Dopamine Transporter Knockdown Mice Research Articles

Chronic antipsychotic treatment exerts limited effects on the mania-like behavior of dopamine transporter knockdown mice

BackgroundBipolar disorder is a life-threatening disorder linked to dopamine transporter (DAT) polymorphisms, with reduced DAT levels seen in positron emission tomography and postmortem brains. AimsThe purpose of this study was to examine the effects of approved antipsychotics on DAT dysfunction-mediated mania behavior in mice. MethodsDAT knockdown mice received either D2-family receptor antagonist risperidone or asenapine and mania-related behaviors were assessed in the clinically-relevant behavioral pattern monitor to assess spontaneous exploration. ResultsChronic risperidone did not reverse mania-like behavior in DAT knockdown mice. Chronic asenapine reduced mania behavior but this effect was more pronounced in wild-type littermates than in DAT knockdown mice. ConclusionTaken together, these findings suggest that while acute antipsychotic treatment may be beneficial in management of bipolar mania, more targeted therapeutics may be necessary for long-term treatment. Specific investigation into DAT-targeting drugs could improve future treatment of bipolar mania.

Chronic nicotine, but not suramin or resveratrol, partially remediates the mania-like profile of dopamine transporter knockdown mice

Bipolar disorder (BD) is a severe mental illness affecting 2% of the global population. Current pharmacotherapies provide incomplete symptom remediation, highlighting the need for novel therapeutics. BD is characterized by fluctuations between mania and depression, likely driven by shifts between hyperdopaminergia and hypercholinergia, respectively. Hyperdopaminergia may result from insufficient activity of the dopamine transporter (DAT), the primary mediator of synaptic dopamine clearance. The DAT knockdown (DAT KD) mouse recreates this mechanism and exhibits a highly reproducible hyperexploratory profile in the cross-species translatable Behavioral Pattern Monitor (BPM) that is: (a) consistent with that observed in BD mania patients; and (b) partially normalized by chronic lithium and valproate treatment. The DAT KD/BPM model of mania therefore exhibits high levels of face-, construct-, and predictive-validity for the pre-clinical assessment of putative anti-mania drugs. Three different drug regimens – chronic nicotine (nicotinic acetylcholine receptor (nAChR) agonist; 40 mg/kg/d, 26 d), subchronic suramin (anti-purinergic; 20 mg/kg, 1 × /wk, 4 wks), and subchronic resveratrol (striatal DAT upregulator; 20 mg/kg/d, 4 d) – were administered to separate cohorts of male and female DAT KD- and wildtype (WT) littermate mice, and exploration was assessed in the BPM. Throughout, DAT KD mice exhibited robust hyperexploratory profiles relative to WTs. Nicotine partially normalized this behavior. Resveratrol modestly upregulated DAT expression but did not normalize DAT KD behavior. These results support the mania-like profile of DAT KD mice, which may be partially remediated by nAChR agonists via restoration of disrupted catecholaminergic/cholinergic equilibrium. Delineating the precise mechanism of action of nicotine could identify more selective therapeutic targets.

Lack of dopamine D4 receptor participation in mouse hyperdopaminergic locomotor response

Chronic methamphetamine (METH) treatment induces behavioral sensitization in rodents. During this process, hyperactivation of the mesolimbic dopamine system plays a central role, and dopamine D2-like receptor-based antipsychotics are known to alleviate the behavioral hyperactivity. The atypical antipsychotic, clozapine (Clz), acts partially as a dopamine D4 receptor (D4R) antagonist and mitigates hyperdopaminergic drug addiction and/or comorbid psychotic symptoms; however, it remains unclear whether D4R blockade contributes to the therapeutic effects of Clz. Here, we evaluated the potential role of D4R in regulating hyperdopaminergia-induced behavioral hyperactivity in METH behavioral sensitization and dopamine transporter (DAT) knockdown (KD) mice. Clz or a D4R-selective antagonist, L-745,870, were co-administered to mice with daily METH in a METH sensitization model, and Clz or L-745,870 were administered alone in a DAT KD hyperactivity model. Locomotor activity and accumbal D4R expression were analyzed. Clz suppressed both the initiation and expression of METH behavioral sensitization, as well as DAT KD hyperactivity. However, repetitive Clz treatment induced tolerance to the suppression effect on METH sensitization initiation. In contrast, D4R inhibition by L-745,870 had no effect on METH sensitization or DAT KD hyperactivity. Accumbal D4R expression was similar between METH-sensitized mice with and without Clz co-treatment. In sum, our results suggest the mesolimbic D4R does not participate in behavioral sensitization encoded by hyperdopaminergia, a finding which likely extends to the therapeutic effects of Clz. Therefore, molecular targets other than D4R should be prioritized in the development of future therapeutics for treatment of hyperdopaminergia-dependent neuropsychiatric disorders.

Vulnerable and Resilient Phenotypes in a Mouse Model of Anorexia Nervosa

BackgroundIncreased physical activity is a common feature of anorexia nervosa (AN). Although high activity levels are associated with greater risk of developing AN, particularly when combined with dieting, most individuals who diet and exercise maintain a healthy body weight. It is unclear why some individuals develop AN while most do not. A rodent model of resilience and vulnerability to AN would be valuable to research. Dopamine, which is believed to play a crucial role in AN, regulates both reward and activity and may modulate vulnerability. MethodsAdolescent and young adult female C57BL/6N mice were tested in the activity-based anorexia (ABA) model, with an extended period of food restriction in adult mice. ABA was also tested in dopamine transporter knockdown mice and wild-type littermates. Mice that adapted to conditions and maintained a stable body weight were characterized as resilient. ResultsIn adults, vulnerable and resilient phenotypes emerged in both the ABA and food-restricted mice without wheels. Vulnerable mice exhibited a pronounced increase in running throughout the light cycle, which dramatically peaked prior to requiring removal from the experiment. Resilient mice exhibited an adaptive decrease in total running, appropriate food anticipatory activity, and increased consumption, thereby achieving stable body weight. Hyperdopaminergia accelerated progression of the vulnerable phenotype. ConclusionsOur demonstration of distinct resilient and vulnerable phenotypes in mouse ABA significantly advances the utility of the model for identifying genes and neural substrates mediating AN risk and resilience. Modulation of dopamine may play a central role in the underlying circuit.

Dopamine D3 receptor and GSK3\u03b2 signaling mediate deficits in novel object recognition memory within dopamine transporter knockdown mice

BackgroundOver-stimulation of dopamine signaling is thought to underlie the pathophysiology of a list of mental disorders, such as psychosis, mania and attention-deficit/hyperactivity disorder. These disorders are frequently associated with cognitive deficits in attention or learning and memory, suggesting that persistent activation of dopamine signaling may change neural plasticity to induce cognitive or emotional malfunction.MethodsDopamine transporter knockdown (DAT-KD) mice were used to mimic a hyper-dopamine state. Novel object recognition (NOR) task was performed to assess the recognition memory. To test the role of dopamine D3 receptor (D3R) on NOR, DAT-KD mice were treated with either a D3R antagonist, FAUC365 or by deletion of D3R. Total or phospho-GSK3 and –ERK1/2 signals in various brain regions were measured by Western blot analyses. To examine the impact of GSK3 signal on NOR, wild-type mice were systemically treated with GSK3 inhibitor SB216763 or, micro-injected with lentiviral shRNA of GSK3β or GSK3α in the medial prefrontal cortex (mPFC).ResultsWe confirmed our previous findings that DAT-KD mice displayed a deficit in NOR memory, which could be prevented by deletion of D3R or exposure to FAUC365. In WT mice, p-GSK3α and p-GSK3β were significantly decreased in the mPFC after exposure to novel objects; however, the DAT-KD mice exhibited no such change in mPFC p-GSK3α/β levels. DAT-KD mice treated with FAUC365 or with D3R deletion exhibited restored novelty-induced GSK3 dephosphorylation in the mPFC. Moreover, inhibition of GSK3 in WT mice diminished NOR performance and impaired recognition memory. Lentiviral shRNA knockdown of GSK3β, but not GSK3α, in the mPFC of WT mice also impaired NOR.ConclusionThese findings suggest that D3R acts via GSK3β signaling in the mPFC to play a functional role in NOR memory. In addition, treatment with D3R antagonists may be a reasonable approach for ameliorating cognitive impairments or episodic memory deficits in bipolar disorder patients.

Convergent neural substrates of inattention in bipolar disorder patients and dopamine transporter-deficient mice using the 5-choice CPT.

Bipolar disorder (BD) is a debilitating psychiatric illness affecting 2%-5% of the population. Although mania is the cardinal feature of BD, inattention and related cognitive dysfunction are observed across all stages. Since cognitive dysfunction confers poor functional outcome in patients, understanding the relevant neural mechanisms remains key to developing novel-targeted therapeutics. The 5-choice continuous performance test (5C-CPT) is a mouse and fMRI-compatible human attentional task, requiring responding to target stimuli while inhibiting responding to nontarget stimuli, as in clinical CPTs. This task was used to delineate systems-level neural deficits in BD contributing to inattentive performance in human subjects with BD as well as mouse models with either parietal cortex (PC) lesions or reduced dopamine transporter (DAT) expression. Mania BD participants exhibited severe 5C-CPT impairment. Euthymic BD patients exhibited modestly impaired 5C-CPT. High impulsivity BD subjects exhibited reduced PC activation during target and nontarget responding compared with healthy participants. In mice, bilateral PC lesions impaired both target and nontarget responding. In the DAT knockdown mouse model of BD mania, knockdown mice exhibited severely impaired 5C-CPT performance versus wildtype littermates. These data support the role of the PC in inattention in BD-specifically regarding identifying the appropriate response to target vs nontarget stimuli. Moreover, the findings indicate that severely reduced DAT function/hyperdopaminergia recreates the attentional deficits observed in BD mania patients. Determining the contribution of DAT in the PC to attention may provide a future target for treatment development.

Dopamine D3 receptor blockade rescues hyper-dopamine activity-induced deficit in novel object recognition memory

Patients afflicted with bipolar disorder demonstrate significant impairments in recognition and episodic memory during acute depressive and manic episodes. These impairments and the related pathophysiology may result from over-activation of the brain dopamine (DA) system. In order to model overactive DA transmission in a well-established novel object recognition (NOR) memory test, we used DA transporter knockdown (DAT-KD) mice, which exhibit reduced DAT expression and display hyper-dopaminergic phenotypes. DAT-KD mice exhibited impaired NOR memory compared to wild-type (WT) mice. This impairment was prevented by administration of FAUC365, a DA D3 receptor (D3R) selective antagonist, prior to object learning. Similarly, D3R knockout (KO)/DAT-KD double mutant mice displayed performance in the NOR test that was comparable to WT mice, suggesting that deficiencies in NOR performance in DAT-KD mice can be compensated by diminishing D3R signaling. GBR12909, a DAT blocker, also impaired NOR performance in WT mice, but not in D3R KO mice. Impaired NOR performance in GBR12909-treated WT mice was also prevented by pretreatment with FAUC365. Together, these findings indicate that reduced DAT activity can impair recognition memory in the NOR test, and D3R appears to be necessary to mediate this effect.

Dopamine transporter knockdown mice in the behavioral pattern monitor: A robust, reproducible model for mania-relevant behaviors

Efforts to replicate results from both basic and clinical models have highlighted problems with reproducibility in science. In psychiatry, reproducibility issues are compounded because the complex behavioral syndromes make many disorders challenging to model. We develop translatable tasks that quantitatively measure psychiatry-relevant behaviors across species. The behavioral pattern monitor (BPM) was designed to analyze exploratory behaviors, which are altered in patients with bipolar disorder (BD), especially during mania episodes. We have repeatedly assessed the behavioral effects of reduced dopamine transporter (DAT) expression in the BPM using a DAT knockdown (KD) mouse line (~10% normal expression). DAT KD mice exhibit a profile in the BPM consistent with acutely manic BD patients in the human version of the task—hyperactivity, increased exploratory behavior, and reduced spatial d (Perry et al., 2009). We collected data from multiple DAT KD BPM experiments in our laboratory to assess the reproducibility of behavioral outcomes across experiments. The four outcomes analyzed were: 1) transitions (amount of locomotor activity); 2) rearings (exploratory activity); 3) holepokes (exploratory activity); and 4) spatial d (geometrical pattern of locomotor activity). By comparing DAT KD mice to wildtype (WT) littermates in every experiment, we calculated effect sizes for each of the four outcomes and then calculated a mean effect size using a random effects model. DAT KD mice exhibited robust, reproducible changes in each of the four outcomes, including increased transitions, rearings, and holepokes, and reduced spatial d, vs. WT littermates. Our results demonstrate that the DAT KD mouse line in the BPM is a consistent, reproducible model of mania-relevant behaviors. More work must be done to assess reproducibility of behavioral outcomes across experiments in order to advance the field of psychiatry and develop more effective therapeutics for patients.

Blockade of dopamine D1-family receptors attenuates the mania-like hyperactive, risk-preferring, and high motivation behavioral profile of mice with low dopamine transporter levels.

Patients with bipolar disorder mania exhibit poor cognition, impulsivity, risk-taking, and goal-directed activity that negatively impact their quality of life. To date, existing treatments for bipolar disorder do not adequately remediate cognitive dysfunction. Reducing dopamine transporter expression recreates many bipolar disorder mania-relevant behaviors (i.e. hyperactivity and risk-taking). The current study investigated whether dopamine D1-family receptor blockade would attenuate the risk-taking, hypermotivation, and hyperactivity of dopamine transporter knockdown mice. Dopamine transporter knockdown and wild-type littermate mice were tested in mouse versions of the Iowa Gambling Task (risk-taking), Progressive Ratio Breakpoint Test (effortful motivation), and Behavioral Pattern Monitor (activity). Prior to testing, the mice were treated with the dopamine D1-family receptor antagonist SCH 23390 hydrochloride (0.03, 0.1, or 0.3 mg/kg), or vehicle. Dopamine transporter knockdown mice exhibited hyperactivity and hyperexploration, hypermotivation, and risk-taking preference compared with wild-type littermates. SCH 23390 hydrochloride treatment decreased premature responding in dopamine transporter knockdown mice and attenuated their hypermotivation. SCH 23390 hydrochloride flattened the safe/risk preference, while reducing activity and exploratory levels of both genotypes similarly. Dopamine transporter knockdown mice exhibited mania-relevant behavior compared to wild-type mice. Systemic dopamine D1-family receptor antagonism attenuated these behaviors in dopamine transporter knockdown, but not all effects were specific to only the knockdown mice. The normalization of behavior via blockade of dopamine D1-family receptors supports the hypothesis that D1 and/or D5 receptors could contribute to the mania-relevant behaviors of dopamine transporter knockdown mice.

Brexpiprazole reduces hyperactivity, impulsivity, and risk-preference behavior in mice with dopamine transporter knockdown—a model of mania

Bipolar disorder (BD) is a unique mood disorder defined by periods of depression and mania. The defining diagnosis of BD is the presence of mania/hypomania, with symptoms including hyperactivity and risk-taking. Since current treatments do not ameliorate cognitive deficits such as risky decision-making, and impulsivity that can negatively affect a patient's quality of life, better treatments are needed. Here, we tested whether acute treatment with brexpiprazole, a serotonin-dopamine activity modulator with partial agonist activity at D2/3 and 5-HT1A receptors, would attenuate the BD mania-relevant behaviors of the dopamine transporter (DAT) knockdown mouse model of mania. The effects of brexpiprazole on DAT knockdown and wild-type littermate mice were examined in the behavioral pattern monitor (BPM) and Iowa gambling task (IGT) to quantify activity/exploration and impulsivity/risk-taking behavior respectively. DAT knockdown mice exhibited hyper-exploratory behavior in the BPM and made fewer safe choices in the IGT. Brexpiprazole attenuated the mania-like hyper-exploratory phenotype and increased safe choices in risk-preferring DAT knockdown mice. Brexpiprazole also reduced safe choices in safe-preferring mice irrespective of genotype. Finally, brexpiprazole reduced premature (impulsive-like) responses in both groups of mice. Consistent with earlier reports, DAT knockdown mice exhibited hyper-exploratory, risk-preferring, and impulsive-like profiles consistent with patients with BD mania in these tasks. These behaviors were attenuated after brexpiprazole treatment. These data therefore indicate that brexpiprazole could be a novel treatment for BD mania and/or risk-taking/impulsivity disorders, since it remediates some relevant behavioral abnormalities in this mouse model.

Effects of decreased dopamine transporter levels on nigrostriatal neurons and paraquat/maneb toxicity in mice

How genetic variations in the dopamine transporter (DAT) combined with exposure to environmental toxins modulate the risk of Parkinson's disease remains unclear. Using unbiased stereology in DAT knock-down mice (DAT-KD) and wild-type (WT) littermates, we found that decreased DAT caused a loss of tyrosine hydroxylase–positive (dopaminergic) neurons in subregions of the substantia nigra pars compacta at 3–4 days, 5 weeks, and 18 months of age. Both genotypes lost dopaminergic neurons with age and remaining neurons at 11 months were resilient to paraquat/maneb. In 5-week-old mice, the toxins decreased substantia nigra pars compacta dopaminergic neurons in both genotypes but less in DAT-KD. Regional analysis revealed striking differences in the subsets of neurons affected by low DAT, paraquat/maneb, and aging. In particular, we show that a potentially protective effect of low DAT against toxin exposure is not sufficient to reduce death of all nigrostriatal dopaminergic neurons. Thus, different regional vulnerability of nigrostriatal dopaminergic neurons may contribute to an increased risk of developing Parkinson's disease when multiple factors are combined.

The effects of reduced dopamine transporter function and chronic lithium on motivation, probabilistic learning, and neurochemistry in mice: Modeling bipolar mania

BackgroundBipolar disorder (BD) mania patients exhibit poor cognition and reward-seeking/hypermotivation, negatively impacting a patient's quality of life. Current treatments (e.g., lithium), do not treat such deficits. Treatment development has been limited due to a poor understanding of the neural mechanisms underlying these behaviors. Here, we investigated putative mechanisms underlying cognition and reward-seeking/motivational changes relevant to BD mania patients using two validated mouse models and neurochemical analyses. MethodsThe effects of reducing dopamine transporter (DAT) functioning via genetic (knockdown vs. wild-type littermates), or pharmacological (GBR12909- vs. vehicle-treated C57BL/6J mice) means were assessed in the probabilistic reversal learning task (PRLT), and progressive ratio breakpoint (PRB) test, during either water or chronic lithium treatment. These tasks quantify reward learning and effortful motivation, respectively. Neurochemistry was performed on brain samples of DAT mutants ± chronic lithium using high performance liquid chromatography. ResultsReduced DAT functioning increased reversals in the PRLT, an effect partially attenuated by chronic lithium. Chronic lithium alone slowed PRLT acquisition. Reduced DAT functioning increased motivation (PRB), an effect attenuated by lithium in GBR12909-treated mice. Neurochemical analyses revealed that DAT knockdown mice exhibited elevated homovanillic acid levels, but that lithium had no effect on these elevated levels. ConclusionsReducing DAT functioning recreates many aspects of BD mania including hypermotivation and improved reversal learning (switching), as well as elevated homovanillic acid levels. Chronic lithium only exerted main effects, impairing learning and elevating norepinephrine and serotonin levels of mice, not specifically treating the underlying mechanisms identified in these models.

The Dopamine Transporter Expression Level Differentially Affects Responses to Cocaine and Amphetamine

Although both cocaine and amphetamine mainly target the dopamine transporter (DAT) and cause psychomotor effects, they have very different mechanisms of actions. The authors examined whether responses to cocaine and amphetamine were affected differentially by changes in DAT expression levels using transgenic mice with different DAT expression levels. In the constitutive DAT knockdown mice, reduced DAT expression enhanced cocaine's locomotor stimulatory effects and at the same time diminished amphetamine's locomotor stimulatory effects. Similar effects were observed in the inducible DAT knockdown mice, ruling out the contribution of developmental compensations in DAT knockdown mice. Extracellular dopamine levels in response to psychostimulants were assessed by in vivo microdialysis. Whereas amphetamine-induced increase in extracellular dopamine was drastically diminished in constitutive DAT knockdown mice, cocaine-induced increase in extracellular dopamine had a faster onset in knockdown mice compared with wild-type controls. Postsynaptically, D1 agonist–stimulated c-fos expression was significantly attenuated in constitutive DAT knockdown mice compared with wild-type controls. The authors propose that responses to cocaine and amphetamine depend on psychostimulant drug type, drug dose, as well as DAT expression level. DAT expression level affects presynaptic responses to psychostimulants directly and postsynaptic responses to psychostimulants indirectly via changes in receptor signaling. These data imply that individual differences in DAT expression (either genetically or pharmacologically induced) may affect susceptibility to addiction of different types of psychostimulants.

Chronic valproate attenuates some, but not all, facets of mania-like behaviour in mice

Bipolar disorder (BD) mania is a psychiatric disorder with multifaceted symptoms. Development of targeted treatments for BD mania may benefit from animal models that mimic multiple symptoms, as opposed to hyperactivity alone. Using the reverse-translated multivariate exploratory paradigm, the behavioural pattern monitor (BPM), we reported that patients with BD mania exhibit hyperactivity as well as increased specific exploration and more linear movements through space. This abnormal profile is also observed in mice with reduced function of the dopamine transporter (DAT) through either constitutive genetic [knockdown (KD)] or acute pharmacological (GBR12909) means. Here, we assessed the pharmacological predictive validity of these models by administering the BD-treatment valproic acid (VPA) for 28 d. After 1.5% VPA- or regular-chow treatment for 28 d, C57BL/6J mice received GBR12909 (9mg/kg) or saline and were tested in the BPM. Similarly, DAT KD and wild type (WT) littermates were treated with VPA-chow and tested in the BPM. GBR12909-treated and DAT KD mice on regular chow were hyperactive, exhibited increased specific exploration and moved in straighter patterns compared to saline-treated and WT mice respectively. Chronic 1.5% VPA-chow treatment resulted in therapeutic concentrations of VPA and ameliorated hyperactivity in both models, while specific exploration and behavioural organization remained unaffected. Hence, the mania-like profile of mice with reduced functional DAT was partially attenuated by chronic VPA treatment, consistent with the incomplete symptomatic effect of VPA treatment in BD patients. Both DAT models may help to identify therapeutics that impact the full spectrum of BD mania.

Dopaminergic enhancement of local food‐seeking is under global homeostatic control

Recent work has implicated dopaminergic mechanisms in overeating and obesity with some researchers suggesting parallels between the dopamine dysregulation associated with addiction and an analogous dysregulation in obesity. The precise role of dopamine in mediating reward and reinforcement, however, remains controversial. In contrast to drugs of abuse, pursuit of a natural reward, such as food, is regulated by homeostatic processes that putatively maintain a stable energy balance keeping unrestrained consumption and reward pursuit in check. Understanding how the reward system is constrained by or escapes homeostatic regulation is a critical question. The widespread use of food restriction to motivate animal subjects in behavior paradigms precludes investigation of this relationship as the homeostatic system is locked into deficit mode. In the present study, we examined the role of dopamine in modulating adaptive feeding behavior in semi-naturalistic homecage paradigms where mice earn all of their food from lever pressing. We compared consumption and meal patterning between hyperdopaminergic dopamine transporter knock-down and wild-type mice in two paradigms that introduce escalating costs for procuring food. We found that hyperdopaminergic mice exhibited similar demand elasticity, weight loss and energy balance in response to cost. However, the dopamine transporter knock-down mice showed clear differences in meal patterning. Consistent with expectations of enhanced motivation, elevated dopamine increased the meal size and reduced intrameal cost sensitivity. Nonetheless, this did not alter the overall energy balance. We conclude that elevated dopamine enhances the incentive or willingness to work locally within meals without shifting the energy balance, enhancing global food-seeking or generating an energy surplus.

Increased risk-taking behavior in dopamine transporter knockdown mice: further support for a mouse model of mania

Reduced functioning of the dopamine transporter (DAT) has been linked to bipolar disorder (BD). Mice with reduced DAT functioning (knockdown, KD) exhibit a behavioral profile in the mouse Behavioral Pattern Monitor (BPM) consistent with patients with BD mania in the human BPM. Patients with BD also exhibit increased risk taking, which can be quantified using the Iowa Gambling Task (IGT). We hypothesized that DAT KD mice would exhibit increased risk-taking behavior in a novel mouse version of the IGT. DAT KD and wildtype (WT) littermates were trained in the mouse IGT. In session 1, KD mice initially made riskier choices, but later performed comparably to WT mice. Once trained to stable choice performance, DAT KD mice continued to exhibit a trend to choose the riskier options more than WT mice. Finally, we confirmed that these DAT KD mice also exhibited an exploratory profile in the BPM consistent with patients with BD mania, where risky choice behavior modestly correlated with specific exploration. These data demonstrate that DAT KD mice chose the riskier options more than WT mice, providing further support for the use of DAT KD mice as a model of BD mania.

Behavioural phenotyping of dopamine transporter knockdown mice using local small interference RNA

The objective of this study was to evaluate basic behavioural phenotypes and methamphetamine-induced hyperactivity in dopamine transporter (DAT) knockdown and control mice after local injections of small interference RNA into ventral tegmental area/substantia nigra once every other day for 14 days. Local knockdown of DAT expression in the mouse brain did not affect basic behavioural functions, but these mice displayed elevated depression-like immobility and a ∼35% reduction of methamphetamine-induced hyperactivity.

Motivational effects on interval timing in dopamine transporter (DAT) knockdown mice

We examined interval timing in mice that underexpress the dopamine transporter (DAT) and have chronically higher levels of extracellular dopamine (Zhuang et al., 2001). The dopaminergic system has been proposed as a neural substrate for an internal clock, with transient elevations of dopaminergic activity producing underestimation of temporal intervals. A group of DAT knockdown (KD) and littermate wild type (WT) mice were tested with a dual peak procedure. Mice obtained reinforcement by pressing one of two levers after a fixed amount of time (30 or 45 s) had elapsed since lever extension. Only one lever was available at a time, and each lever was associated with a single duration. On occasional probe trials, the DAT KD mice began responding earlier in the interval than WT mice, but showed maximal responding and terminated responding around the same time as the WT mice. Administration of raclopride (0.2, 0.6, and 1.2 mg/kg), a D2 antagonist, eliminated most of the differences between DAT KD and WT mice, suggesting that the effects of chronic DAT downregulation on interval timing were mediated by the D2 receptors. Another cohort of DAT KD mice was trained on a visual attention task, and no deficits were observed, confirming that the changes in timed behavior were not attentionally mediated. Our data are consistent with the view that tonic dopamine affects the sensitivity of an organism to external reward signals, and that this increased motivation for reward of DAT KD mice lowers the threshold for initiating responding in a timing task.

Cocaine reward and locomotion stimulation in mice with reduced dopamine transporter expression

BackgroundThe dopamine transporter (DAT) plays a critical role in regulating dopamine neurotransmission. Variations in DAT or changes in basal dopaminergic tone have been shown to alter behavior and drug responses. DAT is one of the three known high affinity targets for cocaine, a powerful psychostimulant that produces reward and stimulates locomotor activity in humans and animals. We have shown that cocaine no longer produces reward in knock-in mice with a cocaine insensitive mutant DAT (DAT-CI), suggesting that cocaine inhibition of DAT is critical for its rewarding effect. However, in DAT-CI mice, the mutant DAT has significantly reduced uptake activity resulting in elevated basal dopaminergic tone, which might cause adaptive changes that alter responses to cocaine. Therefore, the objective of this study is to determine how elevated dopaminergic tone affects how mice respond to cocaine.ResultsWe examined the cocaine induced behavior of DAT knockdown mice that have DAT expression reduced by 90% when compared to the wild type mice. Despite a dramatic reduction of DAT expression and marked elevation in basal dopamine tone, cocaine produced reward, as measured by conditioned place preference, and stimulated locomotor activity in these mice.ConclusionA reduction in DAT expression and elevation of dopaminergic tone do not lead to adaptive changes that abolish the rewarding and stimulating effects of cocaine. Therefore, the lack of reward to cocaine observed in DAT-CI mice is unlikely to have resulted from the reduced DAT activity but instead is likely due to the inability of cocaine to block the mutated DAT and increase extracellular dopamine. This study supports the conclusion that the blockade of DAT is required for cocaine reward and locomotor stimulation.

Altered Corticostriatal Neurotransmission and Modulation in Dopamine Transporter Knock-Down Mice

Dopamine (DA) modulates glutamate neurotransmission in the striatum. Abnormal DA modulation has been implicated in neurological and psychiatric disorders. The development of DA transporter knock-down (DAT-KD) mice has permitted modeling of these disorders and has shed new light on DA modulation. DAT-KD mice exhibit increased extracellular DA, hyperactivity, and alterations in habituation. We used whole cell patch-clamp recordings from visually identified striatal neurons in slices to examine the effects of DAT-KD on corticostriatal transmission. Electrophysiological recordings from medium-sized spiny neurons in the dorsal striatum revealed alterations in both amplitude and frequency, of spontaneous glutamate receptor-mediated synaptic currents in cells from DAT-KD mice. Furthermore, kinetic analyses revealed that these currents had shorter half-amplitude durations and faster decay times. In contrast, GABA-receptor-mediated synaptic currents were not altered. Striatal neurons from DAT-KD mice also responded differently to amphetamine, cocaine, and DA D2-receptor agonists or antagonists compared with wildtype (WT) littermate controls. In WTs amphetamine and cocaine reduced the frequency of spontaneous glutamate currents and these effects appeared to be mediated by activation of D2 receptors. In contrast, in DAT-KD mice either no changes or only small increases in frequency occurred. D2-receptor agonists or antagonists also had opposing effects in WT and DAT-KD mice. Together, these results indicate that chronically increased extracellular DA produces long-lasting changes in corticostriatal communication that may be mediated by changes in D2-receptor function. These findings have implications for understanding mechanisms underlying attention deficit hyperactivity disorder and Tourette's syndrome and may provide insights into novel therapeutic approaches.