41 ( 20 0 7 ) 3 7 –4 7
a v a i l a b l e a t w w w. s c i e n c e d i r e c t . c o m
w w w. e l s e v i e r. c o m / l o c a t e / b r a i n r e s
Research Report
Transmitter balances in the olfactory cortex: Adaptations to early methamphetamine trauma and rearing environment☆ Konrad Lehmann⁎, Devrim Lehmann Institute for General Zoology and Animal Physiology, Erbertstr. 1, 07743 Jena, Germany
A R T I C LE I N FO
AB S T R A C T
Article history:
The olfactory cortex, comprising the anterior olfactory cortex (AOC) and the anterior
Accepted 5 January 2007
piriform cortex (PirC), is a model system for the study of neural plasticity. We investigated
Available online 12 January 2007
the structural imbalances of different transmitter systems induced in this area by an early traumatisation (methamphetamine [MA] intoxication) and/or environmental deprivation
Keywords:
(isolated rearing [IR]), with the working hypothesis that such alterations will not occur in
Piriform cortex
an isolated fashion, but in mutual interaction. Indeed, acetylcholine fibre density is
Brain laterality
increased by IR in both hemispheres of the PirC (left: +22%, p < 0.01, right: + 21%, p < 0.05)
Isolation rearing
and the left hemisphere of the AOC (+13%, p < 0.05), while an early MA intoxication
Trauma
increases it in afterwards enriched–reared animals in the PirC (+ 14%/+ 17%, p < 0.05), but
Structural plasticity
decreases it in the AOC (− 18%/−22%, p < 0.001). The serotonin fibre density is increased by
Methamphetamine
IR in the right PirC of saline-treated (+ 13%, p < 0.01), but not of MA-traumatised gerbils. GABA and dopamine in the AOC show an inverse correlation, with dopamine innervation density being increased by IR (+ 30%, p < 0.001) and MA (+ 26%, p < 0.01), and GABA neuropil density being reduced. Furthermore, switches in hemispheric laterality occur in the AOC. These results demonstrate the complex recursive interactions in structural cortical plasticity. © 2007 Elsevier B.V. All rights reserved.
1.
Introduction
Neuromodulatory fibre systems interact with each other and with intrinsic targets by presynaptic and non-synaptic mechanisms. In this complex mobile, almost every transmitter can influence the activity of every other, such that alterations in one parameter can pull the whole neural system into a novel state. As the activity of neurons grinds into their structure especially during development, abnormal
physiological states during early life will shape an anatomically unbalanced brain (e.g. Hall, 1998 for review). Traditionally, most attention has been paid to the interactions of acetylcholine (ACh) and serotonin (5-HT) (Cassel and Jeltsch, 1995; Richter-Levin and Segal, 1993; Steckler and Sahgal, 1995), of dopamine (DA) and 5-HT (Daw et al., 2002; Kahn and Davidson, 1993; Poeggel et al., 2003; Reader and Dewar, 1999), and of DA and GABA (Benes, 1996; Nossoll et al., 1997). These four transmitters are also, next to glutamate, those with the
☆
We wish to dedicate this work to our daughter Clara who assisted the data acquisition by her perfect behaviour both before and after birth. ⁎ Corresponding author. Fax: +49 3641 949102. E-mail address: [email protected] (K. Lehmann). Abbreviations: 5-HT, 5-hydroxytryptamine, serotonin; ACh, acetylcholine; AOC, anterior olfactory cortex; DA, dopamine; ER, enriched rearing; GABA, γ-amino-butyric acid; IR, isolated rearing; MA, methamphetamine; PD, postnatal day; PirC, piriform cortex 0006-8993/$ – see front matter © 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.brainres.2007.01.020
38
BR A IN RE S EA RCH 1 1 41 ( 20 0 7 ) 3 7 –47
most significant impact on learning functions (Myhrer, 2003). But as has been pointed out before (Decker and McGaugh, 1991), focusing on just one or two transmitters in this complex net of interrelations will render an incomplete picture in which the emerging correlations may seem completely arbitrary. The anterior olfactory cortex (AOC, formerly known as anterior olfactory nucleus) and anterior piriform cortex (PirC) are very suitable structures for the study of hemispheric and transmitter interactions. The AOC, although not clearly layered, is based on radially oriented pyramidal cells (Haberly and Price, 1978), and is nowadays seen as a secondary sensory cortex. The PirC, which receives most of the AOC's output, handles olfactory information at the level of an association cortex (Haberly, 2001; Johnson et al., 2000). As a three-layered paleocortical structure, the PirC presents an old, simple and very clearly organised cytoarchitecture that invites the study of learning and plasticity (Haberly and Bower, 1998; Hasselmo and McGaughy, 2004). Pharmacological challenges have been shown to target the PirC preferentially among all cortical structures. A single systemic injection of methamphetamine (MA) induces a higher expression of c-Fos in the PirC than anywhere else in the brain (Umino et al., 1995), and escalating doses of cocaine increased dopamine and decreased serotonin levels in the PirC in a comprehensive study that measured
dopamine, serotonin and acetylcholine levels in several areas of the brain (Heidbreder et al., 1999). While these studies looked at acute effects of DA agonists, we have previously found long-lasting developmental alterations in various transmitter systems induced by a single early intoxication by methamphetamine (Dawirs et al., 1994; Lehmann et al., 2004; Neddens et al., 2003; Nossoll et al., 1997), as well as by isolated rearing (Lehmann et al., 2004; Neddens et al., 2001, 2003; Winterfeld et al., 1998). We therefore investigated the effects of these treatments on adult acetylcholine, serotonin, dopamine and GABA fibre densities in the AOC and PirC.
2.
Results
2.1.
Anterior piriform cortex
Acetylcholine, 5-HT and GABA innervate the anterior piriform cortex in distinct layers (Fig. 1). The ACh innervation appears dark and dense immediately below the lateral olfactory tract, which is followed by more loosely woven fibres in layer Ib and a dense, uniform innervation throughout the pyramidal layers (Fig. 1A). The 5-HT innervation also starts densely at the surface, but soon becomes weaker towards layer II (Fig. 1B). GABA neurites form a very fine web throughout the olfactory
Fig. 1 – Frontal sections through the anterior piriform cortex of an enriched reared (ER, left) and an isolation reared (IR, right) gerbil at 200× magnification. (A) Cholinergic innervation visualised by acetylcholine esterase staining. (B) Serotonergic innervation visualised by immunohistochemistry. (C) GABA neuropil visualised by immunohistochemistry. Arrowheads indicate exemplary baskets around non-stained, putatively glutamatergic cells. (D) Nissl staining provided for orientation. The frames indicate the width of measurement windows used. Please note that due to different shrinking, the sections may not be perfectly congruent.
39
BR A IN RE S E A RCH 1 1 41 ( 20 0 7 ) 3 7 –4 7
bulb, which is uniform in layer I of the PirC, whereas it spares non-GABAergic neurons in the pyramidal layers, showing typical baskets (Fig. 1C). In the quantitative comparison (cf. Table 1), the cholinergic innervation density is increased by isolation rearing and MA intoxication in both hemispheres of the PirC (Fig. 2A). ANOVA shows a highly significant effect of rearing (p < 0.01) and of layer (p < 0.001), and significant (p < 0.05) interactions of rearing with MA, layers and both. Contrast analysis proves the increase by rearing to be significant at a level of p < 0.01 in all layers of the left hemisphere, and at levels between 0.05 and 0.001 in the right hemisphere, except for layer Ib. The increase effected by early MA intoxication is significant in layer Ia (p < 0.001) and II/III (p < 0.05) of both hemispheres, but not detectable in layer Ib. Fibre densities in MA-treated IR animals are significantly different from those in ER control animals throughout (data not shown), but not from those in any other group. There is no indication of asymmetric ACh innervation in any group, nor of any alterations in such asymmetries. The density of 5-HT fibres in the PirC (Fig. 2B) is influenced by rearing and MA-intoxication (both p < 0.05), layer (p < 0.001) and an interaction of rearing, MA and hemisphere (p < 0.05), according to ANOVA. In detail, as revealed by post-hoc testing and contrast analysis, IR of saline-treated animals increases the innervation density in layer I of the right hemisphere (p < 0.01), with a similar, nearly significant effect in layer II/III (p = 0.065), which averages to a highly significant increase in the right PirC as a whole (p < 0.01). Compared to saline-injected IR animals, MA-intoxicated animals have less 5-HT fibres in the outer layer (p < 0.01) and in the average of both layers (p < 0.05) of the right hemisphere. There are no changes in the left hemisphere or in ER–MA animals. The one-sided increase in 5-HT fibre density in IR animals slightly distorts the
hemispheric symmetry, such that the lateralisation index in the outer layer of IR gerbils differs from that of ER gerbils at a level of p < 0.05 and from MA-intoxicated IR gerbils at a level of p = 0.08. GABA fibres in the PirC (Fig. 2C) are not detectably altered by rearing conditions or MA intoxication in our study, nor is there any lateralisation effect. The trend that is apparent from the graphics, i.e. that after MA intoxication, ER reduces, but IR increases the fibre density, achieves probability values below 0.1.
2.2.
Anterior olfactory cortex
The acetylcholine innervation in the medial AOC is dense and homogeneous, as is the 5-HT innervation. Dopamine fibres travel rostrally in this medial area and descend towards the ventromedial surface in a patch that spans just a few slices (i.e., approx. 300–400 μm in the rostrocaudal direction). GABA neurites and cells are present here as well as everywhere else in the olfactory bulb, with both dendritic and somatic innervation. For the quantification of the cholinergic innervation (Fig. 3A), ANOVA reveals highly significant effect of rearing (p < 0.001) and MA (p < 0.01), and a significant interaction of the two (p < 0.05). In contrast analysis, IR increases the ACh innervation of control animals in the left hemisphere (p < 0.05), and of MA-intoxicated animals in both hemispheres (p < 0.001). MA intoxication leads to a reduced fibre maturation in ER animals (p < 0.001), with no effect in IR animals. The serotonin innervation of the AOC (Fig. 3B) is not changed by IR or MA-intoxication, as far as the direct groupto-group comparison is concerned. ANOVA detects an interaction effect of rearing, MA and hemisphere (p < 0.05), but
Table 1 – Means ± S.E.M. for all evaluated transmitter densities in the piriform cortex (PirC) and anterior olfactory cortex (AOC) ER
PirC LI
ACh LIa ACh LIb 5-HT GABA PirC ACh LII/III 5-HT GABA PirC ACh total 5-HT GABA AOC ACh 5-HT GABA DA medial DA TT
ER–MA
IR
IR–MA
Left
Right
Left
Right
Left
Right
Left
Right
5.55± 0.19r**m*** 4.96± 0.27r** 9.44± 0.3 4.91± 0.34 6.84± 0.18r**m* 5.28± 0.26 4.79± 0.25 6.3 ± 0.17r**m* 6.76± 0.26 4.85± 0.29 8.56± 0.39r*m***
5.07± 0.3r***m*** 5.21± 0.44 9.31± 0.34r** 4.81± 0.26 6.67± 0.37r*m* 5.24± 0.26 4.9 ± 0.2 6.14± 0.34r*m* 6.69± 0.26r**
6.92± 0.25m*** 5.82± 0.37 8.88± 0.24 4.76± 0.18 7.73± 0.33m* 4.75± 0.15 4.53± 0.26 7.21± 0.32m* 6.22± 0.16 4.65± 0.21 7.0 ± 0.32r***m***
7.0 ± 0.23m*** 5.64± 0.39 9.18± 0.17 4.7 ± 0.19 7.74± 0.43m* 4.97± 0.13 4.73± 0.22 7.17± 0.38m* 6.47± 0.1 4.72± 0.19 6.58± 0.35r***m***
6.76± 0.36r** 6.36± 0.34r** 9.9 ± 0.53 4.73± 0.2 8.34± 0.52r** 5.77± 0.36 4.49± 0.24 7.69± 0.43r** 7.25± 0.41 4.57± 0.2 9.64± 0.21r*
6.95± 0.38r*** 6.07± 0.35 10.68 ± 0.24r**m* 4.8 ± 0.25 7.98± 0.34r* 5.81± 0.28 5.15± 0.19 7.42± 0.32r* 7.54± 0.25r**m*
6.54± 0.21 6.32± 0.18 9.32± 0.35m* 5.21± 0.34 8.2 ± 0.13 5.27± 0.17 5.09± 0.15 7.59± 0.13 6.72± 0.22m*
5.53± 0.21 5.5 ± 0.14 5.67± 0.26m**
5.72± 0.23 5.57± 0.23 5.59± 0.21r***
4.8 ± 0.3 8.73± 0.43 6.17± 0.38 5.36± 0.21 5.11± 0.34r***
6.69± 0.18 6.28± 0.24 9.24± 0.22 5.39± 0.14 8.22± 0.26 5.36± 0.35 5.31± 0.25 7.52± 0.2 6.74± 0.29 5.35± 0.18 8.89± 0.37r*** 5.84± 0.24 5.32± 0.21 4.61± 0.41
5.13± 0.23m*
5.53± 0.24r*
5.38± 0.15
5.71± 0.23 6.11± 0.37 4.76± 0.26r***
4.86± 0.21 8.44± 0.4m*** 5.72± 0.2 5.18± 0.22 5.68± 0.17 5.66± 0.26 4.55± 0.24r***m** 5.01± 0.24
4.23± 0.26r*
4.07± 0.21m*
4.61± 0.14
5.6 ± 0.22m*
5.14± 0.21 9.6 ± 0.24r*** 5.57± 0.21 5.59± 0.23 4.54± 0.34 5.25± 0.13
Superscript r indicates significant differences compared between rearing conditions, superscript m significant differences by methamphetamine treatment. For abbreviations see main text. *p < 0.05; **p < 0.01; ***p < 0.001.
40
BR A IN RE S EA RCH 1 1 41 ( 20 0 7 ) 3 7 –47
Fig. 2 – Fibre densities in the anterior piriform cortex. Values are given as means ± S.E.M. for the cholinergic (A), serotonergic (B) and GABAergic (C) innervation. *p < 0.05; **p < 0.01; ***p < 0.001. Asterisks without brackets refer to the neighbouring bars below them. Abbreviations: ER: enriched rearing; ER–MA: methamphetamine treatment (PD14) followed by enriched rearing; IR: isolated rearing; IRMA: methamphetamine treatment followed by isolated rearing.
there is no significant effect of either factor alone. The same holds true for the GABA fibres (Fig. 2C). Neither rearing conditions nor MA intoxication nor laterality induce any difference in their density by themselves, although the leftsided decline from ER controls to IR–MA animals is significant at a level of p = 0.07. Dopamine fibre densities were assessed separately in the medial AOC (same area as the other transmitters, Fig. 3D) and the ventral olfactory cortex/taenia tecta (Fig. 3E). The two regions were treated as repeated measurements in ANOVA, which points out an effect of rearing both by itself (p < 0.05) and in interaction with MA intoxication and region (p < 0.05/0.01, respectively). In the medial AOC, contrast analysis reveals an increase in fibre density by IR of saline-injected animals (p < 0.001) in both hemispheres, and by MA-intoxication of ER animals in the right hemisphere (p < 0.01), with no corresponding effect at all in the left AOC. Similarly, DA innervation
densities are increased by IR of control animals in the left ventral AOC and by MA in the right AOC (both p < 0.05), whereas in MA-intoxicated IR animals, the DA fibre density is reduced in the left hemisphere. Regression analysis of GABA and DA fibre densities in the medial AOC showed that there is a negative correlation (β = −0.36) of the two parameters in the left hemisphere at a level of p < 0.06, as seen across all animals. Within single groups, no significant correlations could be detected. The correlation of group means across both hemispheres is significant (p < 0.05, R2 = 0.585).
2.3.
Lateralisation of effects in the anterior olfactory cortex
The comparison of lateralisation indices revealed some astonishing effects in the AOC (Fig. 4). The ACh innervation, which is normally symmetrical with a leftward tendency,
BR A IN RE S E A RCH 1 1 41 ( 20 0 7 ) 3 7 –4 7
switches significantly (p = 0.06) from left to right by isolated rearing of MA-intoxicated animals, as compared to IR controls (Fig. 4A). Exactly the opposite effect can be observed in the 5-
41
HT innervation, which reverses from right > left in IR controls to left > right in IR–MA animals (p < 0.05, Fig. 4B). The GABA fibres, which showed no significant alteration by direct investigation, undergo a similar rightward switch as ACh fibres, with IR–MA animals in this case being significantly different from all other groups (Fig. 4C). In contrast to these conspicuous results, the hemispheric distribution of the DA innervation is not affected by any influence in either region (Figs. 4D, E). Likewise, none of the transmitter systems undergoes alterations in laterality in the PirC. The interhemispheric coupling is also influenced by epigenetic factors (Table 2). ACh fibre densities of the left and right PirC correlate positively in layer I of IR animals, whereas in the total mean of the PirC, there is an interhemispheric correlation in ER control animals that is lost by any treatment. In the AOC, ER control and MA animals show a positive correlation that is absent in IR gerbils. Serotonin fibre densities of the PirC are positively correlated in all groups, although not necessarily significantly so in all layers. As a rule, the correlation is strong in saline-injected animals of both rearing conditions and is weakened by MA intoxication. GABA and DA innervation show little or no interhemispheric correlation.
3.
Discussion
In the present study, we assessed the densities of acetylcholine (ACh), serotonin (5-HT), dopamine (DA) and GABA fibres in the anterior olfactory cortex (AOC) and anterior piriform cortex (PirC), as they adapt to an early methamphetamine (MA) intoxication and isolated rearing (IR). Isolation rearing increases the cholinergic innervation density of salineinjected animals in both areas, but only in the AOC of MApretreated gerbils, such that MA-intoxicated ER animals attain a higher fibre density than their saline-treated counterparts in the PirC, but a lower one in the AOC. There is a denser 5-HT innervation in the right PirC after IR, which is reversed or prevented by concomitant MA intoxication. GABAergic interneurons do not seem to react significantly to either influence, but their neuritic density is negatively correlated to the DA fibre density both across animals and across groups in the AOC. The DA fibres, finally, become denser in both evaluated areas after IR and, in the right hemisphere, after MA intoxication, while the innervation density is reduced in the left ventral AOC (taenia tecta) in MA-treated IR animals.
Fig. 3 – Fibre densities in the anterior olfactory cortex (AOC). Values are given as means ± S.E.M. for the cholinergic (A), serotonergic (B) GABAergic (C) and dopaminergic (D) innervation of the medial AOC and the dopaminergic innervation of the ventral AOC/taenia tecta (E). *p < 0.05; **p < 0.01; ***p < 0.001. Asterisks without brackets refer to the neighbouring bars below them. Abbreviations: ER: enriched rearing; ER–MA: methamphetamine treatment (PD14) followed by enriched rearing; IR: isolated rearing; IRMA: methamphetamine treatment followed by isolated rearing.
42
BR A IN RE S EA RCH 1 1 41 ( 20 0 7 ) 3 7 –47
This complex set of results presents a confusing puzzle at first glance, but on closer inspection reveals an interesting pattern, characterised principally by the following traits: a) Interrelations exist in the first place between ACh and 5-HT, on the one hand, and DA and GABA on the other. b) The olfactory cortices of the two hemispheres are morphologically interdependent.
3.1. cortex
Acetylcholine–serotonin interactions in the olfactory
The close interrelation of ACh and 5-HT in the brain has been the topic of much research (see Cassel and Jeltsch, 1995; Steckler and Sahgal, 1995 for review). The two neuromodulatory systems densely innervate the whole brain and together form a necessary basis for cortical wakefulness (Dringenberg and Vanderwolf, 1997, 1998). In the present study, both innervations of the PirC respond to rearing conditions in control animals, but remain inert after MA intoxication. It seems that the single pharmacological damage shifts the whole system into a new state in which stimulations produce no effect. On the other hand, ACh and 5-HT fibres respond complementarily in several ways. In the PirC, ACh fibres adapt to epigenetic influences most significantly in the left hemisphere, whereas the 5-HT innervation is responsive only on the right. IR of MA-pretreated animals antithetically switches the lateralisation indices of ACh and 5-HT in the AOC. And the interhemispheric coupling of ACh fibres is mostly dependent on rearing conditions, that of 5-HT fibres, in turn, on MA intoxication. It seems indeed that the two neuromodulators carve out their territories, or share their responsibilities. This is in line with previous findings from our group which show that the ACh innervation will react to epigenetic influences in the hemisphere which receives the relatively weaker 5-HT innervation, and in both hemispheres only if the 5-HT fibres are symmetrical (Lehmann et al., 2004; Neddens et al., 2003, 2004).
Table 2 – Interhemispheric coupling in the AOC
ACh
5-HT
GABA
Fig. 4 – Lateralisation indices (percent deviation from interhemispheric mean) for the cholinergic (A), serotonergic (B), GABAergic (C) and dopaminergic (D) innervation of the AOC and the dopaminergic innervation of the ventral AOC/ taenia tecta (E). Values are given as means ± S.E.M. *p < 0.05; **p < 0.01; ***p < 0.001. Abbreviations: ER: enriched rearing; ER–MA: methamphetamine treatment (PD14) followed by enriched rearing; IR: isolated rearing; IR MA: methamphetamine treatment followed by isolated rearing.
DA
PirC LIa PirC LIb PirC LII/III PirC total AOC PirC LI PirC LII/III PirC total AOC PirC LI PirC LII/III PirC total AOC AOC AOC/TT
ER–Saline
ER–MA
IR–Saline
IR–MA
0.419 0.632 0.621 0.773* 0.691* 0.664* 0.588 0.793** 0.564 0.468 0.628 0.632 0.595 0.348 0.593
0.271 0.580 0.530 0.521 0.817** 0.788** 0.353 0.465 0.245 0.678 0.421 0.571 0.606 −0.450 0.393
0.874** 0.547 0.500 0.633 −0.419 0.776* 0.845** 0.907** 0.641 0.610 0.160 0.709 0.462 0.133 0.441
0.780* 0.631* 0.250 0.537 0.471 0.628 0.876** 0.773* 0.825 0.952 0.475 0.636 0.851* 0.312 0.622
Correlations of left and right fibre densities within a region across all animals in one group. For abbreviations see main text. *p < 0.05; **p < 0.01; ***p < 0.001.
BR A IN RE S E A RCH 1 1 41 ( 20 0 7 ) 3 7 –4 7
In their functional involvement, ACh and 5-HT may not necessarily be seen as complementary. Both take part in olfactory learning (Hasselmo and Barkai, 1995; McLean et al., 1993; Patil et al., 1998; Rosin et al., 1999) (reviewed in McLean et al., 1996; Mendlin et al., 1999; Wilson et al., 2004), but in very different roles: Whereas ACh opens the network for extrinsic signals, 5-HT only facilitates its effect (Nilsson et al., 1988; Riekkinen et al., 1991), which is coherent with the general observation that ACh has a vastly more potent influence on learning abilities than 5-HT (Myhrer, 2003). Correspondingly, ACh and 5-HT exert quite different physiological effects at the neuronal/synaptic level (Hasselmo, 1995). It is also interesting to note that the PirC is reciprocally connected specifically to the orbital prefrontal cortex (Datiche and Cattarelli, 1996), where both 5-HT and ACh react in similar ways (Lehmann et al., 2004; Neddens et al., 2003, 2004). The absence of 5-HT effects in the AOC may be because this structure received fibres from the median raphe nucleus which are generally spared from amphetamine neurotoxicity (Mamounas et al., 1991), whereas the 5-HT projection to the PirC is afforded by the dorsal raphe (Datiche et al., 1995).
3.2. cortex
Dopamine–GABA interactions in the anterior olfactory
Dopamine fibres target only selected brain areas, in which they exert a powerful impact on morphology and function. Their regional appearance makes them a perfect candidate for interactions with local interneurons, although, of course, synaptic, physiological and anatomical interrelations with other transmitter systems have been shown (Kostrzewa et al., 1998; Moore et al., 1999; Nishikawa et al., 2002). The relationship of DAergic and serotoninergic neurons has received special attention, since they have reciprocal connections both between their somata and on the level of synapses (Ferré et al., 1994; Mendlin et al., 1999; Moukhles et al., 1997). A mutual shift of balances in some forebrain regions has been demonstrated both in the present paradigm and in similar combinations of early traumatic experience and isolated rearing (Braun et al., 2000; Lehmann et al., 2003; Poeggel et al., 2003). According to the present results, however, no indication of such a correlation can be found in the olfactory cortex. In contrast, it is conspicuous that the GABA plexus stays completely inert to epigenetic influences in the PirC which is devoid of DA innervation, but correlates inversely with the DA fibre density in the AOC, here even showing a weakly significant reduction in response to combined IR and MA intoxication. Such an interconnection of the two transmitters has been observed before in the medial prefrontal cortex. Here, MA intoxication leads to a suppressed maturation of DA fibres (Dawirs et al., 1994). As their net effect onto the local network is inhibitory, the lack is compensated by a strong sprouting of dendritic GABAergic synapses in layers III and V (Nossoll et al., 1997). Exactly the opposite situation is found in the AOC: IR and (in the right hemisphere) MA traumatisation increase the DA fibre density, with a compensatory reduction of GABA neuropil. This is in line with recent findings in the nucleus
43
accumbens (Lesting et al., 2005; Neddens et al., 2002), which lies directly adjacent to the evaluated area and is obviously the intermediate origin of the investigated DA fibres. As has been speculated earlier (Busche et al., 2004), there seems to be a seesaw relationship between the ventral tegmental area's cortical DA projection, which is reduced by IR and MA (Dawirs et al., 1994; Neddens et al., 2001; Winterfeld et al., 1998), and its subcortical projections, which are enhanced (Busche et al., 2004; Lesting et al., 2005). That lesioning of subcortical DA projections leads to a transient (after 2 weeks) loss, but longterm recovery of fibres has been demonstrated before (Bezard et al., 2000). We here show that this regenerative sprouting is at least partly dependent on environmental conditions, as it is absent in the ventral AOC of IR–MA animals.
3.3.
Epigenetic effects on cortical laterality
An anatomical and functional lateralisation of the brain exists not only in man, but also in rodents (Denenberg, 1983; Glick et al., 1977; Neddens et al., 2004). Put somewhat simplistically, the left hemisphere is more concerned with reward, inhibition of aggression, and learning of abstract stimuli, the right hemisphere, in contrast, with low mood, aggression, and concrete stimuli (Bianki, 1982; Denenberg, 1980; Garbanati et al., 1983; Glick et al., 1980). This pattern also applies to the human PirC, which shows more activation by pleasant tastes in the left side if subjects are hungry, and more right side activation if they are satiated (Del Parigi et al., 2002). Similarly, preferential activation of the left nucleus accumbens by pleasant and of the right accumbens by unpleasant odours has been demonstrated in rats (Besson and Louilot, 1995). Such a separation of tasks requires continuous interhemispheric communication. In the rodent olfactory cortex, this is subserved by strong contralateral projections of the AOC (Shipley et al., 1995). An anatomical lateralisation of brain structures has been found on the population level in rodents both for the prefrontal DA innervation (Slopsema et al., 1982) and for 5HT fibres in frontal and caudal areas of the cortex (Neddens et al., 2004). This latter paper from our work group found 5-HT fibre densities in the right PFC of ER gerbils to be approximately twice as high as in the left cortex, and 50% higher in the left than right entorhinal cortex. In the present study, in contrast, none of the evaluated transmitters shows any lateralisation in ER control animals. Since the medial and dorsal raphe nuclei project to both lateralised and nonlateralised cortex fields in no obvious segregation, it seems that laterality is a neocortical feature, possibly being related to the separation of higher-order functions. Nevertheless, the interhemispheric correlation values obtained in our study confirm that the ACh and 5-HT projections to the PirC and, for ACh, the AOC are interhemispherically coupled, i.e., they interact in a reciprocal feedback loop (Denenberg, 1980). This coupling can be disrupted by epigenetic influences, with ACh being more sensitive to rearing conditions, 5-HT more to MA intoxication. Interestingly, sensitisation with cocaine, which like MA is an indirect DA agonist, abolishes the interhemispheric coupling of DA transmission in the PirC (Heidbreder et al., 1999). Impairments in cerebral laterality have also been found after IR (Franklin
44
BR A IN RE S EA RCH 1 1 41 ( 20 0 7 ) 3 7 –47
and Paxinos, 1997; Heidbreder et al., 2000; Neddens et al., 2004). That an early MA intoxication prevents the adaptation of DA fibres in the AOC to environmental conditions exclusively in the right hemisphere may have to do with the right-sided 5HT-response in neighbouring areas (Neddens et al., 2004). On the other hand, the mesolimbic DA fibres are themselves asymmetrically involved in odour processing (Besson and Louilot, 1995), and also react in a lateralised fashion to early MA intoxication (Lesting et al., 2005). Thus, the unilateral DA fibre responses in the AOC have their origin in the differential functional involvement of the hemispheres. The most conspicuous but, unfortunately, also most confusing finding in this context is that the lateralisation indices for the ACh, 5-HT and GABA innervations of the AOC are reversed by concomitant MA intoxication and IR. This goes along with an induction of interhemispheric coupling in the GABA and, possibly (p < 0.09), the 5-HT innervation. A putative interpretation is that in these animals a fundamental reorganisation of the hemispheric cooperation has happened, by which the two AOCs develop rather as a fused entity than as interrelated individuals. The question whether such an effect can also be detected in other median raphe projection fields is of general relevance in so far as the combination of early traumatisation with later environmental deprivation bears some resemblance to the etiology of some psychiatric disorders (Kaufman and Charney, 2001; McCarley et al., 1999; Read et al., 2001).
3.4.
Conclusions
In this study, we have examined the structural interactions of four prominent transmitter innervations in the AOC and PirC, in brains that had matured under different epigenetic influences. A preferential relationship of ACh and 5-HT fibres on the one hand and DA and GABA innervation on the other hand could be demonstrated, but the results also indicate that the distortion of the neurochemical mobile within a certain brain region can not be sufficiently explained by interactions within that region. Instead, afferent influences always play an important role. Obviously, the findings are limited temporally, spatially and in the choice of transmitters. But they provide a first tentative step towards understanding the complex recurrent interactions of transmitter systems in the brain, which may hopefully be incorporated into a more comprehensive investigation of structural plasticity in the brain.
4.
Experimental procedures
4.1.
Animals and rearing conditions
Male gerbils were bred in our facilities either in standard cages or in semi-naturally structured pens of 1 m2 (for details, see Winterfeld et al., 1998). On postnatal day (PD) 14, some of the pups from each condition were injected with a single dose of 50 mg/kg methamphetamine (MA); the others received a saline injection. Applied at this age, this intoxication preferentially lesions prefrontal DA terminals (Teuchert-Noodt and
Dawirs, 1991) and results in their reduced maturation into adulthood (Dawirs et al., 1994). At weaning (PD30), animals that had been born in standard cages were assigned to isolated rearing (IR) conditions, staying singly in standard makrolon cages (type 3), while those from enriched compounds were reared under enriched conditions (ER) in sibling groups in compounds similar to the ones they were born in. Under both sets of conditions there was a bedding of wood shavings, and food and water were provided ad libitum. All gerbils were kept on natural day/night cycles. The experiments were carried out in accordance with the European Communities Council Directive of 24 November 1986 (86/609/ EEC). Formal approval to conduct the experiments described has been obtained from the animal subjects review board of our institution and could be provided upon request. All efforts were made to minimize the number of animals used and their suffering. For the present study we used samples that had been prepared during previous projects, such that no animal was killed for this study alone. Yet, all samples for one transmitter were processed during a short time span, with groups randomly mixed. Due to the imponderabilities of immunohistochemistry, the sample sizes varied considerably even between hemispheres and neighbouring laminae. Usually, between seven and ten animals were evaluated in each group.
4.2.
DA and GABA immunohistochemistry
The method has been described in detail elsewhere (Busche et al., 2004). In short, the animals were transcardially perfused on PD 90 under deep choralhydrate anesthesia with 0.05 M cacodylate buffer (pH 6.2) containing 1% sodium metabisulfite, followed by 5% glutaraldehyde with 1% sodium metabisulfite in 0.1 M cacodylate buffer (pH 7.5), and finally by 0.05 M Trisbuffered saline (TBS) with 1% sodium metabisulfite (pH 7.5). The brains were dissected, and 40-μm-thick frontal sections
Fig. 5 – Clip out of a schematic drawing of the olfactory peduncle (Franklin and Paxinos, 1997). Evaluated areas are marked by shaded rectangles. Abbreviations: AOCd/v: anterior olfactory cortex, dorsal/ventral part; ca: commissura anterior; TTd/v: dorsal/ventral taenia tecta; PirC: piriform cortex.
BR A IN RE S E A RCH 1 1 41 ( 20 0 7 ) 3 7 –4 7
cut with a vibratome (Leica VT 1000S, Nussloch, Germany) and collected in TBS at 4 °C. Alternating sections were used for GABA and DA immunohistochemistry. They were incubated with the primary antibody (rabbit anti-dopamine, DiaSorin, Stillwater, MN, 1:600; rabbit anti-GABA, DiaSorin, 1:5000) over two nights. Following avidin–biotin reaction (Sigma, St. Louis, MO), staining was done by 0.05% 3,3-diaminobenzidine (DAB, Sigma) and 0.01% H2O2 in TBS.
4.3.
Serotonin immunohistochemistry
The method has been extensively described elsewhere (Neddens et al., 2003). In brief, the animals were transcardially perfused with 100 ml 0.1 M phosphate buffer (pH 7.2), followed by 500 ml of 4% phosphate-buffered paraformaldehyde (pH 7.4) on PD110 under deep choralhydrate anesthesia. Frontal sections of 20 μm were taken on a frigocut and every third section collected in ice-cold phosphate-buffered saline (PBS, pH 7.4). The slices were incubated for 10 min with 1% H2O2 to reduce background staining and blocked with 10% normal goat serum in PBS containing 0.3% Triton X-100. The primary antibody (rabbit anti-serotonin, Incstar) was diluted 1:20,000 in PBS with 1% NGS and 0.3% Triton X-100, and applied for 18 h at 4 °C. Following avidin–biotin reaction (Sigma, St. Louis, MO), the slices were stained by DAB in TBS.
4.4.
Acetylcholine enzyme histochemistry
The method used was described in detail elsewhere (Lehmann et al., 2004). Briefly, animals were transcardially perfused on PD90 under deep choralhydrate anesthesia (1.7 g/kg, i.p.) with 100 ml 0.1 M phosphate buffer (pH 7.2), followed by 500 ml of 4% phosphate-buffered paraformaldehyde (pH 7.4). The brains were removed and postfixed for 4 days at 4 °C, after which they were immersed in 30% sucrose at 4 °C for a further 2 days. Frontal sections of 50 μm were taken on a frigomobile and every other section collected in ice-cold 0.1 M maleate buffer (MB, pH 6.0). The staining procedure was carried out according to the method by Tago et al. (1986). The sections were incubated for 45 min at RT in the staining solution containing, in 50 ml 0.1 M MB, 320 μg K3Fe(CN)6, 940 μg CuSO4, 2,94 mg C6H5Na3O7·2H2O and 1 mg acetylcholine thioiodide as a substrate for acetylcholinesterase. They were stained in a solution containing 0.05% DAB, 0.01% H2O2 and 0,15% ammonium nickel sulfate (Fluka, Milwaukee, WI) in TBS.
4.5.
Computer-aided image analysis
Pictures were taken in one focal plane on a light microscope (for ACh, 5-HT and DA sections: Polyvar, Reichert-Jung; for GABA sections: BX61, Olympus) linked to a digital camera (ProgRes 3008 mF, Jenoptik, Jena or colorview, SOFT imaging systems, Münster, respectively). All images were assessed using a software for image analysis (KS300, Zeiss, Jenoptik, Jena). The investigators were blind to the treatment conditions of the specimens. In the anterior olfactory cortex (AOC), all pictures were taken at a magnification of 250× and a resolution of 768 × 580 square pixels at a position immediately medial to the commissura anterior (Fig. 5). For ACh and 5-HT, which
45
provide a very uniform innervation, the whole area of the image in five consecutive sections was evaluated, while regions-of-interest containing the densest and best-stained innervation were defined for DA (three sections) and GABA (five sections). For DA fibre densities, an additional area in the ventromedial olfactory cortex/taenia tecta (three sections) was investigated, where fibres descend to the surface in a narrow patch. In the piriform cortex (Fig. 5), pictures from five consecutive sections were taken either at a magnification of 250× and a resolution of 768 × 580 square pixels throughout the whole thickness of the cortex (ACh, 5-HT), or at 600× and 2080 × 1544 square pixels in both the molecular and the cellular layer (GABA). In the first case, measurement windows within differentially innervated laminae were defined: For ACh, the densely innervated layer Ia, loosely innervated layer Ib and uniformly, densely innervated layer II/III were assessed separately. For 5-HT, layer I and layer II/III were differentiated. The subsequent measurement procedure differed slightly depending on area and transmitter. In general, a reference area was defined by the exclusion of blood vessels. The image was enhanced by high pass filtering and contrast augmentation. Fibres were then recognised by the so-called “valleys function”, which comes as part of the software package. This function combines a Gaussian filtering and a subsequent so-called “Gerig-operator”, which is an algorithm to compare the grey values of neighbouring pixels irrespective of global patchiness. It implies the setting of a threshold for each picture by the investigator, which was done in constant optical check against the original image. In effect, fibres were depicted as lines of one pixel width, such that possible changes in fibre morphology would not influence the measurement. The fibre area was calculated as a percentage of the reference area.
4.6.
Statistical evaluation
The data of each layer within cortical area were averaged across all sections in each animal. Total average densities across all layers of the piriform cortex were calculated with respect to the layers' different surface areas. Additionally, lateralisation indices were calculated as 100 * (right − left) / [(right + left) / 2] (i.e., as percent deviation from mean) at each position and averaged in the same way. Statistical comparison was done by analysis of variance (ANOVA) with repeated measurements, using two main factors of rearing and treatment, and two repeated measurement factors of hemispheres and (in the PirC) layers. This was followed by post-hoc analysis (LSD test) and contrast analysis. As DA and GABA sections were obtained from the same animals, correlations between the respective fibre densities in the AOC were computed and their strength assessed by regression analysis, both across all animals and within each group. The same was done to determine the interhemispheric coupling of left and right fibre densities (cp. Denenberg, 1980). All calculations were done using the programme Statistica 6 (StatSoft, Tulsa, USA). In the figures, data are depicted as means ± standard error of means (S.E.M.). The levels of significance were set at p < 0.05 (*), p < 0.01 (**) and p < 0.001 (***). In some cases, differences which are significant at a level between 0.05 < p < 0.1 are also drawn into consideration.
46
BR A IN RE S EA RCH 1 1 41 ( 20 0 7 ) 3 7 –47
Acknowledgments The authors thank Dr. Jörg Neddens, Dr. Andrea Busche, Mr. Jörg Lesting and Mrs. Anja Bagorda for technical assistance and inspiring discussions. Thanks are also due to Mr. John O. Ball for correcting the English.
REFERENCES
Benes, F.M., 1996. The role of stress and dopamine–GABA interactions in the vulnerability for schizophrenia. J. Psychiatr. Res. 31 (2), 257–275. Besson, C., Louilot, A., 1995. Asymmetrical involvement of mesolimbic dopaminergic neurons in affective perception. Neuroscience 68, 963–968. Bezard, E., Dovero, S., Imbert, C., Boraud, T., Gross, C.E., 2000. Spontaneous long-term compensatory dopaminergic sprouting in MPTP-treated mice. Synapse 38 (3), 363–368. Bianki, V., 1982. Lateralization of concrete and abstract characteristics analysis in the animal brain. Int. J. Neurosci. 17, 233–241. Braun, K., Lange, E., Metzger, M., Poeggel, G., 2000. Maternal separation followed by early social deprivation affects the development of monoaminergic fiber systems in the medial prefrontal cortex of Octodon degus. Neuroscience 95 (1), 309–318. Busche, A., Polascheck, D., Lesting, J., Neddens, J., Teuchert-Noodt, G., 2004. Developmentally induced imbalance of dopaminergic fibre densities in limbic brain regions of gerbils (Meriones unguiculatus). J. Neural Transm. 111 (4), 451–463. Cassel, J.C., Jeltsch, H., 1995. Serotonergic modulation of cholinergic function in the central nervous system: cognitive implications. Neuroscience 69, 1–41. Datiche, F., Cattarelli, M., 1996. Reciprocal and topographic connections between the piriform and prefrontal cortices in the rat: a tracing study using the B subunit of the cholera toxin. Brain Res. Bull. 41 (6), 391–398. Datiche, F., Luppi, P.H., Cattarelli, M., 1995. Serotonergic and non-serotonergic projections from the raphe nuclei to the piriform cortex in the rat: a cholera toxin B subunit (CTb) and 5-HT immunohistochemical study. Brain Res. 671 (1), 27–37. Daw, N.D., Kakade, S., Dayan, P., 2002. Opponent interactions between serotonin and dopamine. Neural Netw. 15 (4–6), 603–616. Dawirs, R.R., Teuchert-Noodt, G., Czaniera, R., 1994. The postnatal maturation of dopamine innervation in the prefrontal cortex of gerbils (Meriones unguiculatus) is sensitive to an early single dose of methamphetamine. A quantitative immunocytochemical study. J. Hirnforsch. 35 (2), 195–204. Decker, M.W., McGaugh, J.L., 1991. The role of interactions between the cholinergic system and other neuromodulatory systems in learning and memory. Synapse 7 (2), 151–168. Del Parigi, A., Chen, K., Salbe, A.D., Gautier, J.F., Ravussin, E., Reiman, E.M., Tataranni, P.A., 2002. Tasting a liquid meal after a prolonged fast is associated with preferential activation of the left hemisphere. NeuroReport 13 (9), 1141–1145. Denenberg, V.H., 1980. General systems theory, brain organization, and early experiences. Am. J. Physiol. 238 (1), R3–R13. Denenberg, V.H., 1983. Lateralization of function in rats. Am. J. Physiol. 245, R505–R509. Dringenberg, H.C., Vanderwolf, C.H., 1997. Neocortical activation: modulation by multiple pathways acting on central cholinergic and serotonergic systems. Exp. Brain Res. 116, 160–174. Dringenberg, H.C., Vanderwolf, C.H., 1998. Involvement of direct
and indirect pathways in electrocorticographic activation. Neurosci. Biobehav. Rev. 22 (2), 243–257. Ferré, S., Cortés, R., Artigas, F., 1994. Dopaminergic regulation of the serotonergic raphe–striatal pathway: microdialysis studies in freely moving rats. J. Neurosci. 14 (8), 4839–4846. Franklin, K.B.J., Paxinos, G., 1997. The Mouse Brain in Stereotaxic Coordinates. Academic Press, San Diego. Garbanati, J.A., Sherman, G.F., Rosen, G.D., Hofmann, M., Yutzey, D.A., Denenberg, V.H., 1983. Handling in infancy, brain laterality and muricide in rats. Behav. Brain Res. 7 (3), 351–359. Glick, S.D., Zimmerberg, B., Jerussi, T.P., 1977. Adaptive significance of laterality in the rodent. Ann. N. Y. Acad. Sci. 299, 273–280. Glick, S.D., Weaver, L.M., Meibach, R.C., 1980. Lateralization of reward in rats: differences in reinforcing thresholds. Science 207 (4435), 1093–1095. Haberly, L.B., 2001. Parallel-distributed processing in olfactory cortex: new insights from morphological and physiological analysis of neuronal circuitry. Chem. Senses 26 (5), 551–576. Haberly, L.B., Bower, J.M., 1998. Olfactory cortex: model circuit for study of associative memory? Trends Neurosci. 12 (7), 258–264. Haberly, L.B., Price, J.L., 1978. Association and commissural fiber systems of the olfactory cortex of the rat: II. Systems originating in the olfactory peduncle. J. Comp. Neurol. 181, 781–808. Hall, F.S., 1998. Social deprivation of neonatal, adolescent, and adult rats has distinct neurochemical and behavioral consequences. Crit. Rev. Neurobiol. 12 (1 and 2), 129–162. Hasselmo, M.E., 1995. Neuromodulation and cortical function: modeling the physiological basis of behavior. Behav. Brain Res. 67 (1), 1–27. Hasselmo, M.E., Barkai, E., 1995. Cholinergic modulation of activity-dependent synaptic plasticity in the piriform cortex and associative memory function in a network biophysical simulation. J. Neurosci. 15 (10), 6592–6604. Hasselmo, M.E., McGaughy, J., 2004. High acetylcholine levels set circuit dynamics for attention and encoding and low acetylcholine levels set dynamics for consolidation. Prog. Brain Res. 145, 207–231. Heidbreder, C.A., Oertle, T., Feldon, J., 1999. Dopamine and serotonin imbalances in the left anterior cingulate and pyriform cortices following the repeated intermittent administration of cocaine. Neuroscience 89 (3), 701–715. Heidbreder, C.A., Weiss, I.C., Domeney, A.M., Pryce, C., Homberg, J., Hedou, G., Feldon, J., Moran, M.C., 2000. Behavioral, neurochemical and endocrinological characterization of the early social isolation syndrome. Neuroscience 100 (4), 749–768. Johnson, D.M., Illig, K.R., Behan, M., Haberly, L.B., 2000. New features of connectivity in piriform cortex visualized by intracellular injection of pyramidal cells suggests that “primary” olfactory cortex functions like “association” cortex in other sensory systems. J. Neurosci. 20 (18), 6974–6982. Kahn, R.S., Davidson, M., 1993. Serotonin, dopamine and their interactions in schizophrenia. Psychopharmacology 112 (1 Suppl), S1–S4. Kaufman, J., Charney, D., 2001. Effects of early stress on brain structure and function: implications for understanding the relationship between child maltreatment and depression. Dev. Psychopathol. 13, 451–471. Kostrzewa, R.M., Reader, T.A., Descarries, L., 1998. Serotonin neural adaptations to ontogenetic loss of dopamine neurons in rat brain. J. Neurochem. 70 (3), 889–898. Lehmann, K., Lesting, J., Polascheck, D., Teuchert-Noodt, G., 2003. Serotonin fibre densities in subcortical areas: differential effects of isolated rearing and methamphetamine. Dev. Brain Res. 147 (1–2), 123–133. Lehmann, K., Hundsdörfer, B., Hartmann, T., Teuchert-Noodt, G., 2004. The acetylcholine fiber density of the neocortex is altered
BR A IN RE S E A RCH 1 1 41 ( 20 0 7 ) 3 7 –4 7
by isolated rearing and early methamphetamine intoxication in rodents. Exp. Neurol. 189 (1), 131–140. Lesting, J., Neddens, J., Busche, A., Teuchert-Noodt, G., 2005. Hemisphere-specific effects on serotonin but not dopamine innervation in the nucleus accumbens of gerbils caused by isolated rearing and a single early methamphetamine challenge. Brain Res. 1035, 168–176. Mamounas, L.A., Mullen, C.A., O'Hearn, E., Molliver, M.E., 1991. Dual serotoninergic projections to forebrain in the rat: morphologically distinct 5-HT axon terminals exhibit differential vulnerability to neurotoxic amphetamine derivatives. J. Comp. Neurol. 314 (3), 558–586. McCarley, R.W., Wible, C.G., Frumin, M., Hirayasu, Y., Levitt, J.L., Fischer, I.A., Shenton, M.E., 1999. MRI anatomy of schizophrenia. Biol. Psychiatry 45, 1099–1119. McLean, J.H., Darby-King, A., Sullivan, R.M., King, S.R., 1993. Serotonergic influence on olfactory learning in the neonate rat. Behav. Neural Biol. 60 (2), 152–162. McLean, J.H., Darby-King, A., Hodge, E., 1996. 5-HT2 receptor involvement in conditioned olfactory learning in the neonate rat pup. Behav. Neurosci. 110 (6), 1426–1434. Mendlin, A., Martín, F.J., Jacobs, B.L., 1999. Dopaminergic input is required for increases in serotonin output produced by behavioral activation: an in vivo microdialysis study in rat forebrain. Neuroscience 93 (3), 897–905. Moore, H., Fadel, J., Sarter, M., Bruno, J.P., 1999. Role of accumbens and cortical dopamine receptors in the regulation of cortical acetylcholine release. Neuroscience 88 (3), 811–822. Moukhles, H., Bosler, O., Bolam, J.P., Vallée, A., Umbriaco, D., Geffards, M., Doucet, G., 1997. Quantitative and morphometric data indicate precise cellular interactions between serotonin terminals and postsynaptic targets in rat substantia nigra. Neuroscience 76 (4), 1159–1171. Myhrer, T., 2003. Neurotransmitter systems involved in learning and memory in the rat: a meta-analysis based on studies of four behavioral tasks. Brain Res. Rev. 41 (2–3), 268–287. Neddens, J., Brandenburg, K., Teuchert-Noodt, G., Dawirs, R.R., 2001. Differential environment alters ontogeny of dopamine innervation of the orbital prefrontal cortex in gerbils. J. Neurosci. Res. 63 (2), 209–213. Neddens, J., Lesting, J., Dawirs, R.R., Teuchert-Noodt, G., 2002. An early methamphetamine challenge suppresses the maturation of dopamine fibres in the nucleus accumbens of gerbils: on the significance of rearing conditions. J. Neural Transm. 109 (2), 141–155. Neddens, J., Bagorda, F., Busche, A., Horstmann, S., Moll, G.H., Dawirs, R.R., Teuchert-Noodt, G., 2003. Epigenetic factors differentially influence postnatal maturation of serotonin (5-HT) innervation in cerebral cortex of gerbils: interaction of rearing conditions and early methamphetamine challenge. Dev. Brain Res. 146 (1–2), 119–130. Neddens, J., Dawirs, R.R., Bagorda, F., Busche, A., Horstmann, S., Teuchert-Noodt, G., 2004. Postnatal maturation of cortical serotonin lateral asymmetry in gerbils is vulnerable to both environmental and pharmacological epigenetic challenges. Brain Res. 1021, 200–208. Nilsson, O.G., Strecker, R.E., Daszuta, A., Bjorklund, A., 1988. Combined cholinergic and serotonergic denervation of the forebrain produces severe deficits in a spatial learning task in the rat. Brain Res. 453 (1–2), 235–246.
47
Nishikawa, J., Chung, Y.-C., Li, Z., Dai, J., Meltzer, H.Y., 2002. Cholinergic modulation of basal and amphetamine-induced dopamine release in rat medial prefrontal cortex and nucleus accumbens. Brain Res. 958, 176–184. Nossoll, M., Teuchert-Noodt, G., Dawirs, R.R., 1997. A single dose of methamphetamine in neonatal gerbils affects adult prefrontal gamma-aminobutyric acid innervation. Eur. J. Pharmacol. 340 (2–3), R3–R5. Patil, M.M., Linster, C., Lubenov, E., Hasselmo, M.E., 1998. Cholinergic agonist carbachol enables associative long-term potentiation in piriform cortex slices. J. Neurophysiol. 80 (5), 2467–2474. Poeggel, G., Nowicki, L., Braun, K., 2003. Early social deprivation alters monoaminergic afferents in the orbital prefrontal cortex of Octodon degus. Neuroscience 116 (3), 617–620. Reader, T.A., Dewar, K.M., 1999. Effects of denervation and hyperinnervation on dopamine and serotonin systems in the rat neostriatum: implications for human Parkinson's disease. Neurochem. Int. 34, 1–21. Read, J., Perry, B.D., Moskowitz, A., Connolly, J., 2001. The contribution of early traumatic events to schizophrenia in some patients: a traumagenic neurodevelopmental model. Psychiatry 64 (4), 319–345. Richter-Levin, G., Segal, M., 1993. Age-related cognitive deficits in rats are associated with a combined loss of cholinergic and serotonergic functions. Ann. N. Y. Acad. Sci. 695, 254–257. Riekkinen Jr., P., Sirvio, J., Valjakka, A., Miettinen, R., Riekkinen, P., 1991. Pharmacological consequences of cholinergic plus serotonergic manipulations. Brain Res. 552 (1), 23–26. Rosin, J.F., Datiche, F., Cattarelli, M., 1999. Modulation of the piriform cortex activity by the basal forebrain: an optical recording study in the rat. Brain Res. 820 (1–2), 105–111. Shipley, M.T., McLean, J.H., Ennis, M., 1995. Olfactory system. In: Paxinos, G. (Ed.), The Rat Brain. Academic Press, San Diego, pp. 899–926. Slopsema, J.S., Van der Gugten, J., de Bruin, J.P., 1982. Regional concentrations of noradrenaline and dopamine in the frontal cortex of the rat: dopaminergic innervation of the prefrontal subareas and lateralization of prefrontal dopamine. Brain Res. 250 (1), 197–200. Steckler, T., Sahgal, A., 1995. The role of serotonergic–cholinergic interactions in the mediation of cognitive behaviour. Behav. Brain Res. 67, 165–199. Tago, H., Kimura, H., Maeda, T., 1986. Visualization of detailed acetylcholinesterase fiber and neuron staining in rat brain by a sensitive histochemical procedure. J. Histochem. Cytochem. 34, 1431–1438. Teuchert-Noodt, G., Dawirs, R.R., 1991. Age-related toxicity in prefrontal cortex and caudate–putamen complex of gerbils (Meriones unguiculatus). Neuropharmacology 30 (7), 733–743. Umino, A., Nishikawa, T., Takahashi, K., 1995. Methamphetamine-induced nuclear c-fos in rat brain regions. Neurochem. Int. 26 (1), 85–90. Wilson, D.A., Fletcher, M.L., Sullivan, R.M., 2004. Acetylcholine and olfactory perceptual learning. Learn. Mem. 11 (1), 28–34. Winterfeld, K.T., Teuchert-Noodt, G., Dawirs, R.R., 1998. Social environment alters both ontogeny of dopamine innervation of the medial prefrontal cortex and maturation of working memory in gerbils (Meriones unguiculatus). J. Neurosci. Res. 52 (2), 201–209.
a v a i l a b l e a t w w w. s c i e n c e d i r e c t . c o m
w w w. e l s e v i e r. c o m / l o c a t e / b r a i n r e s
Research Report
Transmitter balances in the olfactory cortex: Adaptations to early methamphetamine trauma and rearing environment☆ Konrad Lehmann⁎, Devrim Lehmann Institute for General Zoology and Animal Physiology, Erbertstr. 1, 07743 Jena, Germany
A R T I C LE I N FO
AB S T R A C T
Article history:
The olfactory cortex, comprising the anterior olfactory cortex (AOC) and the anterior
Accepted 5 January 2007
piriform cortex (PirC), is a model system for the study of neural plasticity. We investigated
Available online 12 January 2007
the structural imbalances of different transmitter systems induced in this area by an early traumatisation (methamphetamine [MA] intoxication) and/or environmental deprivation
Keywords:
(isolated rearing [IR]), with the working hypothesis that such alterations will not occur in
Piriform cortex
an isolated fashion, but in mutual interaction. Indeed, acetylcholine fibre density is
Brain laterality
increased by IR in both hemispheres of the PirC (left: +22%, p < 0.01, right: + 21%, p < 0.05)
Isolation rearing
and the left hemisphere of the AOC (+13%, p < 0.05), while an early MA intoxication
Trauma
increases it in afterwards enriched–reared animals in the PirC (+ 14%/+ 17%, p < 0.05), but
Structural plasticity
decreases it in the AOC (− 18%/−22%, p < 0.001). The serotonin fibre density is increased by
Methamphetamine
IR in the right PirC of saline-treated (+ 13%, p < 0.01), but not of MA-traumatised gerbils. GABA and dopamine in the AOC show an inverse correlation, with dopamine innervation density being increased by IR (+ 30%, p < 0.001) and MA (+ 26%, p < 0.01), and GABA neuropil density being reduced. Furthermore, switches in hemispheric laterality occur in the AOC. These results demonstrate the complex recursive interactions in structural cortical plasticity. © 2007 Elsevier B.V. All rights reserved.
1.
Introduction
Neuromodulatory fibre systems interact with each other and with intrinsic targets by presynaptic and non-synaptic mechanisms. In this complex mobile, almost every transmitter can influence the activity of every other, such that alterations in one parameter can pull the whole neural system into a novel state. As the activity of neurons grinds into their structure especially during development, abnormal
physiological states during early life will shape an anatomically unbalanced brain (e.g. Hall, 1998 for review). Traditionally, most attention has been paid to the interactions of acetylcholine (ACh) and serotonin (5-HT) (Cassel and Jeltsch, 1995; Richter-Levin and Segal, 1993; Steckler and Sahgal, 1995), of dopamine (DA) and 5-HT (Daw et al., 2002; Kahn and Davidson, 1993; Poeggel et al., 2003; Reader and Dewar, 1999), and of DA and GABA (Benes, 1996; Nossoll et al., 1997). These four transmitters are also, next to glutamate, those with the
☆
We wish to dedicate this work to our daughter Clara who assisted the data acquisition by her perfect behaviour both before and after birth. ⁎ Corresponding author. Fax: +49 3641 949102. E-mail address: [email protected] (K. Lehmann). Abbreviations: 5-HT, 5-hydroxytryptamine, serotonin; ACh, acetylcholine; AOC, anterior olfactory cortex; DA, dopamine; ER, enriched rearing; GABA, γ-amino-butyric acid; IR, isolated rearing; MA, methamphetamine; PD, postnatal day; PirC, piriform cortex 0006-8993/$ – see front matter © 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.brainres.2007.01.020
38
BR A IN RE S EA RCH 1 1 41 ( 20 0 7 ) 3 7 –47
most significant impact on learning functions (Myhrer, 2003). But as has been pointed out before (Decker and McGaugh, 1991), focusing on just one or two transmitters in this complex net of interrelations will render an incomplete picture in which the emerging correlations may seem completely arbitrary. The anterior olfactory cortex (AOC, formerly known as anterior olfactory nucleus) and anterior piriform cortex (PirC) are very suitable structures for the study of hemispheric and transmitter interactions. The AOC, although not clearly layered, is based on radially oriented pyramidal cells (Haberly and Price, 1978), and is nowadays seen as a secondary sensory cortex. The PirC, which receives most of the AOC's output, handles olfactory information at the level of an association cortex (Haberly, 2001; Johnson et al., 2000). As a three-layered paleocortical structure, the PirC presents an old, simple and very clearly organised cytoarchitecture that invites the study of learning and plasticity (Haberly and Bower, 1998; Hasselmo and McGaughy, 2004). Pharmacological challenges have been shown to target the PirC preferentially among all cortical structures. A single systemic injection of methamphetamine (MA) induces a higher expression of c-Fos in the PirC than anywhere else in the brain (Umino et al., 1995), and escalating doses of cocaine increased dopamine and decreased serotonin levels in the PirC in a comprehensive study that measured
dopamine, serotonin and acetylcholine levels in several areas of the brain (Heidbreder et al., 1999). While these studies looked at acute effects of DA agonists, we have previously found long-lasting developmental alterations in various transmitter systems induced by a single early intoxication by methamphetamine (Dawirs et al., 1994; Lehmann et al., 2004; Neddens et al., 2003; Nossoll et al., 1997), as well as by isolated rearing (Lehmann et al., 2004; Neddens et al., 2001, 2003; Winterfeld et al., 1998). We therefore investigated the effects of these treatments on adult acetylcholine, serotonin, dopamine and GABA fibre densities in the AOC and PirC.
2.
Results
2.1.
Anterior piriform cortex
Acetylcholine, 5-HT and GABA innervate the anterior piriform cortex in distinct layers (Fig. 1). The ACh innervation appears dark and dense immediately below the lateral olfactory tract, which is followed by more loosely woven fibres in layer Ib and a dense, uniform innervation throughout the pyramidal layers (Fig. 1A). The 5-HT innervation also starts densely at the surface, but soon becomes weaker towards layer II (Fig. 1B). GABA neurites form a very fine web throughout the olfactory
Fig. 1 – Frontal sections through the anterior piriform cortex of an enriched reared (ER, left) and an isolation reared (IR, right) gerbil at 200× magnification. (A) Cholinergic innervation visualised by acetylcholine esterase staining. (B) Serotonergic innervation visualised by immunohistochemistry. (C) GABA neuropil visualised by immunohistochemistry. Arrowheads indicate exemplary baskets around non-stained, putatively glutamatergic cells. (D) Nissl staining provided for orientation. The frames indicate the width of measurement windows used. Please note that due to different shrinking, the sections may not be perfectly congruent.
39
BR A IN RE S E A RCH 1 1 41 ( 20 0 7 ) 3 7 –4 7
bulb, which is uniform in layer I of the PirC, whereas it spares non-GABAergic neurons in the pyramidal layers, showing typical baskets (Fig. 1C). In the quantitative comparison (cf. Table 1), the cholinergic innervation density is increased by isolation rearing and MA intoxication in both hemispheres of the PirC (Fig. 2A). ANOVA shows a highly significant effect of rearing (p < 0.01) and of layer (p < 0.001), and significant (p < 0.05) interactions of rearing with MA, layers and both. Contrast analysis proves the increase by rearing to be significant at a level of p < 0.01 in all layers of the left hemisphere, and at levels between 0.05 and 0.001 in the right hemisphere, except for layer Ib. The increase effected by early MA intoxication is significant in layer Ia (p < 0.001) and II/III (p < 0.05) of both hemispheres, but not detectable in layer Ib. Fibre densities in MA-treated IR animals are significantly different from those in ER control animals throughout (data not shown), but not from those in any other group. There is no indication of asymmetric ACh innervation in any group, nor of any alterations in such asymmetries. The density of 5-HT fibres in the PirC (Fig. 2B) is influenced by rearing and MA-intoxication (both p < 0.05), layer (p < 0.001) and an interaction of rearing, MA and hemisphere (p < 0.05), according to ANOVA. In detail, as revealed by post-hoc testing and contrast analysis, IR of saline-treated animals increases the innervation density in layer I of the right hemisphere (p < 0.01), with a similar, nearly significant effect in layer II/III (p = 0.065), which averages to a highly significant increase in the right PirC as a whole (p < 0.01). Compared to saline-injected IR animals, MA-intoxicated animals have less 5-HT fibres in the outer layer (p < 0.01) and in the average of both layers (p < 0.05) of the right hemisphere. There are no changes in the left hemisphere or in ER–MA animals. The one-sided increase in 5-HT fibre density in IR animals slightly distorts the
hemispheric symmetry, such that the lateralisation index in the outer layer of IR gerbils differs from that of ER gerbils at a level of p < 0.05 and from MA-intoxicated IR gerbils at a level of p = 0.08. GABA fibres in the PirC (Fig. 2C) are not detectably altered by rearing conditions or MA intoxication in our study, nor is there any lateralisation effect. The trend that is apparent from the graphics, i.e. that after MA intoxication, ER reduces, but IR increases the fibre density, achieves probability values below 0.1.
2.2.
Anterior olfactory cortex
The acetylcholine innervation in the medial AOC is dense and homogeneous, as is the 5-HT innervation. Dopamine fibres travel rostrally in this medial area and descend towards the ventromedial surface in a patch that spans just a few slices (i.e., approx. 300–400 μm in the rostrocaudal direction). GABA neurites and cells are present here as well as everywhere else in the olfactory bulb, with both dendritic and somatic innervation. For the quantification of the cholinergic innervation (Fig. 3A), ANOVA reveals highly significant effect of rearing (p < 0.001) and MA (p < 0.01), and a significant interaction of the two (p < 0.05). In contrast analysis, IR increases the ACh innervation of control animals in the left hemisphere (p < 0.05), and of MA-intoxicated animals in both hemispheres (p < 0.001). MA intoxication leads to a reduced fibre maturation in ER animals (p < 0.001), with no effect in IR animals. The serotonin innervation of the AOC (Fig. 3B) is not changed by IR or MA-intoxication, as far as the direct groupto-group comparison is concerned. ANOVA detects an interaction effect of rearing, MA and hemisphere (p < 0.05), but
Table 1 – Means ± S.E.M. for all evaluated transmitter densities in the piriform cortex (PirC) and anterior olfactory cortex (AOC) ER
PirC LI
ACh LIa ACh LIb 5-HT GABA PirC ACh LII/III 5-HT GABA PirC ACh total 5-HT GABA AOC ACh 5-HT GABA DA medial DA TT
ER–MA
IR
IR–MA
Left
Right
Left
Right
Left
Right
Left
Right
5.55± 0.19r**m*** 4.96± 0.27r** 9.44± 0.3 4.91± 0.34 6.84± 0.18r**m* 5.28± 0.26 4.79± 0.25 6.3 ± 0.17r**m* 6.76± 0.26 4.85± 0.29 8.56± 0.39r*m***
5.07± 0.3r***m*** 5.21± 0.44 9.31± 0.34r** 4.81± 0.26 6.67± 0.37r*m* 5.24± 0.26 4.9 ± 0.2 6.14± 0.34r*m* 6.69± 0.26r**
6.92± 0.25m*** 5.82± 0.37 8.88± 0.24 4.76± 0.18 7.73± 0.33m* 4.75± 0.15 4.53± 0.26 7.21± 0.32m* 6.22± 0.16 4.65± 0.21 7.0 ± 0.32r***m***
7.0 ± 0.23m*** 5.64± 0.39 9.18± 0.17 4.7 ± 0.19 7.74± 0.43m* 4.97± 0.13 4.73± 0.22 7.17± 0.38m* 6.47± 0.1 4.72± 0.19 6.58± 0.35r***m***
6.76± 0.36r** 6.36± 0.34r** 9.9 ± 0.53 4.73± 0.2 8.34± 0.52r** 5.77± 0.36 4.49± 0.24 7.69± 0.43r** 7.25± 0.41 4.57± 0.2 9.64± 0.21r*
6.95± 0.38r*** 6.07± 0.35 10.68 ± 0.24r**m* 4.8 ± 0.25 7.98± 0.34r* 5.81± 0.28 5.15± 0.19 7.42± 0.32r* 7.54± 0.25r**m*
6.54± 0.21 6.32± 0.18 9.32± 0.35m* 5.21± 0.34 8.2 ± 0.13 5.27± 0.17 5.09± 0.15 7.59± 0.13 6.72± 0.22m*
5.53± 0.21 5.5 ± 0.14 5.67± 0.26m**
5.72± 0.23 5.57± 0.23 5.59± 0.21r***
4.8 ± 0.3 8.73± 0.43 6.17± 0.38 5.36± 0.21 5.11± 0.34r***
6.69± 0.18 6.28± 0.24 9.24± 0.22 5.39± 0.14 8.22± 0.26 5.36± 0.35 5.31± 0.25 7.52± 0.2 6.74± 0.29 5.35± 0.18 8.89± 0.37r*** 5.84± 0.24 5.32± 0.21 4.61± 0.41
5.13± 0.23m*
5.53± 0.24r*
5.38± 0.15
5.71± 0.23 6.11± 0.37 4.76± 0.26r***
4.86± 0.21 8.44± 0.4m*** 5.72± 0.2 5.18± 0.22 5.68± 0.17 5.66± 0.26 4.55± 0.24r***m** 5.01± 0.24
4.23± 0.26r*
4.07± 0.21m*
4.61± 0.14
5.6 ± 0.22m*
5.14± 0.21 9.6 ± 0.24r*** 5.57± 0.21 5.59± 0.23 4.54± 0.34 5.25± 0.13
Superscript r indicates significant differences compared between rearing conditions, superscript m significant differences by methamphetamine treatment. For abbreviations see main text. *p < 0.05; **p < 0.01; ***p < 0.001.
40
BR A IN RE S EA RCH 1 1 41 ( 20 0 7 ) 3 7 –47
Fig. 2 – Fibre densities in the anterior piriform cortex. Values are given as means ± S.E.M. for the cholinergic (A), serotonergic (B) and GABAergic (C) innervation. *p < 0.05; **p < 0.01; ***p < 0.001. Asterisks without brackets refer to the neighbouring bars below them. Abbreviations: ER: enriched rearing; ER–MA: methamphetamine treatment (PD14) followed by enriched rearing; IR: isolated rearing; IRMA: methamphetamine treatment followed by isolated rearing.
there is no significant effect of either factor alone. The same holds true for the GABA fibres (Fig. 2C). Neither rearing conditions nor MA intoxication nor laterality induce any difference in their density by themselves, although the leftsided decline from ER controls to IR–MA animals is significant at a level of p = 0.07. Dopamine fibre densities were assessed separately in the medial AOC (same area as the other transmitters, Fig. 3D) and the ventral olfactory cortex/taenia tecta (Fig. 3E). The two regions were treated as repeated measurements in ANOVA, which points out an effect of rearing both by itself (p < 0.05) and in interaction with MA intoxication and region (p < 0.05/0.01, respectively). In the medial AOC, contrast analysis reveals an increase in fibre density by IR of saline-injected animals (p < 0.001) in both hemispheres, and by MA-intoxication of ER animals in the right hemisphere (p < 0.01), with no corresponding effect at all in the left AOC. Similarly, DA innervation
densities are increased by IR of control animals in the left ventral AOC and by MA in the right AOC (both p < 0.05), whereas in MA-intoxicated IR animals, the DA fibre density is reduced in the left hemisphere. Regression analysis of GABA and DA fibre densities in the medial AOC showed that there is a negative correlation (β = −0.36) of the two parameters in the left hemisphere at a level of p < 0.06, as seen across all animals. Within single groups, no significant correlations could be detected. The correlation of group means across both hemispheres is significant (p < 0.05, R2 = 0.585).
2.3.
Lateralisation of effects in the anterior olfactory cortex
The comparison of lateralisation indices revealed some astonishing effects in the AOC (Fig. 4). The ACh innervation, which is normally symmetrical with a leftward tendency,
BR A IN RE S E A RCH 1 1 41 ( 20 0 7 ) 3 7 –4 7
switches significantly (p = 0.06) from left to right by isolated rearing of MA-intoxicated animals, as compared to IR controls (Fig. 4A). Exactly the opposite effect can be observed in the 5-
41
HT innervation, which reverses from right > left in IR controls to left > right in IR–MA animals (p < 0.05, Fig. 4B). The GABA fibres, which showed no significant alteration by direct investigation, undergo a similar rightward switch as ACh fibres, with IR–MA animals in this case being significantly different from all other groups (Fig. 4C). In contrast to these conspicuous results, the hemispheric distribution of the DA innervation is not affected by any influence in either region (Figs. 4D, E). Likewise, none of the transmitter systems undergoes alterations in laterality in the PirC. The interhemispheric coupling is also influenced by epigenetic factors (Table 2). ACh fibre densities of the left and right PirC correlate positively in layer I of IR animals, whereas in the total mean of the PirC, there is an interhemispheric correlation in ER control animals that is lost by any treatment. In the AOC, ER control and MA animals show a positive correlation that is absent in IR gerbils. Serotonin fibre densities of the PirC are positively correlated in all groups, although not necessarily significantly so in all layers. As a rule, the correlation is strong in saline-injected animals of both rearing conditions and is weakened by MA intoxication. GABA and DA innervation show little or no interhemispheric correlation.
3.
Discussion
In the present study, we assessed the densities of acetylcholine (ACh), serotonin (5-HT), dopamine (DA) and GABA fibres in the anterior olfactory cortex (AOC) and anterior piriform cortex (PirC), as they adapt to an early methamphetamine (MA) intoxication and isolated rearing (IR). Isolation rearing increases the cholinergic innervation density of salineinjected animals in both areas, but only in the AOC of MApretreated gerbils, such that MA-intoxicated ER animals attain a higher fibre density than their saline-treated counterparts in the PirC, but a lower one in the AOC. There is a denser 5-HT innervation in the right PirC after IR, which is reversed or prevented by concomitant MA intoxication. GABAergic interneurons do not seem to react significantly to either influence, but their neuritic density is negatively correlated to the DA fibre density both across animals and across groups in the AOC. The DA fibres, finally, become denser in both evaluated areas after IR and, in the right hemisphere, after MA intoxication, while the innervation density is reduced in the left ventral AOC (taenia tecta) in MA-treated IR animals.
Fig. 3 – Fibre densities in the anterior olfactory cortex (AOC). Values are given as means ± S.E.M. for the cholinergic (A), serotonergic (B) GABAergic (C) and dopaminergic (D) innervation of the medial AOC and the dopaminergic innervation of the ventral AOC/taenia tecta (E). *p < 0.05; **p < 0.01; ***p < 0.001. Asterisks without brackets refer to the neighbouring bars below them. Abbreviations: ER: enriched rearing; ER–MA: methamphetamine treatment (PD14) followed by enriched rearing; IR: isolated rearing; IRMA: methamphetamine treatment followed by isolated rearing.
42
BR A IN RE S EA RCH 1 1 41 ( 20 0 7 ) 3 7 –47
This complex set of results presents a confusing puzzle at first glance, but on closer inspection reveals an interesting pattern, characterised principally by the following traits: a) Interrelations exist in the first place between ACh and 5-HT, on the one hand, and DA and GABA on the other. b) The olfactory cortices of the two hemispheres are morphologically interdependent.
3.1. cortex
Acetylcholine–serotonin interactions in the olfactory
The close interrelation of ACh and 5-HT in the brain has been the topic of much research (see Cassel and Jeltsch, 1995; Steckler and Sahgal, 1995 for review). The two neuromodulatory systems densely innervate the whole brain and together form a necessary basis for cortical wakefulness (Dringenberg and Vanderwolf, 1997, 1998). In the present study, both innervations of the PirC respond to rearing conditions in control animals, but remain inert after MA intoxication. It seems that the single pharmacological damage shifts the whole system into a new state in which stimulations produce no effect. On the other hand, ACh and 5-HT fibres respond complementarily in several ways. In the PirC, ACh fibres adapt to epigenetic influences most significantly in the left hemisphere, whereas the 5-HT innervation is responsive only on the right. IR of MA-pretreated animals antithetically switches the lateralisation indices of ACh and 5-HT in the AOC. And the interhemispheric coupling of ACh fibres is mostly dependent on rearing conditions, that of 5-HT fibres, in turn, on MA intoxication. It seems indeed that the two neuromodulators carve out their territories, or share their responsibilities. This is in line with previous findings from our group which show that the ACh innervation will react to epigenetic influences in the hemisphere which receives the relatively weaker 5-HT innervation, and in both hemispheres only if the 5-HT fibres are symmetrical (Lehmann et al., 2004; Neddens et al., 2003, 2004).
Table 2 – Interhemispheric coupling in the AOC
ACh
5-HT
GABA
Fig. 4 – Lateralisation indices (percent deviation from interhemispheric mean) for the cholinergic (A), serotonergic (B), GABAergic (C) and dopaminergic (D) innervation of the AOC and the dopaminergic innervation of the ventral AOC/ taenia tecta (E). Values are given as means ± S.E.M. *p < 0.05; **p < 0.01; ***p < 0.001. Abbreviations: ER: enriched rearing; ER–MA: methamphetamine treatment (PD14) followed by enriched rearing; IR: isolated rearing; IR MA: methamphetamine treatment followed by isolated rearing.
DA
PirC LIa PirC LIb PirC LII/III PirC total AOC PirC LI PirC LII/III PirC total AOC PirC LI PirC LII/III PirC total AOC AOC AOC/TT
ER–Saline
ER–MA
IR–Saline
IR–MA
0.419 0.632 0.621 0.773* 0.691* 0.664* 0.588 0.793** 0.564 0.468 0.628 0.632 0.595 0.348 0.593
0.271 0.580 0.530 0.521 0.817** 0.788** 0.353 0.465 0.245 0.678 0.421 0.571 0.606 −0.450 0.393
0.874** 0.547 0.500 0.633 −0.419 0.776* 0.845** 0.907** 0.641 0.610 0.160 0.709 0.462 0.133 0.441
0.780* 0.631* 0.250 0.537 0.471 0.628 0.876** 0.773* 0.825 0.952 0.475 0.636 0.851* 0.312 0.622
Correlations of left and right fibre densities within a region across all animals in one group. For abbreviations see main text. *p < 0.05; **p < 0.01; ***p < 0.001.
BR A IN RE S E A RCH 1 1 41 ( 20 0 7 ) 3 7 –4 7
In their functional involvement, ACh and 5-HT may not necessarily be seen as complementary. Both take part in olfactory learning (Hasselmo and Barkai, 1995; McLean et al., 1993; Patil et al., 1998; Rosin et al., 1999) (reviewed in McLean et al., 1996; Mendlin et al., 1999; Wilson et al., 2004), but in very different roles: Whereas ACh opens the network for extrinsic signals, 5-HT only facilitates its effect (Nilsson et al., 1988; Riekkinen et al., 1991), which is coherent with the general observation that ACh has a vastly more potent influence on learning abilities than 5-HT (Myhrer, 2003). Correspondingly, ACh and 5-HT exert quite different physiological effects at the neuronal/synaptic level (Hasselmo, 1995). It is also interesting to note that the PirC is reciprocally connected specifically to the orbital prefrontal cortex (Datiche and Cattarelli, 1996), where both 5-HT and ACh react in similar ways (Lehmann et al., 2004; Neddens et al., 2003, 2004). The absence of 5-HT effects in the AOC may be because this structure received fibres from the median raphe nucleus which are generally spared from amphetamine neurotoxicity (Mamounas et al., 1991), whereas the 5-HT projection to the PirC is afforded by the dorsal raphe (Datiche et al., 1995).
3.2. cortex
Dopamine–GABA interactions in the anterior olfactory
Dopamine fibres target only selected brain areas, in which they exert a powerful impact on morphology and function. Their regional appearance makes them a perfect candidate for interactions with local interneurons, although, of course, synaptic, physiological and anatomical interrelations with other transmitter systems have been shown (Kostrzewa et al., 1998; Moore et al., 1999; Nishikawa et al., 2002). The relationship of DAergic and serotoninergic neurons has received special attention, since they have reciprocal connections both between their somata and on the level of synapses (Ferré et al., 1994; Mendlin et al., 1999; Moukhles et al., 1997). A mutual shift of balances in some forebrain regions has been demonstrated both in the present paradigm and in similar combinations of early traumatic experience and isolated rearing (Braun et al., 2000; Lehmann et al., 2003; Poeggel et al., 2003). According to the present results, however, no indication of such a correlation can be found in the olfactory cortex. In contrast, it is conspicuous that the GABA plexus stays completely inert to epigenetic influences in the PirC which is devoid of DA innervation, but correlates inversely with the DA fibre density in the AOC, here even showing a weakly significant reduction in response to combined IR and MA intoxication. Such an interconnection of the two transmitters has been observed before in the medial prefrontal cortex. Here, MA intoxication leads to a suppressed maturation of DA fibres (Dawirs et al., 1994). As their net effect onto the local network is inhibitory, the lack is compensated by a strong sprouting of dendritic GABAergic synapses in layers III and V (Nossoll et al., 1997). Exactly the opposite situation is found in the AOC: IR and (in the right hemisphere) MA traumatisation increase the DA fibre density, with a compensatory reduction of GABA neuropil. This is in line with recent findings in the nucleus
43
accumbens (Lesting et al., 2005; Neddens et al., 2002), which lies directly adjacent to the evaluated area and is obviously the intermediate origin of the investigated DA fibres. As has been speculated earlier (Busche et al., 2004), there seems to be a seesaw relationship between the ventral tegmental area's cortical DA projection, which is reduced by IR and MA (Dawirs et al., 1994; Neddens et al., 2001; Winterfeld et al., 1998), and its subcortical projections, which are enhanced (Busche et al., 2004; Lesting et al., 2005). That lesioning of subcortical DA projections leads to a transient (after 2 weeks) loss, but longterm recovery of fibres has been demonstrated before (Bezard et al., 2000). We here show that this regenerative sprouting is at least partly dependent on environmental conditions, as it is absent in the ventral AOC of IR–MA animals.
3.3.
Epigenetic effects on cortical laterality
An anatomical and functional lateralisation of the brain exists not only in man, but also in rodents (Denenberg, 1983; Glick et al., 1977; Neddens et al., 2004). Put somewhat simplistically, the left hemisphere is more concerned with reward, inhibition of aggression, and learning of abstract stimuli, the right hemisphere, in contrast, with low mood, aggression, and concrete stimuli (Bianki, 1982; Denenberg, 1980; Garbanati et al., 1983; Glick et al., 1980). This pattern also applies to the human PirC, which shows more activation by pleasant tastes in the left side if subjects are hungry, and more right side activation if they are satiated (Del Parigi et al., 2002). Similarly, preferential activation of the left nucleus accumbens by pleasant and of the right accumbens by unpleasant odours has been demonstrated in rats (Besson and Louilot, 1995). Such a separation of tasks requires continuous interhemispheric communication. In the rodent olfactory cortex, this is subserved by strong contralateral projections of the AOC (Shipley et al., 1995). An anatomical lateralisation of brain structures has been found on the population level in rodents both for the prefrontal DA innervation (Slopsema et al., 1982) and for 5HT fibres in frontal and caudal areas of the cortex (Neddens et al., 2004). This latter paper from our work group found 5-HT fibre densities in the right PFC of ER gerbils to be approximately twice as high as in the left cortex, and 50% higher in the left than right entorhinal cortex. In the present study, in contrast, none of the evaluated transmitters shows any lateralisation in ER control animals. Since the medial and dorsal raphe nuclei project to both lateralised and nonlateralised cortex fields in no obvious segregation, it seems that laterality is a neocortical feature, possibly being related to the separation of higher-order functions. Nevertheless, the interhemispheric correlation values obtained in our study confirm that the ACh and 5-HT projections to the PirC and, for ACh, the AOC are interhemispherically coupled, i.e., they interact in a reciprocal feedback loop (Denenberg, 1980). This coupling can be disrupted by epigenetic influences, with ACh being more sensitive to rearing conditions, 5-HT more to MA intoxication. Interestingly, sensitisation with cocaine, which like MA is an indirect DA agonist, abolishes the interhemispheric coupling of DA transmission in the PirC (Heidbreder et al., 1999). Impairments in cerebral laterality have also been found after IR (Franklin
44
BR A IN RE S EA RCH 1 1 41 ( 20 0 7 ) 3 7 –47
and Paxinos, 1997; Heidbreder et al., 2000; Neddens et al., 2004). That an early MA intoxication prevents the adaptation of DA fibres in the AOC to environmental conditions exclusively in the right hemisphere may have to do with the right-sided 5HT-response in neighbouring areas (Neddens et al., 2004). On the other hand, the mesolimbic DA fibres are themselves asymmetrically involved in odour processing (Besson and Louilot, 1995), and also react in a lateralised fashion to early MA intoxication (Lesting et al., 2005). Thus, the unilateral DA fibre responses in the AOC have their origin in the differential functional involvement of the hemispheres. The most conspicuous but, unfortunately, also most confusing finding in this context is that the lateralisation indices for the ACh, 5-HT and GABA innervations of the AOC are reversed by concomitant MA intoxication and IR. This goes along with an induction of interhemispheric coupling in the GABA and, possibly (p < 0.09), the 5-HT innervation. A putative interpretation is that in these animals a fundamental reorganisation of the hemispheric cooperation has happened, by which the two AOCs develop rather as a fused entity than as interrelated individuals. The question whether such an effect can also be detected in other median raphe projection fields is of general relevance in so far as the combination of early traumatisation with later environmental deprivation bears some resemblance to the etiology of some psychiatric disorders (Kaufman and Charney, 2001; McCarley et al., 1999; Read et al., 2001).
3.4.
Conclusions
In this study, we have examined the structural interactions of four prominent transmitter innervations in the AOC and PirC, in brains that had matured under different epigenetic influences. A preferential relationship of ACh and 5-HT fibres on the one hand and DA and GABA innervation on the other hand could be demonstrated, but the results also indicate that the distortion of the neurochemical mobile within a certain brain region can not be sufficiently explained by interactions within that region. Instead, afferent influences always play an important role. Obviously, the findings are limited temporally, spatially and in the choice of transmitters. But they provide a first tentative step towards understanding the complex recurrent interactions of transmitter systems in the brain, which may hopefully be incorporated into a more comprehensive investigation of structural plasticity in the brain.
4.
Experimental procedures
4.1.
Animals and rearing conditions
Male gerbils were bred in our facilities either in standard cages or in semi-naturally structured pens of 1 m2 (for details, see Winterfeld et al., 1998). On postnatal day (PD) 14, some of the pups from each condition were injected with a single dose of 50 mg/kg methamphetamine (MA); the others received a saline injection. Applied at this age, this intoxication preferentially lesions prefrontal DA terminals (Teuchert-Noodt and
Dawirs, 1991) and results in their reduced maturation into adulthood (Dawirs et al., 1994). At weaning (PD30), animals that had been born in standard cages were assigned to isolated rearing (IR) conditions, staying singly in standard makrolon cages (type 3), while those from enriched compounds were reared under enriched conditions (ER) in sibling groups in compounds similar to the ones they were born in. Under both sets of conditions there was a bedding of wood shavings, and food and water were provided ad libitum. All gerbils were kept on natural day/night cycles. The experiments were carried out in accordance with the European Communities Council Directive of 24 November 1986 (86/609/ EEC). Formal approval to conduct the experiments described has been obtained from the animal subjects review board of our institution and could be provided upon request. All efforts were made to minimize the number of animals used and their suffering. For the present study we used samples that had been prepared during previous projects, such that no animal was killed for this study alone. Yet, all samples for one transmitter were processed during a short time span, with groups randomly mixed. Due to the imponderabilities of immunohistochemistry, the sample sizes varied considerably even between hemispheres and neighbouring laminae. Usually, between seven and ten animals were evaluated in each group.
4.2.
DA and GABA immunohistochemistry
The method has been described in detail elsewhere (Busche et al., 2004). In short, the animals were transcardially perfused on PD 90 under deep choralhydrate anesthesia with 0.05 M cacodylate buffer (pH 6.2) containing 1% sodium metabisulfite, followed by 5% glutaraldehyde with 1% sodium metabisulfite in 0.1 M cacodylate buffer (pH 7.5), and finally by 0.05 M Trisbuffered saline (TBS) with 1% sodium metabisulfite (pH 7.5). The brains were dissected, and 40-μm-thick frontal sections
Fig. 5 – Clip out of a schematic drawing of the olfactory peduncle (Franklin and Paxinos, 1997). Evaluated areas are marked by shaded rectangles. Abbreviations: AOCd/v: anterior olfactory cortex, dorsal/ventral part; ca: commissura anterior; TTd/v: dorsal/ventral taenia tecta; PirC: piriform cortex.
BR A IN RE S E A RCH 1 1 41 ( 20 0 7 ) 3 7 –4 7
cut with a vibratome (Leica VT 1000S, Nussloch, Germany) and collected in TBS at 4 °C. Alternating sections were used for GABA and DA immunohistochemistry. They were incubated with the primary antibody (rabbit anti-dopamine, DiaSorin, Stillwater, MN, 1:600; rabbit anti-GABA, DiaSorin, 1:5000) over two nights. Following avidin–biotin reaction (Sigma, St. Louis, MO), staining was done by 0.05% 3,3-diaminobenzidine (DAB, Sigma) and 0.01% H2O2 in TBS.
4.3.
Serotonin immunohistochemistry
The method has been extensively described elsewhere (Neddens et al., 2003). In brief, the animals were transcardially perfused with 100 ml 0.1 M phosphate buffer (pH 7.2), followed by 500 ml of 4% phosphate-buffered paraformaldehyde (pH 7.4) on PD110 under deep choralhydrate anesthesia. Frontal sections of 20 μm were taken on a frigocut and every third section collected in ice-cold phosphate-buffered saline (PBS, pH 7.4). The slices were incubated for 10 min with 1% H2O2 to reduce background staining and blocked with 10% normal goat serum in PBS containing 0.3% Triton X-100. The primary antibody (rabbit anti-serotonin, Incstar) was diluted 1:20,000 in PBS with 1% NGS and 0.3% Triton X-100, and applied for 18 h at 4 °C. Following avidin–biotin reaction (Sigma, St. Louis, MO), the slices were stained by DAB in TBS.
4.4.
Acetylcholine enzyme histochemistry
The method used was described in detail elsewhere (Lehmann et al., 2004). Briefly, animals were transcardially perfused on PD90 under deep choralhydrate anesthesia (1.7 g/kg, i.p.) with 100 ml 0.1 M phosphate buffer (pH 7.2), followed by 500 ml of 4% phosphate-buffered paraformaldehyde (pH 7.4). The brains were removed and postfixed for 4 days at 4 °C, after which they were immersed in 30% sucrose at 4 °C for a further 2 days. Frontal sections of 50 μm were taken on a frigomobile and every other section collected in ice-cold 0.1 M maleate buffer (MB, pH 6.0). The staining procedure was carried out according to the method by Tago et al. (1986). The sections were incubated for 45 min at RT in the staining solution containing, in 50 ml 0.1 M MB, 320 μg K3Fe(CN)6, 940 μg CuSO4, 2,94 mg C6H5Na3O7·2H2O and 1 mg acetylcholine thioiodide as a substrate for acetylcholinesterase. They were stained in a solution containing 0.05% DAB, 0.01% H2O2 and 0,15% ammonium nickel sulfate (Fluka, Milwaukee, WI) in TBS.
4.5.
Computer-aided image analysis
Pictures were taken in one focal plane on a light microscope (for ACh, 5-HT and DA sections: Polyvar, Reichert-Jung; for GABA sections: BX61, Olympus) linked to a digital camera (ProgRes 3008 mF, Jenoptik, Jena or colorview, SOFT imaging systems, Münster, respectively). All images were assessed using a software for image analysis (KS300, Zeiss, Jenoptik, Jena). The investigators were blind to the treatment conditions of the specimens. In the anterior olfactory cortex (AOC), all pictures were taken at a magnification of 250× and a resolution of 768 × 580 square pixels at a position immediately medial to the commissura anterior (Fig. 5). For ACh and 5-HT, which
45
provide a very uniform innervation, the whole area of the image in five consecutive sections was evaluated, while regions-of-interest containing the densest and best-stained innervation were defined for DA (three sections) and GABA (five sections). For DA fibre densities, an additional area in the ventromedial olfactory cortex/taenia tecta (three sections) was investigated, where fibres descend to the surface in a narrow patch. In the piriform cortex (Fig. 5), pictures from five consecutive sections were taken either at a magnification of 250× and a resolution of 768 × 580 square pixels throughout the whole thickness of the cortex (ACh, 5-HT), or at 600× and 2080 × 1544 square pixels in both the molecular and the cellular layer (GABA). In the first case, measurement windows within differentially innervated laminae were defined: For ACh, the densely innervated layer Ia, loosely innervated layer Ib and uniformly, densely innervated layer II/III were assessed separately. For 5-HT, layer I and layer II/III were differentiated. The subsequent measurement procedure differed slightly depending on area and transmitter. In general, a reference area was defined by the exclusion of blood vessels. The image was enhanced by high pass filtering and contrast augmentation. Fibres were then recognised by the so-called “valleys function”, which comes as part of the software package. This function combines a Gaussian filtering and a subsequent so-called “Gerig-operator”, which is an algorithm to compare the grey values of neighbouring pixels irrespective of global patchiness. It implies the setting of a threshold for each picture by the investigator, which was done in constant optical check against the original image. In effect, fibres were depicted as lines of one pixel width, such that possible changes in fibre morphology would not influence the measurement. The fibre area was calculated as a percentage of the reference area.
4.6.
Statistical evaluation
The data of each layer within cortical area were averaged across all sections in each animal. Total average densities across all layers of the piriform cortex were calculated with respect to the layers' different surface areas. Additionally, lateralisation indices were calculated as 100 * (right − left) / [(right + left) / 2] (i.e., as percent deviation from mean) at each position and averaged in the same way. Statistical comparison was done by analysis of variance (ANOVA) with repeated measurements, using two main factors of rearing and treatment, and two repeated measurement factors of hemispheres and (in the PirC) layers. This was followed by post-hoc analysis (LSD test) and contrast analysis. As DA and GABA sections were obtained from the same animals, correlations between the respective fibre densities in the AOC were computed and their strength assessed by regression analysis, both across all animals and within each group. The same was done to determine the interhemispheric coupling of left and right fibre densities (cp. Denenberg, 1980). All calculations were done using the programme Statistica 6 (StatSoft, Tulsa, USA). In the figures, data are depicted as means ± standard error of means (S.E.M.). The levels of significance were set at p < 0.05 (*), p < 0.01 (**) and p < 0.001 (***). In some cases, differences which are significant at a level between 0.05 < p < 0.1 are also drawn into consideration.
46
BR A IN RE S EA RCH 1 1 41 ( 20 0 7 ) 3 7 –47
Acknowledgments The authors thank Dr. Jörg Neddens, Dr. Andrea Busche, Mr. Jörg Lesting and Mrs. Anja Bagorda for technical assistance and inspiring discussions. Thanks are also due to Mr. John O. Ball for correcting the English.
REFERENCES
Benes, F.M., 1996. The role of stress and dopamine–GABA interactions in the vulnerability for schizophrenia. J. Psychiatr. Res. 31 (2), 257–275. Besson, C., Louilot, A., 1995. Asymmetrical involvement of mesolimbic dopaminergic neurons in affective perception. Neuroscience 68, 963–968. Bezard, E., Dovero, S., Imbert, C., Boraud, T., Gross, C.E., 2000. Spontaneous long-term compensatory dopaminergic sprouting in MPTP-treated mice. Synapse 38 (3), 363–368. Bianki, V., 1982. Lateralization of concrete and abstract characteristics analysis in the animal brain. Int. J. Neurosci. 17, 233–241. Braun, K., Lange, E., Metzger, M., Poeggel, G., 2000. Maternal separation followed by early social deprivation affects the development of monoaminergic fiber systems in the medial prefrontal cortex of Octodon degus. Neuroscience 95 (1), 309–318. Busche, A., Polascheck, D., Lesting, J., Neddens, J., Teuchert-Noodt, G., 2004. Developmentally induced imbalance of dopaminergic fibre densities in limbic brain regions of gerbils (Meriones unguiculatus). J. Neural Transm. 111 (4), 451–463. Cassel, J.C., Jeltsch, H., 1995. Serotonergic modulation of cholinergic function in the central nervous system: cognitive implications. Neuroscience 69, 1–41. Datiche, F., Cattarelli, M., 1996. Reciprocal and topographic connections between the piriform and prefrontal cortices in the rat: a tracing study using the B subunit of the cholera toxin. Brain Res. Bull. 41 (6), 391–398. Datiche, F., Luppi, P.H., Cattarelli, M., 1995. Serotonergic and non-serotonergic projections from the raphe nuclei to the piriform cortex in the rat: a cholera toxin B subunit (CTb) and 5-HT immunohistochemical study. Brain Res. 671 (1), 27–37. Daw, N.D., Kakade, S., Dayan, P., 2002. Opponent interactions between serotonin and dopamine. Neural Netw. 15 (4–6), 603–616. Dawirs, R.R., Teuchert-Noodt, G., Czaniera, R., 1994. The postnatal maturation of dopamine innervation in the prefrontal cortex of gerbils (Meriones unguiculatus) is sensitive to an early single dose of methamphetamine. A quantitative immunocytochemical study. J. Hirnforsch. 35 (2), 195–204. Decker, M.W., McGaugh, J.L., 1991. The role of interactions between the cholinergic system and other neuromodulatory systems in learning and memory. Synapse 7 (2), 151–168. Del Parigi, A., Chen, K., Salbe, A.D., Gautier, J.F., Ravussin, E., Reiman, E.M., Tataranni, P.A., 2002. Tasting a liquid meal after a prolonged fast is associated with preferential activation of the left hemisphere. NeuroReport 13 (9), 1141–1145. Denenberg, V.H., 1980. General systems theory, brain organization, and early experiences. Am. J. Physiol. 238 (1), R3–R13. Denenberg, V.H., 1983. Lateralization of function in rats. Am. J. Physiol. 245, R505–R509. Dringenberg, H.C., Vanderwolf, C.H., 1997. Neocortical activation: modulation by multiple pathways acting on central cholinergic and serotonergic systems. Exp. Brain Res. 116, 160–174. Dringenberg, H.C., Vanderwolf, C.H., 1998. Involvement of direct
and indirect pathways in electrocorticographic activation. Neurosci. Biobehav. Rev. 22 (2), 243–257. Ferré, S., Cortés, R., Artigas, F., 1994. Dopaminergic regulation of the serotonergic raphe–striatal pathway: microdialysis studies in freely moving rats. J. Neurosci. 14 (8), 4839–4846. Franklin, K.B.J., Paxinos, G., 1997. The Mouse Brain in Stereotaxic Coordinates. Academic Press, San Diego. Garbanati, J.A., Sherman, G.F., Rosen, G.D., Hofmann, M., Yutzey, D.A., Denenberg, V.H., 1983. Handling in infancy, brain laterality and muricide in rats. Behav. Brain Res. 7 (3), 351–359. Glick, S.D., Zimmerberg, B., Jerussi, T.P., 1977. Adaptive significance of laterality in the rodent. Ann. N. Y. Acad. Sci. 299, 273–280. Glick, S.D., Weaver, L.M., Meibach, R.C., 1980. Lateralization of reward in rats: differences in reinforcing thresholds. Science 207 (4435), 1093–1095. Haberly, L.B., 2001. Parallel-distributed processing in olfactory cortex: new insights from morphological and physiological analysis of neuronal circuitry. Chem. Senses 26 (5), 551–576. Haberly, L.B., Bower, J.M., 1998. Olfactory cortex: model circuit for study of associative memory? Trends Neurosci. 12 (7), 258–264. Haberly, L.B., Price, J.L., 1978. Association and commissural fiber systems of the olfactory cortex of the rat: II. Systems originating in the olfactory peduncle. J. Comp. Neurol. 181, 781–808. Hall, F.S., 1998. Social deprivation of neonatal, adolescent, and adult rats has distinct neurochemical and behavioral consequences. Crit. Rev. Neurobiol. 12 (1 and 2), 129–162. Hasselmo, M.E., 1995. Neuromodulation and cortical function: modeling the physiological basis of behavior. Behav. Brain Res. 67 (1), 1–27. Hasselmo, M.E., Barkai, E., 1995. Cholinergic modulation of activity-dependent synaptic plasticity in the piriform cortex and associative memory function in a network biophysical simulation. J. Neurosci. 15 (10), 6592–6604. Hasselmo, M.E., McGaughy, J., 2004. High acetylcholine levels set circuit dynamics for attention and encoding and low acetylcholine levels set dynamics for consolidation. Prog. Brain Res. 145, 207–231. Heidbreder, C.A., Oertle, T., Feldon, J., 1999. Dopamine and serotonin imbalances in the left anterior cingulate and pyriform cortices following the repeated intermittent administration of cocaine. Neuroscience 89 (3), 701–715. Heidbreder, C.A., Weiss, I.C., Domeney, A.M., Pryce, C., Homberg, J., Hedou, G., Feldon, J., Moran, M.C., 2000. Behavioral, neurochemical and endocrinological characterization of the early social isolation syndrome. Neuroscience 100 (4), 749–768. Johnson, D.M., Illig, K.R., Behan, M., Haberly, L.B., 2000. New features of connectivity in piriform cortex visualized by intracellular injection of pyramidal cells suggests that “primary” olfactory cortex functions like “association” cortex in other sensory systems. J. Neurosci. 20 (18), 6974–6982. Kahn, R.S., Davidson, M., 1993. Serotonin, dopamine and their interactions in schizophrenia. Psychopharmacology 112 (1 Suppl), S1–S4. Kaufman, J., Charney, D., 2001. Effects of early stress on brain structure and function: implications for understanding the relationship between child maltreatment and depression. Dev. Psychopathol. 13, 451–471. Kostrzewa, R.M., Reader, T.A., Descarries, L., 1998. Serotonin neural adaptations to ontogenetic loss of dopamine neurons in rat brain. J. Neurochem. 70 (3), 889–898. Lehmann, K., Lesting, J., Polascheck, D., Teuchert-Noodt, G., 2003. Serotonin fibre densities in subcortical areas: differential effects of isolated rearing and methamphetamine. Dev. Brain Res. 147 (1–2), 123–133. Lehmann, K., Hundsdörfer, B., Hartmann, T., Teuchert-Noodt, G., 2004. The acetylcholine fiber density of the neocortex is altered
BR A IN RE S E A RCH 1 1 41 ( 20 0 7 ) 3 7 –4 7
by isolated rearing and early methamphetamine intoxication in rodents. Exp. Neurol. 189 (1), 131–140. Lesting, J., Neddens, J., Busche, A., Teuchert-Noodt, G., 2005. Hemisphere-specific effects on serotonin but not dopamine innervation in the nucleus accumbens of gerbils caused by isolated rearing and a single early methamphetamine challenge. Brain Res. 1035, 168–176. Mamounas, L.A., Mullen, C.A., O'Hearn, E., Molliver, M.E., 1991. Dual serotoninergic projections to forebrain in the rat: morphologically distinct 5-HT axon terminals exhibit differential vulnerability to neurotoxic amphetamine derivatives. J. Comp. Neurol. 314 (3), 558–586. McCarley, R.W., Wible, C.G., Frumin, M., Hirayasu, Y., Levitt, J.L., Fischer, I.A., Shenton, M.E., 1999. MRI anatomy of schizophrenia. Biol. Psychiatry 45, 1099–1119. McLean, J.H., Darby-King, A., Sullivan, R.M., King, S.R., 1993. Serotonergic influence on olfactory learning in the neonate rat. Behav. Neural Biol. 60 (2), 152–162. McLean, J.H., Darby-King, A., Hodge, E., 1996. 5-HT2 receptor involvement in conditioned olfactory learning in the neonate rat pup. Behav. Neurosci. 110 (6), 1426–1434. Mendlin, A., Martín, F.J., Jacobs, B.L., 1999. Dopaminergic input is required for increases in serotonin output produced by behavioral activation: an in vivo microdialysis study in rat forebrain. Neuroscience 93 (3), 897–905. Moore, H., Fadel, J., Sarter, M., Bruno, J.P., 1999. Role of accumbens and cortical dopamine receptors in the regulation of cortical acetylcholine release. Neuroscience 88 (3), 811–822. Moukhles, H., Bosler, O., Bolam, J.P., Vallée, A., Umbriaco, D., Geffards, M., Doucet, G., 1997. Quantitative and morphometric data indicate precise cellular interactions between serotonin terminals and postsynaptic targets in rat substantia nigra. Neuroscience 76 (4), 1159–1171. Myhrer, T., 2003. Neurotransmitter systems involved in learning and memory in the rat: a meta-analysis based on studies of four behavioral tasks. Brain Res. Rev. 41 (2–3), 268–287. Neddens, J., Brandenburg, K., Teuchert-Noodt, G., Dawirs, R.R., 2001. Differential environment alters ontogeny of dopamine innervation of the orbital prefrontal cortex in gerbils. J. Neurosci. Res. 63 (2), 209–213. Neddens, J., Lesting, J., Dawirs, R.R., Teuchert-Noodt, G., 2002. An early methamphetamine challenge suppresses the maturation of dopamine fibres in the nucleus accumbens of gerbils: on the significance of rearing conditions. J. Neural Transm. 109 (2), 141–155. Neddens, J., Bagorda, F., Busche, A., Horstmann, S., Moll, G.H., Dawirs, R.R., Teuchert-Noodt, G., 2003. Epigenetic factors differentially influence postnatal maturation of serotonin (5-HT) innervation in cerebral cortex of gerbils: interaction of rearing conditions and early methamphetamine challenge. Dev. Brain Res. 146 (1–2), 119–130. Neddens, J., Dawirs, R.R., Bagorda, F., Busche, A., Horstmann, S., Teuchert-Noodt, G., 2004. Postnatal maturation of cortical serotonin lateral asymmetry in gerbils is vulnerable to both environmental and pharmacological epigenetic challenges. Brain Res. 1021, 200–208. Nilsson, O.G., Strecker, R.E., Daszuta, A., Bjorklund, A., 1988. Combined cholinergic and serotonergic denervation of the forebrain produces severe deficits in a spatial learning task in the rat. Brain Res. 453 (1–2), 235–246.
47
Nishikawa, J., Chung, Y.-C., Li, Z., Dai, J., Meltzer, H.Y., 2002. Cholinergic modulation of basal and amphetamine-induced dopamine release in rat medial prefrontal cortex and nucleus accumbens. Brain Res. 958, 176–184. Nossoll, M., Teuchert-Noodt, G., Dawirs, R.R., 1997. A single dose of methamphetamine in neonatal gerbils affects adult prefrontal gamma-aminobutyric acid innervation. Eur. J. Pharmacol. 340 (2–3), R3–R5. Patil, M.M., Linster, C., Lubenov, E., Hasselmo, M.E., 1998. Cholinergic agonist carbachol enables associative long-term potentiation in piriform cortex slices. J. Neurophysiol. 80 (5), 2467–2474. Poeggel, G., Nowicki, L., Braun, K., 2003. Early social deprivation alters monoaminergic afferents in the orbital prefrontal cortex of Octodon degus. Neuroscience 116 (3), 617–620. Reader, T.A., Dewar, K.M., 1999. Effects of denervation and hyperinnervation on dopamine and serotonin systems in the rat neostriatum: implications for human Parkinson's disease. Neurochem. Int. 34, 1–21. Read, J., Perry, B.D., Moskowitz, A., Connolly, J., 2001. The contribution of early traumatic events to schizophrenia in some patients: a traumagenic neurodevelopmental model. Psychiatry 64 (4), 319–345. Richter-Levin, G., Segal, M., 1993. Age-related cognitive deficits in rats are associated with a combined loss of cholinergic and serotonergic functions. Ann. N. Y. Acad. Sci. 695, 254–257. Riekkinen Jr., P., Sirvio, J., Valjakka, A., Miettinen, R., Riekkinen, P., 1991. Pharmacological consequences of cholinergic plus serotonergic manipulations. Brain Res. 552 (1), 23–26. Rosin, J.F., Datiche, F., Cattarelli, M., 1999. Modulation of the piriform cortex activity by the basal forebrain: an optical recording study in the rat. Brain Res. 820 (1–2), 105–111. Shipley, M.T., McLean, J.H., Ennis, M., 1995. Olfactory system. In: Paxinos, G. (Ed.), The Rat Brain. Academic Press, San Diego, pp. 899–926. Slopsema, J.S., Van der Gugten, J., de Bruin, J.P., 1982. Regional concentrations of noradrenaline and dopamine in the frontal cortex of the rat: dopaminergic innervation of the prefrontal subareas and lateralization of prefrontal dopamine. Brain Res. 250 (1), 197–200. Steckler, T., Sahgal, A., 1995. The role of serotonergic–cholinergic interactions in the mediation of cognitive behaviour. Behav. Brain Res. 67, 165–199. Tago, H., Kimura, H., Maeda, T., 1986. Visualization of detailed acetylcholinesterase fiber and neuron staining in rat brain by a sensitive histochemical procedure. J. Histochem. Cytochem. 34, 1431–1438. Teuchert-Noodt, G., Dawirs, R.R., 1991. Age-related toxicity in prefrontal cortex and caudate–putamen complex of gerbils (Meriones unguiculatus). Neuropharmacology 30 (7), 733–743. Umino, A., Nishikawa, T., Takahashi, K., 1995. Methamphetamine-induced nuclear c-fos in rat brain regions. Neurochem. Int. 26 (1), 85–90. Wilson, D.A., Fletcher, M.L., Sullivan, R.M., 2004. Acetylcholine and olfactory perceptual learning. Learn. Mem. 11 (1), 28–34. Winterfeld, K.T., Teuchert-Noodt, G., Dawirs, R.R., 1998. Social environment alters both ontogeny of dopamine innervation of the medial prefrontal cortex and maturation of working memory in gerbils (Meriones unguiculatus). J. Neurosci. Res. 52 (2), 201–209.