Effects of context-drug learning on synaptic connectivity in the basolateral nucleus of the amygdala in rats.

Context-drug learning produces structural and functional synaptic changes in the circuitry of the basolateral nucleus of the amygdala (BLA). However, how the synaptic changes translated to the neuronal targets was not established. Thus, in the present study, immunohistochemistry with a cell-specific marker and the stereological quantification of synapses was used to determine if context-drug learning increases the number of excitatory and inhibitory/modulatory synapses contacting the gamma-aminobutyric acid (GABA) interneurons and/or the pyramidal neurons in the BLA circuitry. Amphetamine-conditioned place preference increased the number of asymmetric (excitatory) synapses contacting the spines and dendrites of pyramidal neurons and the number of multisynaptic boutons contacting pyramidal neurons and GABA interneurons. Context-drug learning increased asymmetric (excitatory) synapses onto dendrites of GABA interneurons and increased symmetric (inhibitory or modulatory) synapses onto dendrites but not perikarya of these same interneurons. The formation of context-drug associations alters the synaptic connectivity in the BLA circuitry, findings that have important implications for drug-seeking behavior.

Aberrant restoration of spines and their synapses in L-DOPA-induced dyskinesia: involvement of corticostriatal but not thalamostriatal synapses.

We examined the structural plasticity of excitatory synapses from corticostriatal and thalamostriatal pathways and their postsynaptic targets in adult Sprague-Dawley rats to understand how these striatal circuits change in l-DOPA-induced dyskinesias (LIDs). We present here detailed electron and light microscopic analyses that provide new insight into the nature of the structural and synaptic remodeling of medium spiny neurons in response to LIDs. Numerous studies have implicated enhanced glutamate signaling and persistent long-term potentiation as central to the behavioral sensitization phenomenon of LIDs. Moreover, experience-dependent alterations in behavior are thought to involve structural modifications, specifically alterations in patterns of synaptic connectivity. Thus, we hypothesized that in the striatum of rats with LIDs, one of two major glutamatergic pathways would form new or altered contacts, especially onto the spines of medium spiny neuron (MSNs). Our data provide compelling evidence for a dramatic rewiring of the striatum of dyskinetic rats and that this rewiring involves corticostriatal but not thalamostriatal contacts onto MSNs. There is a dramatic increase in corticostriatal contacts onto spines and dendrites that appear to be directly linked to dyskinetic behaviors, since they were not seen in the striatum of animals that did not develop dyskinesia. There is also an aberrant increase in spines receiving more than one excitatory contact(i.e., multisynaptic spines) in the dyskinetic animals compared with the 6-hydroxydopamine-treated and control rats. Such alterations could substantially impair the ability of striatal neurons to gate cortically driven signals and contribute to the loss of bidirectional synaptic plasticity.

Different roles of BDNF in nucleus accumbens core versus shell during the incubation of cue-induced cocaine craving and its long-term maintenance.

Brain-derived neurotrophic factor (BDNF) contributes to diverse types of plasticity, including cocaine addiction. We investigated the role of BDNF in the rat nucleus accumbens (NAc) in the incubation of cocaine craving over 3 months of withdrawal from extended access cocaine self-administration. First, we confirmed by immunoblotting that BDNF levels are elevated after this cocaine regimen on withdrawal day 45 (WD45) and showed that BDNF mRNA levels are not altered. Next, we explored the time course of elevated BDNF expression using immunohistochemistry. Elevation of BDNF in the NAc core was detected on WD45 and further increased on WD90, whereas elevation in shell was not detected until WD90. Surface expression of activated tropomyosin receptor kinase B (TrkB) was also enhanced on WD90. Next, we used viral vectors to attenuate BDNF-TrkB signaling. Virus injection into the NAc core enhanced cue-induced cocaine seeking on WD1 compared with controls, whereas no effect was observed on WD30 or WD90. Attenuating BDNF-TrkB signaling in shell did not affect cocaine seeking on WD1 or WD45 but significantly decreased cocaine seeking on WD90. These results suggest that basal levels of BDNF transmission in the NAc core exert a suppressive effect on cocaine seeking in early withdrawal (WD1), whereas the late elevation of BDNF protein in NAc shell contributes to incubation in late withdrawal (WD90). Finally, BDNF protein levels in the NAc were significantly increased after ampakine treatment, supporting the novel hypothesis that the gradual increase of BDNF levels in NAc accompanying incubation could be caused by increased AMPAR transmission during withdrawal.

MPTP mouse models of Parkinson's disease: an update.

Among the most widely used models of Parkinson's disease (PD) are those that employ toxins, especially 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Depending on the protocol used, MPTP yields large variations in nigral cell loss, striatal dopamine loss and behavioral deficits. Motor deficits do not fully replicate those seen in PD. Nonetheless, MPTP mouse models mimic many aspects of the disease and are therefore important tools for understanding PD. In this review, we will discuss the ability of MPTP mouse models to replicate the pathophysiology of PD, the mechanisms of MPTP-induced neurotoxicity, strain differences in susceptibility to MPTP, and the models' roles in testing therapeutic approaches.

Amphetamine-associated contextual learning is accompanied by structural and functional plasticity in the basolateral amygdala.

Drug seeking and the vulnerability to relapse occur when individuals are exposed to an environment with sensory cues in which drug taking has occurred. Memory formation is thought to require plasticity in synaptic circuits, and so we examined whether the memory for a drug-paired environment correlates with changes in the synaptic circuits of the basolateral amygdala (BLA), in which emotional learning is a recognized phenomenon. We used amphetamine (AMPH) as the unconditioned stimulus in the conditioned place preference (CPP) paradigm. Rats were conditioned with 1.0 mg/kg AMPH and tested, drug free, 72 h after the last conditioning session. Controls included a saline-conditioned group and a home cage AMPH injection group, whose exposure to the CPP apparatus was delayed by 4 h, long enough to clear the AMPH from the brain. We counted excitatory synapses in the BLA using the electron microscope and the physical disector design (stereology). Rats that expressed AMPH CPP had an increase in excitatory synapses compared with controls. Excitatory synaptic activity was measured using in vivo intracellular recordings from the BLA in anesthetized rats. We found that AMPH CPP, but not drug alone, increased measures of synaptic drive, including the frequency of synaptic events, and the paired-pulse ratio of synaptic inputs to BLA pyramidal neurons. The in vivo findings suggest that the increase in BLA neuronal excitatory drive reflects the change in excitatory synapse number. Thus, context-drug associations are accompanied by structural and functional plasticity in the BLA, findings that have important implications for drug-seeking behavior.

Fos after single and repeated self-administration of cocaine and saline in the rat: emphasis on the Basal forebrain and recalibration of expression.

The effects of addictive psychostimulant drugs on the brain change over repeated administrations. We evaluated a large sample of brain structures, particularly ones comprising basal forebrain macrosystems, and determined in which the immediate-early gene product, Fos, is expressed following a single and repeated self-administrations of cocaine. The caudate-putamen and accumbens, comprising the basal ganglia input structures, and the hypothalamic supraoptic and paraventricular nuclei, lateral and medial habenula, mesopontine rostromedial tegmental nucleus and anterior cingulate cortex exhibited Fos expression enhanced by acute self-administration of cocaine (SAC), but desensitized after repeated administrations. Fos expression was mainly enhanced by acutely self-administered cocaine in basal ganglia output and intrinsic structures and the intermediate nucleus of lateral septum, medial division of the central amygdaloid nucleus and zona incerta, but, in contrast, was sensitized in these structures after repeated administrations. Acute and repeated SAC left Fos expression unaffected or marginally enhanced in most extended amygdala structures, of which nearly all, however, exhibited robustly increased Fos expression after repeated saline self-administration, occasionally to levels exceeding those elicited by cocaine. Thus, self-administered cocaine mainly elicits Fos expression, which persists or increases with repeated administrations in some structures, but declines in others. In addition, Fos expression is sensitized in most extended amygdala structures merely by the act of repeated self-administering. Similar spatiotemporal patterns of cocaine- or saline-elicited Fos expression characterize functionally related clusters of structures, such as, eg, basal ganglia input structures, basal ganglia output structures, extended amygdala and structures in the brainstem to which forebrain macrosystems project.

Increased synapses in the medial prefrontal cortex are associated with repeated amphetamine administration.

Psychostimulant drug experience leads not only to long-lasting changes in behavior but also modifications in the activity and morphology of pyramidal neurons in the medial prefrontal cortex (mPFC). The objective of this study was to establish whether repeated treatment of rats with amphetamine (AMPH) is accompanied by changes in the pattern or types of synapses in the mPFC and, specifically, onto neurons that project to the lateral hypothalamus, where our earlier work has shown increased markers of neuronal activity after repeated AMPH treatment (Morshedi and Meredith [2008] Psychopharmacology (Berl) 197:179-189). Rats were treated with a behaviorally sensitizing regimen of AMPH, following which synapses in the infralimbic and prelimbic cortices of the mPFC, were analyzed with unbiased stereology (physical disector and electron microscopy). All synapses were counted and their targets were identified by standard methodological criteria. Repeated AMPH administration was associated with a significant increase in the number of asymmetric axospinous synapses, no change in axodendritic or axosomatic contacts, and no change in the total number of synapses on corticolateral hypothalamic pyramidal neurons compared to vehicle-treated rats. Therefore, behavioral sensitization as a result of repeated exposure to AMPH is accompanied by the increased formation of spine, but not dendritic, synapses onto pyramidal neurons in the mPFC.

Modeling PD pathogenesis in mice: advantages of a chronic MPTP protocol.

Formidable challenges for Parkinson's disease (PD) research are to understand the processes underlying nigrostriatal degeneration and how to protect dopamine neurons. Fundamental research relies on good animal models that demonstrate the pathological hallmarks and motor deficits of PD. Using a chronic regimen of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine and probenecid (MPTP/p) in mice, dopamine cell loss exceeds 60%, extracellular glutamate is elevated, cytoplasmic inclusions are formed and inflammation is chronic. Nevertheless, isradipine, an L-type calcium-channel blocker, attenuates the degeneration. These data support the validity of the MPTP/p model for unravelling the degenerative processes in PD and testing therapies that slow their progress.

Animal models of Parkinson's disease progression.

Parkinson's disease (PD) is a progressive neurodegenerative disorder whose etiology is not understood. This disease occurs both sporadically and through inheritance of single genes, although the familial types are rare. Over the past decade or so, experimental and clinical data suggest that PD could be a multifactorial, neurodegenerative disease that involves strong interactions between the environment and genetic predisposition. Our understanding of the pathophysiology and motor deficits of the disease relies heavily on fundamental research on animal models and the last few years have seen an explosion of toxin-, inflammation-induced and genetically manipulated models. The insight gained from the use of such models has strongly advanced our understanding of the progression and stages of the disease. The models have also aided the development of novel therapies to improve symptomatic management, and they are critical for the development of neuroprotective strategies. This review critically evaluates these in vivo models and the roles they play in mimicking the progression of PD.

The structural basis for mapping behavior onto the ventral striatum and its subdivisions.

The striatum can be divided into dorsal (caudate-putamen) and ventral parts. In the ventral division, the nucleus accumbens, which subserves adaptive and goal-directed behaviors, is further subdivided into shell and core. Accumbal neurons show different types of experience-dependent plasticity: those in the core seem to discriminate the motivational value of conditioned stimuli, features that rely on the integration of information and enhanced synaptic plasticity at the many spines on these cells, whereas shell neurons seem to be involved with the release of predetermined behavior patterns in relation to unconditioned stimuli, and the behavioral consequences of repeated administration of addictive drugs. In the core, the principal neurons are medium sized and densely spiny, but in the medial shell, these same neurons are much smaller and their dendrites, significantly less spiny, suggesting that morphological differences could mediate unique neuroadaptations associated with each region. This review is focused on evaluating the structural differences in nucleus accumbens core and shell neurons and discusses how such different morphologies could underlie distinguishable behavioral processes.

Repeated amphetamine administration induces Fos in prefrontal cortical neurons that project to the lateral hypothalamus but not the nucleus accumbens or basolateral amygdala.

RATIONALE: The development of sensitization to amphetamine (AMPH) is dependent on increases in excitatory outflow from the medial prefrontal cortex (mPFC) to subcortical centers. These projections are clearly important for the progressive enhancement of the behavioral response during drug administration that persists through withdrawal. OBJECTIVES: The objective of this study was to identify the mPFC subcortical pathway(s) activated by a sensitizing regimen of AMPH. MATERIALS AND METHODS: Using retrograde labeling techniques, Fos activation was evaluated in the predominant projection pathways of the mPFC of sensitized rats after a challenge injection of AMPH. RESULTS: There was a significant increase in Fos-immunoreactive cells in the mPFC, nucleus accumbens (NAc), basolateral amygdala (BLA), and lateral hypothalamus (LH) of rats treated repeatedly with AMPH when compared to vehicle-treated controls. The mPFC pyramidal neurons that project to the LH but not the NAc or BLA show a significant induction of Fos after repeated AMPH treatment. In addition, we found a dramatic increase in Fos-activated orexin neurons. CONCLUSIONS: The LH, a region implicated in natural and drug reward processes, may play a role in the development and persistence of sensitization to repeated AMPH through its connections with the mPFC and possibly through its orexin neurons.

Prominent activation of brainstem and pallidal afferents of the ventral tegmental area by cocaine.

Blockade of monoamine transporters by cocaine should not necessarily lead to certain observed consequences of cocaine administration, including increased firing of ventral mesencephalic dopamine (DA) neurons and accompanying impulse-stimulated release of DA in the forebrain and cortex. Accordingly, we hypothesize that the dopaminergic-activating effect of cocaine requires stimulation of the dopaminergic neurons by afferents of the ventral tegmental area (VTA). We sought to determine if afferents of the VTA are activated following cocaine administration. Rats were injected in the VTA with retrogradely transported Fluoro-Gold and, after 1 week, were allowed to self-administer cocaine or saline via jugular catheters for 2 h on 6 consecutive days. Other rats received a similar amount of investigator-administered cocaine through jugular catheters. Afterward, the rats were killed and the brains processed immunohistochemically for retrogradely transported tracer and Fos, the protein product of the neuronal activation-associated immediate early gene, c-fos. Forebrain neurons exhibiting both Fos and tracer immunoreactivity were enriched in both cocaine groups relative to the controls only in the globus pallidus and ventral pallidum, which, together, represented a minor part of total forebrain retrogradely labeled neurons. In contrast, both modes of cocaine administration strongly increased double-labeling relative to the controls in the brainstem, specifically in the caudal ventromedial mesencephalon and rostromedial pontine tegmentum. It is concluded that a previously unappreciated activation of pallidal and brainstem afferents may contribute to the modulation of dopaminergic neuronal activity following cocaine administration.

Context modulates the expression of conditioned motor sensitization, cellular activation and synaptophysin immunoreactivity.

We tested the hypothesis that amphetamine (AMPH)-induced conditioned motor sensitization is accompanied by cellular activation (measured by Fos immunoreactivity) and synaptophysin immunoreactivity in reward-related brain areas. Forty-eight rats were tested for conditioned motor sensitization using a conditioning paradigm that was performed in a three-chambered apparatus. Rats underwent two drug pairings with 1.0 mg/kg AMPH in one outer chamber and, on alternate days, were paired with saline in the other. On the fifth day, relative to the first AMPH treatment, AMPH administration increased motor activity in the AMPH-paired context but not in the saline-paired context. Relative to the first saline treatment, saline on the fifth day produced a conditioned increase in motor activity when given in the chamber previously paired with AMPH, and saline given in the saline-paired context produced a conditioned decrease in motor activity. AMPH administered in the AMPH-paired context increased the density of both Fos and synaptophysin immunoreactivity in the dentate gyrus, cornu ammonis (CA)1, CA3, basolateral amygdala and dorsolateral striatum. This pairing between context and drug increased Fos but not synaptophysin immunoreactivity in the nucleus accumbens core and shell. Saline administered in the AMPH-paired context increased the density of Fos immunoreactivity in the basolateral amygdala and nucleus accumbens core. These data indicate that the basolateral amygdala-nucleus accumbens core pathway is necessary for the context-elicited conditioned motor responses, while the hippocampus encodes the spatial context.

MPTP administration in mice changes the ratio of splice isoforms of fosB and rgs9.

Most cases of Parkinson's disease (PD) are sporadic, suggesting an environmental influence on individuals affected by this neurodegenerative disorder. Environmental stresses often lead to changes in the regulation of splicing of pre-mRNA transcripts and this may lead to the pathogenesis of the disease. A 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)/probenecid mouse model was used to examine the changes in the splicing of the fosB and rgs9 transcripts. The ratio of DeltafosB/fosB transcript was decreased in the substantia nigra and unchanged in the striatum after acute MPTP treatment. The DeltafosB/fosB transcript ratio decreased initially and then increased in the striatum of chronically MPTP-treated animals due to different degrees of reduction for the splice variants over time, whereas the ratio was unchanged in the substantia nigra. The ratio of rgs9-2/rgs9-1 transcript decreased in the substantia nigra of mice after acute MPTP treatment and increased temporarily in the striatum after chronic MPTP treatment. There was an increase in the DeltaFosB/FosB and RGS9-2/RGS9-1 protein ratios 3 weeks and 3 days post-treatment, respectively, in chronically treated mice. The data indicate that the pattern of splice isoforms of fosB and rgs9 reflects the brain's immediate and long-term responses to the physiological stress associated with Parkinsonism.

Differential laminar effects of amphetamine on prefrontal parvalbumin interneurons.

The increase in excitatory outflow from the medial prefrontal cortex is critical to the development of sensitization to amphetamine. There is evidence that psychostimulant-induced changes in dopamine-GABA interactions are key to understanding the behaviorally sensitized response. The objective of this study was to characterize the effects of different amphetamine paradigms on the Fos activation of GABAergic interneurons that contain parvalbumin in the medial prefrontal cortex. Although a sensitizing, repeated regimen of amphetamine induced Fos in all cortical layers, only layer V parvalbumin-immunolabeled cells were activated in the infralimbic and prelimbic cortices. Repeated amphetamine treatment was also associated with a loss of parvalbumin immunoreactivity in layer V, but only in the prelimbic cortex. An acute amphetamine injection to naive rats was associated with an increase in Fos, but in parvalbumin-positive neurons of the prelimbic cortex, where it was preferentially induced in layer III. These data indicate that distinct substrates mediate the response to repeated or acute amphetamine treatment. They also suggest that a sensitizing amphetamine regimen directs medial prefrontal cortex (mPFC) outflow, via changes in inhibitory neuron activation, toward subcortical centers important in reward.

'Rejuvenation' protects neurons in mouse models of Parkinson's disease.

Why dopamine-containing neurons of the brain's substantia nigra pars compacta die in Parkinson's disease has been an enduring mystery. Our studies suggest that the unusual reliance of these neurons on L-type Ca(v)1.3 Ca2+ channels to drive their maintained, rhythmic pacemaking renders them vulnerable to stressors thought to contribute to disease progression. The reliance on these channels increases with age, as juvenile dopamine-containing neurons in the substantia nigra pars compacta use pacemaking mechanisms common to neurons not affected in Parkinson's disease. These mechanisms remain latent in adulthood, and blocking Ca(v)1.3 Ca2+ channels in adult neurons induces a reversion to the juvenile form of pacemaking. Such blocking ('rejuvenation') protects these neurons in both in vitro and in vivo models of Parkinson's disease, pointing to a new strategy that could slow or stop the progression of the disease.

Amphetamine-induced place preference and conditioned motor sensitization requires activation of tyrosine kinase receptors in the hippocampus.

The environmental context in which abused drugs are taken contribute to the drug experience and is a powerful and persistent stimulus to elicit memories of that experience even in the abstinent addict. Using amphetamine (AMPH) as the unconditioned stimulus, the present study compared two popular context-dependent paradigms in rats, conditioned motor sensitization (CMS) and conditioned place preference (CPP), to ascertain whether particular brain regions were differentially involved. The neuronal substrates underlying these context-dependent behaviors are poorly understood, but regulators of the neuronal plasticity that accompany learning, such as neurotrophic factors and their cognate tyrosine kinase receptors (e.g., TrkB), are credible candidates. We found a significant elevation of TrkB-like immunoreactivity specifically in CA3/dentate gyrus (DG) subregions of the hippocampus after AMPH (0.3 mg/kg)-induced CPP, but not in the delayed-paired (control) AMPH condition. A higher AMPH dose (1.0 mg/kg) induced both CPP and CMS and elevated TrkB in the CA3/DG as well as in the nucleus accumbens shell. The development of both conditioned behaviors was blocked by intra-CA3/DG infusion of the Trk inhibitor K-252a. These findings reveal that CPP and CMS are induced by different doses of AMPH and are associated with TrkB changes in particular brain regions. Moreover, Trk receptors in the hippocampus are critical mediators of the neuronal changes necessary for inducing both forms of conditioning. Thus, although these two conditioning models are distinct, because they are commonly regulated by the hippocampal Trk system, these receptors may be a therapeutic target for attenuating the significance of contextual cues that otherwise strengthen the addictive properties of abused drugs.

The neural substrates of amphetamine conditioned place preference: implications for the formation of conditioned stimulus-reward associations.

Associations formed between conditioned stimuli and drug reward are major contributors in human drug addiction. To better understand the brain changes that accompany this process, we used immunohistochemistry for c-Fos (a neuronal activity marker), synaptophysin (a marker for synaptogenesis) and tyrosine kinase B receptor (a neurotrophic factor receptor that mediates synaptic plasticity) to investigate the neural substrates of amphetamine-induced conditioned place preference in rats. Conditioned place preference was induced by both 1.0 mg/kg and 0.3 mg/kg doses of amphetamine. Furthermore, amphetamine conditioning increased the density of c-Fos-immunoreactive cells and these cells were fully colocalized with the tyrosine kinase B receptor in the dentate gyrus, CA1 field and basolateral amygdala. Amphetamine conditioning increased the density of synaptophysin-immunoreactive varicosities in all brain regions studied, except the nucleus accumbens shell and dorsolateral striatum. The degree of conditioned place preference was highly correlated with c-Fos-immunoreactive cell density in the basolateral amygdala and with the density of synaptophysin-immunoreactive varicosities in all mesolimbic regions studied. The latter correlation was particularly impressive for the ventral pallidum and basolateral amygdala. The formation of conditioned stimulus-amphetamine reward associations is accompanied by tyrosine kinase B receptor expression in the basolateral amygdala and dentate gyrus, CA1 and CA3 fields of the hippocampus. These data therefore suggest that the formation of conditioned stimulus-reward associations requires, at least in part, activation of amygdalar-hippocampal circuits.

Behavioral models of Parkinson's disease in rodents: a new look at an old problem.

The circuitry important for voluntary movement is influenced by dopamine from the substantia nigra and regulated by the nigrostriatal system. The basal ganglia influence the pyramidal tract and other motor systems, such as the mesopontine nuclei and the rubrospinal tract. Although the neuroanatomical substrates underlying motor control are similar for humans and rodents, the behavioral repertoire mediated by those circuits is not. The principal aim of this review is to evaluate how injury to dopamine-mediated pathways in rodents gives rise to motor dysfunction that mimics human Parkinsonism. We will examine the behavioral tests in common use with rodent models of Parkinson's disease and critically evaluate the appropriateness of each test for detecting motor impairment. We will show how tests of motor performance must be guided by a thorough understanding of the clinical symptoms accompanying the disease, the circuitry mediating dopamine deficits in rodents, and familiarity with the rodent behavioral repertoire. We will explain how investigations in rodents of skilled forepaw actions, including placing, grooming, or foot faults, have clear correlates in Parkinson's disease, and are, therefore, the most sensitive ways of detecting motor impairment following dopamine loss from the basal ganglia of rodents.

The role of oxidative stress in the dysregulation of gene expression and protein metabolism in neurodegenerative disease.

There are few examples for which the genetic basis for neurodegenerative disease has been identified. For the majority of these disorders, the key to their understanding lies in knowledge of the molecular changes that contribute to altered gene expression and the translational modification of the protein products. Environmental factors play a role in the development and chronicity of neurodegenerative disorders. Environmental stimuli such as hypoxia, toxins, or heavy metals, increase production of reactive oxygen species and lower energy reserves. Chronic exposure to oxidative radicals can adversely affect gene expression and proteolysis. This review summarizes what is currently known about some of the changes in gene expression and protein metabolism that occur after oxidative stress which contribute to neurodegeneration, and reveals areas where more research is clearly needed.