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1Department of Experimental Neurophysiology, Centre for Neurogenomics and Cognitive Research, Vrije Universiteit Amsterdam, Amsterdam; and 2Department of Pharmacology and Anatomy, Rudolf Magnus Institute of Neuroscience, University Medical Center, Utrecht, The Netherlands
Submitted 30 March 2006; accepted in final form 28 June 2006
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMETHODSRESULTSDISCUSSIONGRANTSACKNOWLEDGMENTSREFERENCES |
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONGRANTSACKNOWLEDGMENTSREFERENCES |
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). Repeated administration of drugs of abuse causes a persistent increase in dopamine release in the nucleus accumbens (NAc). We recently showed that repeated in vivo amphetamine as well as morphine treatment is associated with an increase in the strength of cholinergic modulation of GABAergic transmission within the NAc shell (de Rover et al. 2004, 2005). It is unclear, however, whether the increase in endogenous cholinergic tonus is directly caused by the repeated administration of drugs of abuse or a secondary effect caused by the increase in dopamine release in the NAc.
An experimental animal model that may shed new light on this issue is the Pitx3-deficient (or aphakia) mouse mutant. Pitx3 is a homeodomain gene that is necessary for the development of substantia nigra pars compacta (SNc) dopaminergic neurons (Smidt et al. 2004a). In Pitx3-deficient mice most dopaminergic neurons in the SNc are lost during development and consequently there is no dopamine release in the caudate putamen in these animals (Hwang et al. 2003; Nunes et al. 2003; Smidt et al. 2004a; Van den Munckhof et al. 2003). Therefore Pitx3-deficient mice have been used as a genetic mouse model of Parkinsonism (e.g., Fleming et al. 2005). However, the behavioral phenotype of the Pitx3-deficient mice is rather mild and mainly motor output and not motor skills are affected (Smits et al. 2006). This suggests that adaptive changes in other brain regions may partly compensate for the loss of dopaminergic innervation of the caudate putamen in these mutant mice. In line with this a significantly higher percentage of the surviving dopaminergic neurons, mainly located in the ventral tegmental area (VTA) and innervating the NAc, was reported to be spontaneously active in Pitx3-deficient compared with wild-type mice (Smits et al. 2005). This indicates that there is more dopamine release in the NAc of Pitx3-deficient mice compared with that of wild-type mice (Gonon and Buda 1985; Kuhr et al. 1987; Suaud-Chagny et al. 1992). Moreover, the gene expression pattern in the NAc of Pitx3-deficient mice (Smits et al. 2005) was found to be similar to the pattern found in DAT –/– mice, known to display a hyperdopaminergic tonus (Dumartin et al. 2000; Fauchey et al. 2000; Giros et al. 1996; Jones et al. 1998). Finally, Pitx3-deficient mice display increased climbing behavior (Smidt et al. 2004a,b), which can also be induced by dopaminergic agonists (Costall et al. 1978). Together, these findings lead to the idea that Pitx3-deficient mice, like DAT –/– mice, have a constitutively increased dopaminergic tonus in their NAc (Smits et al. 2005), which may partly compensate for the loss of dopaminergic tonus in the caudate putamen. Understanding these adaptations may help to find better symptomatic treatments for Parkinson's Disease.
We recently reported that cholinergic neuromodulation of the GABAergic input onto medium spiny neurons, may play an important role in mediating downstream effects of alterations in the dopaminergic input from the VTA to the NAc (de Rover et al. 2002, 2004, 2005). In these previous reports, normal animals were conditioned with repeated in vivo injections with drugs of abuse, which was shown to lead to a conditional increase in the cholinergic modulation of the GABAergic transmission recorded in medium spiny neurons. Here we investigated whether the cholinergic transmission in the NAc of Pitx3-deficient mice was similarly affected.
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METHODS |
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The Pitx3-deficient (Aphakia) strain (Rieger et al. 2001; Semina et al. 2000; Varnum and Stevens 1968) and C57Bl/6-Jico mice (Charles River, Maastricht, The Netherlands) were used. All mice were bred at the Rudolf Magnus Institute for Neuroscience (Utrecht, The Netherlands) and brought to the Vrije Universiteit Amsterdam (The Netherlands) on postnatal day (PN) 7. The animals were housed in Macrolon cages under controlled conditions (lights on from 2:00 P.M. to 2:00 A.M.). Standard food (Hope Farms, Woerden, The Netherlands) and tap water were available ad libitum. On arrival in the laboratory (Vrije Universiteit Amsterdam) the animals were allowed to become accustomed to the housing facilities for 
5 days before the beginning of the experiment. Experiments were done on mice ranging from 12 to 16 days PN. Experiments were done double blind; wild-type and Pitx3-deficient mice were taken from the nest by the experimenter without any prior knowledge about the genotypes. The heads were kept on ice and used for post hoc determination of the genotypes. An expert determined the genotypes on the basis of the presence of a lens in the eyes of the animals, after analysis of the experiments.
Electrophysiology
After decapitation the brain was removed from the skull and placed for 3 min in ice-cold artificial cerebrospinal fluid (ACSF) of the following composition (in mM): NaCl, 125; KCl, 3; NaH2PO4, 1.2; CaCl2, 2.4; MgSO4, 1.3; NaHCO3, 25; and D-glucose, 10 (pH 7.2). Transversal whole brain slices of 300 µm thickness were cut on a Leica vibratome (Nussloch, Germany). The parts containing the striata (dorsal and ventral parts were not separated) were then isolated from the rest of the slice and transferred to a storage chamber, filled with carboxygenated ACSF (5% CO2-95% O2) stored on ice (4°C).
Fifteen minutes before measurements, slices were transferred to the recording chamber, where they were continuously perfused with carboxygenated ACSF at 33°C. Only neurons in the NAc and preferably in the NAc shell were recorded (Paxinos and Franklin 2001). Although the transversal slices that we used provided the best opportunity to distinguish between the NAc core and shell regions using light microscopy, because of the age of the mice and thus the size of their NAc, we could not exclude the possibility that a small minority of our recordings were made from NAc core neurons. Further, because of the size of the NAc of the mice that were used, no discrimination between different subregions within the NAc shell was made. Instead, we aimed at recording throughout the whole NAc shell. Neurons were identified by both morphological and electrophysiological criteria as published previously (Kawaguchi et al. 1995). More specifically, voltage-clamp recordings were made from GABAergic cells, which were identified by their small soma size and compact cell appearance (not elongated). Further, only a slight leakage current was tolerated at a holding potential of –70 mV, indicating that the natural membrane potential was not more depolarized than –70 mV. Current-clamp recordings were made from cholinergic interneurons that were identified by their large soma size, about twice the average (GABAergic cell) soma size, spontaneous AP firing, typical AP shape, that is, with a large (10 mV) and relatively long lasting undershoot. Voltage-clamp recordings of GABAergic neurons were always performed at a holding potential of –70 mV. Only recordings in which the uncompensated series resistance was <20 M
were analyzed. Series resistance was usually compensated by 70% and whole cell capacitances averaged at 7 ± 3 pF, indicating that only GABAergic neurons were recorded (see RESULTS for recording criteria). Pipettes for the voltage-clamp recordings (3–4 M) were filled with (in mM): CsCl, 135.5; N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES), 10; MgATP, 2; ethylene glycol-bis (beta-aminoethyl ether) N,N,N',N'-tetraacetic acid (EGTA), 11; Tris-GTP, 0.1; and CaCl2, 1 (pH 7.2 with CsOH). Cesium was used to block potassium channels to increase the signal-to-noise ratio in these recordings and a high intracellular chloride concentration was used to set the chloride reversal potential at 0 mV to obtain a linear voltage–current relationship for the chloride-gating 
-aminobutyric acid type A (GABAA) receptors.
In contrast, in current-clamp recordings, the intracellular solution contained a low concentration of chloride to establish a more natural chloride reversal potential. Gluconate was used as a replacement of chloride. Furthermore, potassium was used instead of the potassium channel blocker cesium because functioning potassium channels are essential for the occurrence of action potentials. Electrodes for the current-clamp recordings of cholinergic (ACh) neurons were filled with (in mM): K-gluconate, 121; KCl, 9; HEPES, 10; MgATP, 4; phosphocreatine, 10; and Tris-GTP, 0.3 (pH 7.2 with KOH). We used the amplitude of the action potential (AP) as a measure of the quality of the recording. Only cells with constant AP amplitudes were analyzed (max. 10% variation). Mecamylamine was applied through bath perfusion.
HEPES was from Life Technologies (Breda, The Netherlands). NaCl, NaHCO3, KCl, D-glucose, MgATP, EGTA, Tris-GTP, and K-gluconate were from Sigma (St. Louis, MO). 6,7-Dinitroquinoxaline-2,3(1H,4H)-dione (DNQX) and mecamylamine were from RBI (Natick, MA). NaH2PO4 was from Merck (Darmstadt, Germany). CaCl2 and MgSO4 were from Baker (Deventer, The Netherlands). CsCl was from ICN (Zoetermeer, The Netherlands) or Sigma.
Data acquisition and analysis of action potential firing
Data were stored on DAT, digitized at a sampling frequency of 10 kHz, and analyzed off-line by the Strathclyde software EDR and WCP (J. Dempster, University of Strathclyde, Glasgow, UK), using an amplitude-threshold criterion (i.e., ten times the SD of the baseline noise) to detect action potentials.
The first 30 s of each recording were discarded because the firing frequency is usually high right after the transition from the cell attached to the whole cell mode. Lognormal functions were fitted to histograms of the binned interval data obtained from 300 action potentials per cell (starting at 30 s in the recordings). Lognormal or double lognormal fits were made using NLREG (nonlinear regression analysis, P.H. Sherrod, Brentwood, TN), which results in average AP intervals (as tau values) with corresponding SDs depending on the peak width. The histograms were compared by constructing cumulative probability plots followed by Kolmogorov–Smirnov (K-S) testing (plots not shown, but results given in RESULTS).
Data acquisition and analysis of synaptic transmission
Data were stored on DAT, digitized at a sampling frequency of 5 kHz, and analyzed off-line by the Strathclyde software EDR and WCP (J. Dempster, University of Strathclyde), using an amplitude-threshold criterion (i.e., twice the SD of the baseline noise) to detect synaptic events. We used the stability of the rise times to determine the quality of the recording. Only cells with constant rise times throughout the recording were analyzed. Quantitative analysis was performed as previously described in detail (Brussaard et al. 1996), i.e., frequency and amplitude of inhibitory postsynaptic currents (IPSCs) were analyzed. Unimodal lognormal functions were fitted to histograms of binned amplitude data and unimodal exponential functions were fitted to histograms of binned interval data obtained from a minimum of 1,000 IPSCs per histogram. The histograms were compared by constructing cumulative probability plots followed by K-S testing (plots not shown, but results given in RESULTS and Table 1). Further the results of the fits were compared with the Wilcoxon-matched pairs test of which the results are given in RESULTS. Average effects of mecamylamine (Fig. 2) and action potentials (Fig. 3) on amplitudes and frequencies of IPSCs were calculated by normalizing the results of the fits to the result under control conditions (100%) for each cell and averaging the normalized results of the fits per experimental condition (shown as bar graphs in Figs. 2 and 3). As a measure of the variation under control conditions, the SEs under control conditions were expressed as percentages of the average control values and visualized as horizontal bars in each bar graph (Figs. 2 and 3).
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RESULTS |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONGRANTSACKNOWLEDGMENTSREFERENCES |
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First we recorded the spontaneous firing activity of cholinergic interneurons in current clamp. These neurons were identified based on the two-photon imaging of their morphology (de Rover et al. 2004) showing that their cell soma size (cell capacitance) and their spontaneous firing properties (Kawaguchi et al. 1995) are crucially different from GABAergic interneurons and/or medium spiny neurons. The ACh interneurons in slices of wild-type animals showed spontaneous, somewhat irregular firing over a wide range of frequencies of 0.7–13.5 Hz (n = 7). These seven recordings were made in slices from seven different wild-type mice and thus the range of frequencies may be regarded as representative for the firing of cholinergic neurons in the NAc of wild-type mice. The average firing frequency was found to be 9.4 ± 1.8 Hz, and two examples are shown in Fig. 1A. The ACh interneurons in slices from Pitx3-deficient mice also showed a variety of firing frequencies between 1.6 and 24.6 Hz (n = 6). Again, this wide range of frequencies may be regarded as representative of the firing of cholinergic neurons in the NAc of Pitx3-deficient mice because we recorded six different cells from six different Pitx3-deficient mice. The average firing frequency was found to be 9.0 ± 3.9 Hz and three examples are shown in Fig. 1B. The group average of firing frequency did not differ significantly between the two groups of mice (Mann–Whitney; data not shown); however, the range of firing frequencies that we measured appeared to be shifted for the mutant mice (1.6–24.6 Hz) compared with the wild-type mice (0.7–13.5 Hz). This may indicate a more variable action potential firing in the Pitx3-deficient mice.
Thus next we analyzed the firing patterns. For each recorded cell a histogram was made of 300 APs starting after 30 s of cell dialysis at the onset of the recording. Lognormal fits were made to these histograms, except in the cases where a double lognormal fit resulted in a significantly better fit. In the case of the wild-type mice this procedure resulted in seven recordings, all of which were sufficiently described by single lognormal distributions, one with a 
value of 1.36 s and six with average values of about 0.1 s (two example fits in Fig. 1C and overview in Fig. 1E). In the mutant mice, four unimodal lognormal fits and two bimodal lognormal fits were observed (example fits in Fig. 1D and overview in Fig. 1E). Moreover, the four unimodel recordings were variable, whereas the two remaining recordings had bimodal distributions. In contrast, in six of seven recordings in the wild-type mice the average firing frequency was about 0.1 Hz and all (100%) of these distributions were fit best using an unimodal fit only. Together, the firing frequency and the firing pattern data suggested that action potential firing in the mutant mice is more variable and thus intrinsically different from that observed in wild-type mice (Fig. 1, C–E).
Endogenous nicotinic modulation of GABAergic synapses
The spontaneous firing patterns observed in the putative cholinergic interneurons may result in endogenous release of ACh, possibly leading to nicotinic modulation of the GABAergic input of medium spiny neurons in the NAc shell, in line with previous observations in rats (de Rover et al. 2004). To test this hypothesis, we recorded spontaneous inhibitory postsynaptic currents (sIPSCs) from the medium spiny cells, before and after the application of mecamylamine, a nicotinic acetylcholine receptor (nAChR) antagonist (1 µM). The GABAergic medium spiny neurons, which are the output neurons of this brain area, were identified by combined electrophysiological and morphological criteria, i.e., a two-photon laser scanning microscopy reconstruction was matched with electrophysiological properties as previously described (de Rover et al. 2004). Next, GABAergic medium spiny neurons were voltage clamped at –70 mV and sIPSCs were pharmacologically isolated from excitatory postsynaptic currents (EPSCs) using DNQX (20 µM). The mecamylamine effects on sIPSC frequencies were inconsistent (Fig. 2). Overall, mecamylamine did not affect sIPSC frequency in either wild-type mice (Fig. 2E1) or mutant mice (Fig. 2F1). However, there was a considerable amount of variation: in wild-type mice within individual recordings mecamylamine caused a substantial decrease in sIPSC frequency in three cells (P < 0.01, K-S), no effect in three other cells (P > 0.05, K-S) and an increase in sIPSC frequency in two cells (P < 0.01, K-S; Table 1). The mecamylamine effect on sIPSC frequency was variable in mutant mice as well: mecamylamine caused a decrease in sIPSC frequency in two cells (P < 0.01, K-S), no effect in two other cells (P > 0.05, K-S) and an increase in sIPSC frequency in three cells (P < 0.01, K-S; Table 1). In addition, the variation in the effect of mecamylamine on the sIPSC frequency was significantly larger in wild-type mice than in Pitx3-deficient mice (F-max test: P = 0.035; note the error bar in Fig. 2E1 vs. the error bar in Fig. 2 F1).
Within individual recordings mecamylamine caused a significant decrease in sIPSC amplitude in seven of eight cells in controls (Fig. 2, A, C, and E2 and Table 1, significant at P < 0.01 in five cells and at P < 0.05 in two other cells). This would imply the presence of a significant concentration of endogenous ACh in the slices of wild-type mice. The same holds true for four of seven cells recorded in slices from Pitx3-deficient mice, where we observed a similar suppression of the sIPSC amplitude upon application of mecamylamine (Fig. 2, B, D, and F2 and Table 1, significant at P < 0.01 in four cells). In three additional recordings from mutant mice no changes were observed (Table 1).
Evoked cholinergic modulation of GABAergic synapses
Because some effects of mecamylamine were observed in slices from both wild-type and Pitx3-deficient mice—albeit in a rather inconsistent and apparently uncorrelated manner—these results were difficult to interpret. Possibly this was explained by the fact that in these synaptic recording experiments, there were also differences with respect to the variability in firing of nearby ACh interneurons (see Fig. 1). Thus we performed paired recordings of ACh interneurons and medium spiny neurons to gain a better control over the experimental conditions of the cholinergic modulation of the GABA input in this type of experiment. To this end, the ACh neurons were recorded in current clamp, whereas nearby GABAergic medium spiny neurons were recorded in voltage clamp. Initially, the ACh neurons were kept quiescent by hyperpolarizing current injection for a period of 3 min, during which so-called baseline sIPSCs were recorded (Fig. 3, A1 and B1). Next, ACh neurons were allowed to fire at their endogenous frequency (no current injection) and again sIPSCs were recorded during AP firing in the ACh neurons (Fig. 3, A2 and B2). The AP firing in ACh neurons in wild-type slices caused an increase in sIPSC frequency in five of seven recorded cell pairs (Fig. 3, A and C, overall average: 149.5 ± 27.8%, P < 0.05), which was absent in the subsequent presence of mecamylamine (1 µM; data not shown). In contrast, in none of seven cell pairs in Pitx3-deficient mice did we observe any effect (Fig. 3, B and D, overall average: 103.9 ± 5.7%, not significant).
There was no overall effect of ACh neuron firing on the sIPSC amplitudes: in wild-type mice the average sIPSC amplitude during AP firing in ACh neurons was 104.2 ± 16.2% compared with the average baseline sIPSC amplitude (not significant, K-S). In Pitx3-deficient mice the average sIPSC amplitude during AP firing in ACh neurons was 94.5 ± 11.1% compared with the average baseline sIPSC amplitude (not significant, K-S).
In conclusion, ACh released by spontaneous AP firing in ACh neurons causes an increase in sIPSC frequency without affecting the sIPSC amplitude and this effect is not present in Pitx3-deficient mice.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONGRANTSACKNOWLEDGMENTSREFERENCES |
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The first step in the circuit of cholinergic modulation of GABAergic synapses in the NAc is the neuronal activity of the cholinergic interneurons. The endogenous firing pattern of ACh interneurons was found to be different, although the average firing frequency was not significantly different. In wild-type mice we found only unimodal lognormal fits to the interval histograms, whereas in mutant mice the firing frequency appeared more variable. We previously showed that repeated in vivo amphetamine treatment in rats causes more variable firing patterns of ACh neurons in the NAc (de Rover et al. 2004). Bimodal (or burstlike) firing patterns are classically viewed as more reliable in terms of neurotransmission compared with regular firing (reviewed in Cooper 2002; Lisman 1997). This may imply that there is a constant increased release of ACh in Pitx3-deficient mice, which is regulated at the level of the ACh neurons themselves. Because irregular firing patterns in the mutant mice may alternatively have caused a more variable cholinergic tonus in the NAc of Pitx3-deficient mice compared with wild-type mice, we also determined the effects of spontaneous as well as evoked cholinergic modulation of sIPSC frequency. In doing so, we wanted to investigate whether a hyperdopaminergic input toward the NAc of Pitx3-deficient mice (Smits et al. 2005) may cause downstream changes in the microcircuitry, for instance by alterations in the expression and/or function of nAChRs in the NAc.
Endogenous nicotinic modulation of GABAergic synapses
In wild-type and in some, but not all, of the Pitx3-deficient mice, the effects of mecamylamine were qualitatively similar to those observed in control rats, i.e., mecamylamine is likely to produce a consistent reduction in the amplitude of sIPSCs in individual recordings. As argued previously the nicotinic modulation of GABAergic input onto medium spiny neurons is best explained by presynaptic activation of nAChRs on GABAergic interneurons. The endogenous activation of these nACh receptors, most likely at the level of the somata of the interneurons, would induce presynaptic firing (de Rover et al. 2002, 2004; Koos and Tepper 2002), leading to an upregulation of probability of GABA vesicles being released and therefore affect amplitude and/or frequency of the sIPSCs in the medium spiny neurons. Thus it may be concluded that nicotinic modulation of sIPSCs may occur both in wild-type and in Pitx3-deficient mice.
However, the effects of mecamylamine in mice were not as straightforward compared with those previously described for rat slices (de Rover et al. 2002, 2004), possibly arising from the difference in size of the NAc of the animals used (age difference and difference between rats and mice). The smaller NAc used in the present research may have increased the risk that some of our recordings were made in the NAc core instead of the NAc shell as described in METHODS. Further, the increase in sIPSC frequencies caused by the nicotinic receptor blocker mecamylamine that we found in some cells from both types of mice may be the result of indirect effects upstream in the microcircuit.
Evoked cholinergic modulation of GABAergic synapses
In a final attempt to more precisely pinpoint the moment of ACh release and the extent to which nicotinic modulation of sIPSCs in the medium spiny neurons occurs, we manipulated the timing of spontaneous firing of ACh neurons in the paired recording experiments. In wild-type animals, we observed that this caused a consistent increase in sIPSC frequency, without any effect on the sIPSC amplitude. In contrast, in Pitx3-deficient mice no effects whatsoever were observed. Thus we conclude that in Pitx3-deficient mice the cholinergic neuromodulation of inhibitory connectivity in the NAc is strongly affected.
This clear-cut result currently lacks the elucidation of an underlying mechanism, although there are several possible explanations. One possible explanation is that the dopaminergic tonus normally acts as a gating mechanism controlling cholinergic modulation of GABAergic inhibition in the NAc, similar to the role dopamine is considered to play in the prefrontal cortex (Braver et al. 1999; Dreher and Burnod 2002). A hyperdopaminergic tonus was reported to be present in the NAc of Pitx3-deficient mice (Gonon and Buda 1985; Kuhr et al. 1987; Smits et al. 2005; Suaud-Chagny et al. 1992). This may cause "abnormal gating" and thus apparent insensitivity of medium spiny neurons to changes in firing rate of ACh neurons. Alternatively, it is possible that in these mutant mice alterations in the excitation-secretion coupling in ACh neurons and/or desensitization of nAChRs may have occurred. Thus the Pitx3-deficient mice may have undergone developmental alterations of the NAc microcircuitry compared with wild-type mice.
Functional implications
Previously, both amphetamine and morphine pretreatment were shown to induce an increased mecamylamine effect on sIPSC amplitudes, explained by increased ACh release in the NAc (de Rover et al. 2004, 2005). In contrast, in Pitx3-deficient mice the mecamylamine effect on the sIPSC amplitudes was decreased rather than increased. Thus it seems that not only the firing pattern of cholinergic interneurons in the NAc of Pitx3-deficient mice is affected, but also more downstream parts of the microcircuit may be affected by the increased dopaminergic tonus in the NAc of these mice. It remains to be investigated why this effect of an increased dopaminergic tonus in the NAc was found in this study and not in drug-treated rats (de Rover et al. 2004, 2005). Possible explanations could be that the microcircuit is different in the control situation. The present study was done in mice, whereas the previous drug treatment studies were done in rats. Further, in the present study the developing NAc microcircuit was affected by an increased dopaminergic input, whereas in the drug-treated rats an existing NAc microcircuit was affected.
At the behavioral level the present results may have two interesting implications. First, the differently developed NAc microcircuitry may serve as a compensatory mechanism. In wild-type mice, injection of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) leads to a selective loss of SNc dopaminergic neurons (Bradbury et al. 1986), which in turn gives rise to Parkinsonian animal behavior (Fredriksson et al. 1990; Gerlach et al. 1991). Although Pitx3-deficient mice lack dopamine in their dorsal striata, they do not display a severe Parkinson-like phenotype (Smidt et al. 2004a; Smits et al. 2006). The developmental alterations of the NAc microcircuitry in Pitx3-deficient mice may be underlying this compensated behavior. Therefore understanding these adaptations may help to find better symptomatic treatments for Parkinson's disease.
The second implication at the behavioral level is that the changed NAc microcircuitry, as measured here, may be a general effect, downstream of an increased dopaminergic transmission in the NAc, and therefore a so-called common denominator. We previously showed that the average firing frequency of cholinergic interneurons is not different between amphetamine pretreated rats and saline pretreated controls, but there is a more pronounced bimodality in the firing patterns of cholinergic interneurons of amphetamine pretreated rats (de Rover et al. 2004). The present results show a similar change in the firing of the cholinergic interneurons in Pitx3-deficient mice. Therefore we currently consider the possibility that the more pronounced bimodality in the firing pattern of cholinergic interneurons after amphetamine pretreatment is not a substance-specific but rather a secondary effect caused by the increased dopaminergic tonus in the NAc.
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ACKNOWLEDGMENTS |
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Present address of M. de Rover: The University of Cambridge Behavioural and Clinical Neurosciences Institute, Downing St., Cambridge, CB2 3EB, United Kingdom (E-mail: md415@cam.ac.uk).
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FOOTNOTES |
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Address for reprint requests and other correspondence: A. B. Brussaard, Dept. of Experimental Neurophysiology, Centre for Neurogenomics and Cognitive Research (CNCR), Vrije Universiteit Amsterdam, De Boelelaan 1085, 1081 HV Amsterdam, The Netherlands (E-mail: brssrd{at}cncr.vu.nl)
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