RESUMEN
An essential step toward understanding brain function is to establish a structural framework with cellular resolution on which multi-scale datasets spanning molecules, cells, circuits and systems can be integrated and interpreted1. Here, as part of the collaborative Brain Initiative Cell Census Network (BICCN), we derive a comprehensive cell type-based anatomical description of one exemplar brain structure, the mouse primary motor cortex, upper limb area (MOp-ul). Using genetic and viral labelling, barcoded anatomy resolved by sequencing, single-neuron reconstruction, whole-brain imaging and cloud-based neuroinformatics tools, we delineated the MOp-ul in 3D and refined its sublaminar organization. We defined around two dozen projection neuron types in the MOp-ul and derived an input-output wiring diagram, which will facilitate future analyses of motor control circuitry across molecular, cellular and system levels. This work provides a roadmap towards a comprehensive cellular-resolution description of mammalian brain architecture.
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Corteza Motora/anatomía & histología , Corteza Motora/citología , Neuronas/clasificación , Animales , Atlas como Asunto , Femenino , Neuronas GABAérgicas/citología , Neuronas GABAérgicas/metabolismo , Glutamatos/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Neuroimagen , Neuronas/citología , Neuronas/metabolismo , Especificidad de Órganos , Análisis de Secuencia de ARN , Análisis de la Célula IndividualRESUMEN
The anatomy of the mammalian visual system, from the retina to the neocortex, is organized hierarchically1. However, direct observation of cellular-level functional interactions across this hierarchy is lacking due to the challenge of simultaneously recording activity across numerous regions. Here we describe a large, open dataset-part of the Allen Brain Observatory2-that surveys spiking from tens of thousands of units in six cortical and two thalamic regions in the brains of mice responding to a battery of visual stimuli. Using cross-correlation analysis, we reveal that the organization of inter-area functional connectivity during visual stimulation mirrors the anatomical hierarchy from the Allen Mouse Brain Connectivity Atlas3. We find that four classical hierarchical measures-response latency, receptive-field size, phase-locking to drifting gratings and response decay timescale-are all correlated with the hierarchy. Moreover, recordings obtained during a visual task reveal that the correlation between neural activity and behavioural choice also increases along the hierarchy. Our study provides a foundation for understanding coding and signal propagation across hierarchically organized cortical and thalamic visual areas.
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Potenciales de Acción/fisiología , Corteza Visual/anatomía & histología , Corteza Visual/fisiología , Animales , Conjuntos de Datos como Asunto , Electrofisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Estimulación Luminosa , Tálamo/anatomía & histología , Tálamo/citología , Tálamo/fisiología , Corteza Visual/citologíaRESUMEN
The mammalian cortex is a laminar structure containing many areas and cell types that are densely interconnected in complex ways, and for which generalizable principles of organization remain mostly unknown. Here we describe a major expansion of the Allen Mouse Brain Connectivity Atlas resource1, involving around a thousand new tracer experiments in the cortex and its main satellite structure, the thalamus. We used Cre driver lines (mice expressing Cre recombinase) to comprehensively and selectively label brain-wide connections by layer and class of projection neuron. Through observations of axon termination patterns, we have derived a set of generalized anatomical rules to describe corticocortical, thalamocortical and corticothalamic projections. We have built a model to assign connection patterns between areas as either feedforward or feedback, and generated testable predictions of hierarchical positions for individual cortical and thalamic areas and for cortical network modules. Our results show that cell-class-specific connections are organized in a shallow hierarchy within the mouse corticothalamic network.
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Corteza Cerebral/anatomía & histología , Corteza Cerebral/citología , Vías Nerviosas/anatomía & histología , Vías Nerviosas/citología , Tálamo/anatomía & histología , Tálamo/citología , Animales , Axones/fisiología , Corteza Cerebral/fisiología , Femenino , Integrasas/genética , Integrasas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Vías Nerviosas/fisiología , Tálamo/fisiologíaRESUMEN
The maintenance of short-term memories is critical for survival in a dynamically changing world. Previous studies suggest that this memory can be stored in the form of persistent neural activity or using a synaptic mechanism, such as with short-term plasticity. Here, we compare the predictions of these two mechanisms to neural and behavioral measurements in a visual change detection task. Mice were trained to respond to changes in a repeated sequence of natural images while neural activity was recorded using two-photon calcium imaging. We also trained two types of artificial neural networks on the same change detection task as the mice. Following fixed pre-processing using a pretrained convolutional neural network, either a recurrent neural network (RNN) or a feedforward neural network with short-term synaptic depression (STPNet) was trained to the same level of performance as the mice. While both networks are able to learn the task, the STPNet model contains units whose activity are more similar to the in vivo data and produces errors which are more similar to the mice. When images are omitted, an unexpected perturbation which was absent during training, mice often do not respond to the omission but are more likely to respond to the subsequent image. Unlike the RNN model, STPNet produces a similar pattern of behavior. These results suggest that simple neural adaptation mechanisms may serve as an important bottom-up memory signal in this task, which can be used by downstream areas in the decision-making process.
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Adaptación Fisiológica , Memoria a Corto Plazo , Estimulación Luminosa , Percepción Visual , Animales , Conducta Animal , Biología Computacional/métodos , Toma de Decisiones , Ratones , Redes Neurales de la Computación , Análisis y Desempeño de TareasRESUMEN
Cocaine-experienced Wistar and Wistar Kyoto (WKY) rats received four daily repeated forced swim stress sessions (R-FSS), each of which preceded 4-hour cocaine self-administration sessions. Twenty-four hours after the last swim stress, cocaine valuation was assessed during a single-session threshold procedure. Prior exposure to R-FSS significantly altered cocaine responding in Wistar, but not WKY, rats. Behavioral economic analysis of responding revealed that the Wistar rats that had received R-FSS exhibited an increase in the maximum price that they were willing to pay for cocaine (Pmax ). Pre-treatment with the long-lasting kappa opioid receptor (KOR) antagonist norbinaltorphimine prevented the stress-induced increase in Pmax . Thus, R-FSS exposure had strain-dependent effects on cocaine responding during the threshold procedure, and the stress effects on cocaine valuation exhibited by Wistar, but not WKY, required intact KOR signaling.
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Cocaína/administración & dosificación , Inhibidores de Captación de Dopamina/administración & dosificación , Comportamiento de Búsqueda de Drogas/fisiología , Estrés Psicológico/fisiopatología , Animales , Comportamiento de Búsqueda de Drogas/efectos de los fármacos , Economía del Comportamiento , Naltrexona/análogos & derivados , Naltrexona/farmacología , Antagonistas de Narcóticos/farmacología , Ratas , Ratas Endogámicas WKY , Ratas Wistar , Autoadministración , NataciónRESUMEN
In complex environments, animals can adopt diverse strategies to find rewards. How distinct strategies differentially engage brain circuits is not well understood. Here, we investigate this question, focusing on the cortical Vip-Sst disinhibitory circuit between vasoactive intestinal peptide-postive (Vip) interneurons and somatostatin-positive (Sst) interneurons. We characterize the behavioral strategies used by mice during a visual change detection task. Using a dynamic logistic regression model, we find that individual mice use mixtures of a visual comparison strategy and a statistical timing strategy. Separately, mice also have periods of task engagement and disengagement. Two-photon calcium imaging shows large strategy-dependent differences in neural activity in excitatory, Sst inhibitory, and Vip inhibitory cells in response to both image changes and image omissions. In contrast, task engagement has limited effects on neural population activity. We find that the diversity of neural correlates of strategy can be understood parsimoniously as the increased activation of the Vip-Sst disinhibitory circuit during the visual comparison strategy, which facilitates task-appropriate responses.
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Interneuronas , Somatostatina , Péptido Intestinal Vasoactivo , Corteza Visual , Animales , Péptido Intestinal Vasoactivo/metabolismo , Corteza Visual/fisiología , Ratones , Somatostatina/metabolismo , Interneuronas/fisiología , Inhibición Neural/fisiología , Masculino , Ratones Endogámicos C57BL , Estimulación Luminosa/métodos , Percepción Visual/fisiologíaRESUMEN
To understand the neural basis of behavior, it is essential to measure spiking dynamics across many interacting brain regions. Although new technologies, such as Neuropixels probes, facilitate multi-regional recordings, significant surgical and procedural hurdles remain for these experiments to achieve their full potential. Here, we describe skull-shaped hemispheric implants enabling large-scale electrophysiology datasets (SHIELD). These 3D-printed skull-replacement implants feature customizable insertion holes, allowing dozens of cortical and subcortical structures to be recorded in a single mouse using repeated multi-probe insertions over many days. We demonstrate the procedure's high success rate, biocompatibility, lack of adverse effects on behavior, and compatibility with imaging and optogenetics. To showcase SHIELD's scientific utility, we use multi-probe recordings to reveal novel insights into how alpha rhythms organize spiking activity across visual and sensorimotor networks. Overall, this method enables powerful, large-scale electrophysiological experiments for the study of distributed neural computation.
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Encéfalo , Cráneo , Animales , Ratones , Encéfalo/fisiología , Cráneo/cirugía , Optogenética/métodos , Fenómenos Electrofisiológicos/fisiología , Impresión Tridimensional , Potenciales de Acción/fisiología , Electrodos Implantados , Ratones Endogámicos C57BL , Masculino , Electrofisiología/métodosRESUMEN
Multi-electrode arrays such as Neuropixels probes enable electrophysiological recordings from large populations of single neurons with high temporal resolution. By using such probes, the activity from functionally interacting, yet distinct, brain regions can be measured simultaneously by inserting multiple probes into the same subject. However, the use of multiple probes in small animals such as mice requires the removal of a sizable fraction of the skull, while also minimizing tissue damage and keeping the brain stable during the recordings. Here, we describe a step-by-step process designed to facilitate reliable recordings from up to six Neuropixels probes simultaneously in awake, head-fixed mice. The procedure involves four stages: the implantation of a headframe and a removable glass coverslip, the precise positioning of the Neuropixels probes at targeted points on the brain surface, the placement of a perforated plastic imaging window and the insertion of the probes into the brain of an awake mouse. The approach provides access to multiple brain regions and has been successfully applied across hundreds of mice. The procedure has been optimized for dense recordings from the mouse visual system, but it can be adapted for alternative recording configurations to target multiple probes in other brain areas. The protocol is suitable for users with experience in stereotaxic surgery in mice.
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Neuronas , Vigilia , Ratones , Animales , Vigilia/fisiología , Neuronas/fisiología , Encéfalo/fisiología , Electrodos , Cabeza , Electrodos ImplantadosRESUMEN
The classic view that neural populations in sensory cortices preferentially encode responses to incoming stimuli has been strongly challenged by recent experimental studies. Despite the fact that a large fraction of variance of visual responses in rodents can be attributed to behavioral state and movements, trial-history, and salience, the effects of contextual modulations and expectations on sensory-evoked responses in visual and association areas remain elusive. Here, we present a comprehensive experimental and theoretical study showing that hierarchically connected visual and association areas differentially encode the temporal context and expectation of naturalistic visual stimuli, consistent with the theory of hierarchical predictive coding. We measured neural responses to expected and unexpected sequences of natural scenes in the primary visual cortex (V1), the posterior medial higher order visual area (PM), and retrosplenial cortex (RSP) using 2-photon imaging in behaving mice collected through the Allen Institute Mindscope's OpenScope program. We found that information about image identity in neural population activity depended on the temporal context of transitions preceding each scene, and decreased along the hierarchy. Furthermore, our analyses revealed that the conjunctive encoding of temporal context and image identity was modulated by expectations of sequential events. In V1 and PM, we found enhanced and specific responses to unexpected oddball images, signaling stimulus-specific expectation violation. In contrast, in RSP the population response to oddball presentation recapitulated the missing expected image rather than the oddball image. These differential responses along the hierarchy are consistent with classic theories of hierarchical predictive coding whereby higher areas encode predictions and lower areas encode deviations from expectation. We further found evidence for drift in visual responses on the timescale of minutes. Although activity drift was present in all areas, population responses in V1 and PM, but not in RSP, maintained stable encoding of visual information and representational geometry. Instead we found that RSP drift was independent of stimulus information, suggesting a role in generating an internal model of the environment in the temporal domain. Overall, our results establish temporal context and expectation as substantial encoding dimensions in the visual cortex subject to fast representational drift and suggest that hierarchically connected areas instantiate a predictive coding mechanism.
RESUMEN
Identification of structural connections between neurons is a prerequisite to understanding brain function. Here we developed a pipeline to systematically map brain-wide monosynaptic input connections to genetically defined neuronal populations using an optimized rabies tracing system. We used mouse visual cortex as the exemplar system and revealed quantitative target-specific, layer-specific and cell-class-specific differences in its presynaptic connectomes. The retrograde connectivity indicates the presence of ventral and dorsal visual streams and further reveals topographically organized and continuously varying subnetworks mediated by different higher visual areas. The visual cortex hierarchy can be derived from intracortical feedforward and feedback pathways mediated by upper-layer and lower-layer input neurons. We also identify a new role for layer 6 neurons in mediating reciprocal interhemispheric connections. This study expands our knowledge of the visual system connectomes and demonstrates that the pipeline can be scaled up to dissect connectivity of different cell populations across the mouse brain.
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Conectoma , Corteza Visual , Ratones , Animales , Neuronas/fisiología , Encéfalo/fisiología , Corteza Visual/fisiología , Vías VisualesRESUMEN
Although the medial prefrontal cortex (mPFC) has been shown to be integrally involved in extinction of a number of associative behaviors, its role in extinction of alcohol (ethanol)-induced associative learning has received little attention. Previous reports have provided evidence supporting a role for the mPFC in acquisition and extinction of amphetamine-induced conditioned place preference (CPP) in rats, however, it remains unknown if this region is necessary for extinction of ethanol (EtOH)-induced associative learning in mice. Using immunohistochemical analysis of phosphorylated and unphosphorylated cAMP response element-binding protein (CREB), the current set of experiments first showed that the prelimbic (PL) and infralimbic (IL) subregions of the mPFC exhibited dynamic responses in phosphorylation of CREB to a Pavlovian-conditioned, EtOH-paired cue. Interestingly, CREB phosphorylation within these regions was sensitive to manipulations of the EtOH-cue contingency-that is, the cue-induced increase of pCREB in both the PL and IL was absent following extinction. In order to confirm a functional role of the mPFC in regulating the extinction process, we then showed that electrolytic lesions of the mPFC following acquisition blocked subsequent extinction of EtOH-CPP. Together, these experiments indicate a role for the PL and IL subregions of the mPFC in processing changes of the EtOH-cue contingency, as well as in regulating extinction of EtOH-induced associative learning in mice.
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Aprendizaje por Asociación/fisiología , Condicionamiento Clásico/fisiología , Etanol/administración & dosificación , Extinción Psicológica/fisiología , Corteza Prefrontal/fisiología , Animales , Aprendizaje por Asociación/efectos de los fármacos , Conducta Animal/efectos de los fármacos , Conducta Animal/fisiología , Condicionamiento Clásico/efectos de los fármacos , Extinción Psicológica/efectos de los fármacos , Masculino , Ratones , Corteza Prefrontal/efectos de los fármacosRESUMEN
We propose centralized brain observatories for large-scale recordings of neural activity in mice and non-human primates coupled with cloud-based data analysis and sharing. Such observatories will advance reproducible systems neuroscience and democratize access to the most advanced tools and data.
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Encéfalo , Neurociencias , Animales , RatonesRESUMEN
Despite significant progress in understanding neural coding, it remains unclear how the coordinated activity of large populations of neurons relates to what an observer actually perceives. Since neurophysiological differences must underlie differences among percepts, differentiation analysis-quantifying distinct patterns of neurophysiological activity-has been proposed as an "inside-out" approach that addresses this question. This methodology contrasts with "outside-in" approaches such as feature tuning and decoding analyses, which are defined in terms of extrinsic experimental variables. Here, we used two-photon calcium imaging in mice of both sexes to systematically survey stimulus-evoked neurophysiological differentiation (ND) in excitatory neuronal populations in layers (L)2/3, L4, and L5 across five visual cortical areas (primary, lateromedial, anterolateral, posteromedial, and anteromedial) in response to naturalistic and phase-scrambled movie stimuli. We find that unscrambled stimuli evoke greater ND than scrambled stimuli specifically in L2/3 of the anterolateral and anteromedial areas, and that this effect is modulated by arousal state and locomotion. By contrast, decoding performance was far above chance and did not vary substantially across areas and layers. Differentiation also differed within the unscrambled stimulus set, suggesting that differentiation analysis may be used to probe the ethological relevance of individual stimuli.
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Corteza Visual , Animales , Femenino , Locomoción/fisiología , Masculino , Ratones , Neuronas/fisiología , Neurofisiología , Estimulación Luminosa , Corteza Visual/fisiologíaRESUMEN
Extracellular electrophysiology and two-photon calcium imaging are widely used methods for measuring physiological activity with single-cell resolution across large populations of cortical neurons. While each of these two modalities has distinct advantages and disadvantages, neither provides complete, unbiased information about the underlying neural population. Here, we compare evoked responses in visual cortex recorded in awake mice under highly standardized conditions using either imaging of genetically expressed GCaMP6f or electrophysiology with silicon probes. Across all stimulus conditions tested, we observe a larger fraction of responsive neurons in electrophysiology and higher stimulus selectivity in calcium imaging, which was partially reconciled by applying a spikes-to-calcium forward model to the electrophysiology data. However, the forward model could only reconcile differences in responsiveness when restricted to neurons with low contamination and an event rate above a minimum threshold. This work established how the biases of these two modalities impact functional metrics that are fundamental for characterizing sensory-evoked responses.
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Electrofisiología/métodos , Neuronas/fisiología , Animales , Calcio , Señalización del Calcio , Genotipo , Ratones , Ratones Transgénicos , Neuronas/citología , Corteza Visual/citología , Corteza Visual/fisiologíaRESUMEN
The evolutionarily conserved default mode network (DMN) is a distributed set of brain regions coactivated during resting states that is vulnerable to brain disorders. How disease affects the DMN is unknown, but detailed anatomical descriptions could provide clues. Mice offer an opportunity to investigate structural connectivity of the DMN across spatial scales with cell-type resolution. We co-registered maps from functional magnetic resonance imaging and axonal tracing experiments into the 3D Allen mouse brain reference atlas. We find that the mouse DMN consists of preferentially interconnected cortical regions. As a population, DMN layer 2/3 (L2/3) neurons project almost exclusively to other DMN regions, whereas L5 neurons project in and out of the DMN. In the retrosplenial cortex, a core DMN region, we identify two L5 projection types differentiated by in- or out-DMN targets, laminar position, and gene expression. These results provide a multi-scale description of the anatomical correlates of the mouse DMN.
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Encéfalo/diagnóstico por imagen , Red en Modo Predeterminado/diagnóstico por imagen , Red Nerviosa/diagnóstico por imagen , Neuronas/fisiología , Animales , Encéfalo/citología , Conectoma , Red en Modo Predeterminado/citología , Imagen por Resonancia Magnética , Ratones , Red Nerviosa/citología , Neuronas/citologíaRESUMEN
Astrocytic inwardly rectifying K(+) currents (I(KIR)) have an important role in extracellular K(+) homeostasis, which influences neuronal excitability, and serum extravasation has been linked to impaired K(IR)-mediated K(+) buffering and chronic hyperexcitability. Head injury induces acute impairment in astroglial membrane I(KIR) and impaired K(+) buffering in the rat hippocampus, but chronic spontaneous seizures appear in the perilesional neocortex--not the hippocampus--in the early weeks to months after injury. Thus we examined astrocytic K(IR) channel pathophysiology in both neocortex and hippocampus after rostral parasaggital fluid percussion injury (rpFPI). rpFPI induced greater acute serum extravasation and metabolic impairment in the perilesional neocortex than in the underlying hippocampus, and in situ whole cell recordings showed a greater acute loss of astrocytic I(KIR) in neocortex than hippocampus. I(KIR) loss persisted through 1 mo after injury only in the neocortical epileptic focus, but fully recovered in the hippocampus that did not generate chronic seizures. Neocortical cell-attached recordings showed no loss or an increase of I(KIR) in astrocytic somata. Confocal imaging showed depletion of KIR4.1 immunoreactivity especially in processes--not somata--of neocortical astrocytes, whereas hippocampal astrocytes appeared normal. In naïve animals, intracortical infusion of serum, devoid of coagulation-mediating thrombin activity, reproduces the effects of rpFPI both in vivo and at the cellular level. In vivo serum infusion induces partial seizures similar to those induced by rpFPI, whereas bath-applied serum, but not dialyzed albumin, rapidly silenced astrocytic K(IR) membrane currents in whole cell and cell-attached patch-clamp recordings in situ. Thus both acute impairment in astrocytic I(KIR) and chronic spontaneous seizures typical of rpFPI are reproduced by serum extravasation, whereas the chronic impairment in astroglial I(KIR) is specific to the neocortex that develops the epileptic focus.
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Astrocitos/fisiología , Lesiones Encefálicas/fisiopatología , Epilepsias Parciales/fisiopatología , Neocórtex/lesiones , Canales de Potasio de Rectificación Interna/fisiología , Adenosina Difosfato/análisis , Adenosina Trifosfato/análisis , Animales , Lesiones Encefálicas/complicaciones , Cromatografía Líquida de Alta Presión , Electroencefalografía , Epilepsias Parciales/etiología , Exudados y Transudados , Masculino , Microscopía Confocal , Neocórtex/fisiopatología , Técnicas de Placa-Clamp , Ratas , Ratas Sprague-Dawley , Suero , Trombina/análisis , Grabación en VideoRESUMEN
To study the mechanisms of perception and cognition, neural measurements must be made during behavior. A goal of the Allen Brain Observatory is to map the activity of distinct cortical cell classes underlying visual and behavioral processing. Here we describe standardized methodology for training head-fixed mice on a visual change detection task, and we use our paradigm to characterize learning and behavior of five GCaMP6-expressing transgenic lines. We used automated training procedures to facilitate comparisons across mice. Training times varied, but most transgenic mice learned the behavioral task. Motivation levels also varied across mice. To compare mice in similar motivational states we subdivided sessions into over-, under-, and optimally motivated periods. When motivated, the pattern of perceptual decisions were highly correlated across transgenic lines, although overall performance (d-prime) was lower in one line labeling somatostatin inhibitory cells. These results provide important context for using these mice to map neural activity underlying perception and behavior.
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Cortical circuits can flexibly change with experience and learning, but the effects on specific cell types, including distinct inhibitory types, are not well understood. Here we investigated how excitatory and VIP inhibitory cells in layer 2/3 of mouse visual cortex were impacted by visual experience in the context of a behavioral task. Mice learned a visual change detection task with a set of eight natural scene images. Subsequently, during 2-photon imaging experiments, mice performed the task with these familiar images and three sets of novel images. Strikingly, the temporal dynamics of VIP activity differed markedly between novel and familiar images: VIP cells were stimulus-driven by novel images but were suppressed by familiar stimuli and showed ramping activity when expected stimuli were omitted from a temporally predictable sequence. This prominent change in VIP activity suggests that these cells may adopt different modes of processing under novel versus familiar conditions.
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Péptido Intestinal Vasoactivo/metabolismo , Animales , Ratones , Análisis y Desempeño de Tareas , Corteza Visual/metabolismo , Corteza Visual/fisiologíaRESUMEN
To understand how the brain processes sensory information to guide behavior, we must know how stimulus representations are transformed throughout the visual cortex. Here we report an open, large-scale physiological survey of activity in the awake mouse visual cortex: the Allen Brain Observatory Visual Coding dataset. This publicly available dataset includes the cortical activity of nearly 60,000 neurons from six visual areas, four layers, and 12 transgenic mouse lines in a total of 243 adult mice, in response to a systematic set of visual stimuli. We classify neurons on the basis of joint reliabilities to multiple stimuli and validate this functional classification with models of visual responses. While most classes are characterized by responses to specific subsets of the stimuli, the largest class is not reliably responsive to any of the stimuli and becomes progressively larger in higher visual areas. These classes reveal a functional organization wherein putative dorsal areas show specialization for visual motion signals.
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Corteza Visual/anatomía & histología , Corteza Visual/fisiología , Animales , Conjuntos de Datos como Asunto , RatonesRESUMEN
BACKGROUND: D-cycloserine (DCS), a partial N-methyl-D-aspartate receptor agonist, has been shown to enhance the extinction of both cocaine and amphetamine-induced conditioned place preference (CPP). However, there have been no reports of the effects of DCS on the extinction of ethanol-conditioned behaviors in mice. Thus, the current experiments examined the effects of DCS on the extinction and subsequent reconditioning of ethanol-induced CPP in mice. METHODS: Male DBA/2J mice received either 2 or 4 pairings of ethanol (2 g/kg) with a conditioned stimulus (CS+) floor cue (and an equal number of saline pairings with a CS- floor cue on alternate days) resulting in either a weak or strong ethanol CPP, respectively. Following conditioning of a strong ethanol CPP mice received saline or 30 mg/kg DCS prior to each of the twelve 30-minute choice extinction trials administered at 48-hour intervals. Mice that had received conditioning of a weak ethanol CPP received saline, 30 or 60 mg/kg DCS immediately before each of the six 30-minute choice extinction trials. Following successful ethanol CPP extinction, mice received reconditioning trials similar to the initial conditioning trials. A final experiment examined the effects 12 DCS pre-exposures (15, 30, and 60 mg/kg) on initial conditioning of ethanol CPP. RESULTS: First, we showed that 2 doses of DCS (30 and 60 mg/kg) did not have aversive properties that could confound the effects on extinction of CPP (Experiment 1). Second, we showed that DCS (30 and 60 mg/kg) had no effect on the rate of extinction of either strong (Experiment 2) or weak (Experiment 3) ethanol-induced CPP. Interestingly, DCS administered during extinction interfered with reconditioning of ethanol-induced CPP--an effect specific to reconditioning, as DCS pre-exposure did not influence initial ethanol CPP conditioning (Experiment 4). CONCLUSIONS: These experiments show that although DCS showed no effect on extinction behavior, when given during extinction it interfered with subsequent reconditioning of ethanol CPP. The mechanisms of this effect were not, however, due to nonspecific interference with learning because repeated DCS pre-exposures did not impair initial conditioning of ethanol CPP.