Experience-induced Arc/Arg3.1 primes CA1 pyramidal neurons for metabotropic glutamate receptor-dependent long-term synaptic depression.

A novel experience induces the Arc/Arg3.1 gene as well as plasticity of CA1 neural networks. To understand how these are linked, we briefly exposed GFP reporter mice of Arc transcription to a novel environment. Excitatory synaptic function of CA1 neurons with recent in vivo Arc induction (ArcGFP+) was similar to neighboring noninduced neurons. However, in response to group 1 metabotropic glutamate receptor (mGluR) activation, ArcGFP+ neurons preferentially displayed long-term synaptic depression (mGluR-LTD) and robust increases in dendritic Arc protein. mGluR-LTD in ArcGFP+ neurons required rapid protein synthesis and Arc, suggesting that dendritic translation of Arc underlies the priming of mGluR-LTD. In support of this idea, novelty exposure increased Arc messenger RNA in CA1 dendrites and promoted mGluR-induced translation of Arc in hippocampal synaptoneurosomes. Repeated experience suppressed synaptic transmission onto ArcGFP+ neurons and occluded mGluR-LTD ex vivo. mGluR-LTD priming in neurons with similar Arc activation history may contribute to encoding a novel environment.

Dopamine, cognitive function, and gamma oscillations: role of D4 receptors.

Cognitive deficits in individuals with schizophrenia (SCZ) are considered core symptoms of this disorder, and can manifest at the prodromal stage. Antipsychotics ameliorate positive symptoms but only modestly improve cognitive symptoms. The lack of treatments that improve cognitive abilities currently represents a major obstacle in developing more effective therapeutic strategies for this debilitating disorder. While D4 receptor (D4R)-specific antagonists are ineffective in the treatment of positive symptoms, animal studies suggest that D4R drugs can improve cognitive deficits. Moreover, recent work from our group suggests that D4Rs synergize with the neuregulin/ErbB4 signaling pathway, genetically identified as risk factors for SCZ, in parvalbumin (PV)-expressing interneurons to modulate gamma oscillations. These high-frequency network oscillations correlate with attention and increase during cognitive tasks in healthy subjects, and this correlation is attenuated in affected individuals. This finding, along with other observations indicating impaired GABAergic function, has led to the idea that abnormal neural activity in the prefrontal cortex (PFC) in individuals with SCZ reflects a perturbation in the balance of excitation and inhibition. Here we review the current state of knowledge of D4R functions in the PFC and hippocampus, two major brain areas implicated in SCZ. Special emphasis is given to studies focusing on the potential role of D4Rs in modulating GABAergic transmission and to an emerging concept of a close synergistic relationship between dopamine/D4R and neuregulin/ErbB4 signaling pathways that tunes the activity of PV interneurons to regulate gamma frequency network oscillations and potentially cognitive processes.

In vivo two-photon imaging of experience-dependent molecular changes in cortical neurons.

The brain's ability to change in response to experience is essential for healthy brain function, and abnormalities in this process contribute to a variety of brain disorders. To better understand the mechanisms by which brain circuits react to an animal's experience requires the ability to monitor the experience-dependent molecular changes in a given set of neurons, over a prolonged period of time, in the live animal. While experience and associated neural activity is known to trigger gene expression changes in neurons most of the methods to detect such changes do not allow repeated observation of the same neurons over multiple days or do not have sufficient resolution to observe individual neurons. Here, we describe a method that combines in vivo two-photon microscopy with a genetically encoded fluorescent reporter to track experience-dependent gene expression changes in individual cortical neurons over the course of day-to-day experience. One of the well-established experience-dependent genes is Activity-regulated cytoskeletal associated protein (Arc). The transcription of Arc is rapidly and highly induced by intensified neuronal activity and its protein product regulates the endocytosis of glutamate receptors and long-term synaptic plasticity. The expression of Arc has been widely used as a molecular marker to map neuronal circuits involved in specific behaviors. In most of those studies, Arc expression was detected by in situ hybridization or immunohistochemistry in fixed brain sections. Although those methods revealed that the expression of Arc was localized to a subset of excitatory neurons after behavioral experience, how the cellular patterns of Arc expression might change with multiple episodes of repeated or distinctive experiences over days was not investigated. In vivo two-photon microscopy offers a powerful way to examine experience-dependent cellular changes in the living brain. To enable the examination of Arc expression in live neurons by two-photon microscopy, we previously generated a knock-in mouse line in which a GFP reporter is placed under the control of the endogenous Arc promoter. This protocol describes the surgical preparations and imaging procedures for tracking experience-dependent Arc-GFP expression patterns in neuronal ensembles in the live animal. In this method, chronic cranial windows were first implanted in Arc-GFP mice over the cortical regions of interest. Those animals were then repeatedly imaged by two-photon microscopy after desired behavioral paradigms over the course of several days. This method may be generally applicable to animals carrying other fluorescent reporters of experience-dependent molecular changes.

Optogenetic inactivation modifies monkey visuomotor behavior.

A critical technique for understanding how neuronal activity contributes to behavior is determining whether perturbing it changes behavior. The advent of optogenetic techniques allows the immediately reversible alteration of neuronal activity in contrast to chemical approaches lasting minutes to hours. Modification of behavior using optogenetics has had substantial success in rodents but has not been as successful in monkeys. Here, we show how optogenetic inactivation of superior colliculus neurons in awake monkeys leads to clear and repeatable behavioral deficits in the metrics of saccadic eye movements. We used our observations to evaluate principles governing the use of optogenetic techniques in the study of the neuronal bases of behavior in monkeys, particularly how experimental design must address relevant parameters, such as the application of light to subcortical structures, the spread of viral injections, and the extent of neuronal inactivation with light.

Loss of Arc renders the visual cortex impervious to the effects of sensory experience or deprivation.

A myriad of mechanisms have been suggested to account for the full richness of visual cortical plasticity. We found that visual cortex lacking Arc is impervious to the effects of deprivation or experience. Using intrinsic signal imaging and chronic visually evoked potential recordings, we found that Arc(-/-) mice did not exhibit depression of deprived-eye responses or a shift in ocular dominance after brief monocular deprivation. Extended deprivation also failed to elicit a shift in ocular dominance or open-eye potentiation. Moreover, Arc(-/-) mice lacked stimulus-selective response potentiation. Although Arc(-/-) mice exhibited normal visual acuity, baseline ocular dominance was abnormal and resembled that observed after dark-rearing. These data suggest that Arc is required for the experience-dependent processes that normally establish and modify synaptic connections in visual cortex.

Differential gene expression between sensory neocortical areas: potential roles for Ten_m3 and Bcl6 in patterning visual and somatosensory pathways.

Adult neocortical areas are characterized by marked differences in cytoarchitecture and connectivity that underlie their functional roles. The molecular determinants of these differences are largely unknown. We performed a microarray analysis to identify molecules that define the somatosensory and visual areas during the time when afferent and efferent projections are forming. We identified 122 molecules that are differentially expressed between the regions and confirmed by quantitative polymerase chain reaction 95% of the 20 genes tested. Two genes were chosen for further investigation: Bcl6 and Ten_m3. Bcl6 was highly expressed in the superficial cortical plate corresponding to developing layer IV of somatosensory cortex at postnatal day (P) 0. This had diminished by P3, but strong expression was found in layer V pyramidal cells by P7 and was maintained until adulthood. Retrograde tracing showed that Bcl6 is expressed in corticospinal neurons. Ten_m3 was expressed in a graded pattern within layer V of caudal cortex that corresponds well with visual cortex. Retrograde tracing and immunostaining showed that Ten_m3 is highly expressed along axonal tracts of projection neurons of the developing visual pathway. Overexpression demonstrated that Ten_m3 promotes homophilic adhesion and neurite outgrowth in vivo. This suggests an important role for Ten_m3 in the development of the visual pathway.