The rostral intralaminar nuclear complex of the thalamus supports striatally mediated action reinforcement.

The dorsal striatum (DS) mediates the selection of actions for reward acquisition necessary for survival. Striatal pathology contributes to several neuropsychiatric conditions, including aberrant selection of actions for specific rewards in addiction. A major source of glutamate driving striatal activity is the rostral intralaminar nuclei (rILN) of the thalamus. Yet, the information that is relayed to the striatum to support action selection is unknown. Here, we discovered that rILN neurons projecting to the DS are innervated by a range of cortical and subcortical afferents and that rILN→DS neurons stably signaled at two time points in mice performing an action sequence task reinforced by sucrose reward: action initiation and reward acquisition. In vivo activation of this pathway increased the number of successful trials, whereas inhibition decreased the number of successful trials. These findings illuminate a role for the rostral intralaminar nuclear complex in reinforcing actions.

Inducible CRISPR Epigenome Systems Mimic Cocaine Induced Bidirectional Regulation of Nab2 and Egr3.

Substance use disorder is a chronic disease and a leading cause of disability around the world. The NAc is a major brain hub mediating reward behavior. Studies demonstrate exposure to cocaine is associated with molecular and functional imbalance in NAc medium spiny neuron subtypes (MSNs), dopamine receptor 1 and 2 enriched D1-MSNs and D2-MSNs. We previously reported repeated cocaine exposure induced transcription factor early growth response 3 (Egr3) mRNA in NAc D1-MSNs, and reduced it in D2-MSNs. Here, we report our findings of repeated cocaine exposure in male mice inducing MSN subtype-specific bidirectional expression of the Egr3 corepressor NGFI-A-binding protein 2 (Nab2). Using CRISPR activation and interference (CRISPRa and CRISPRi) tools combined with Nab2 or Egr3-targeted sgRNAs, we mimicked these bidirectional changes in Neuro2a cells. Furthermore, we investigated D1-MSN- and D2-MSN-specific expressional changes of histone lysine demethylases Kdm1a, Kdm6a, and Kdm5c in NAc after repeated cocaine exposure in male mice. Since Kdm1a showed bidirectional expression patterns in D1-MSNs and D2-MSNs, like Egr3, we developed a light-inducible Opto-CRISPR-KDM1a system. We were able to downregulate Egr3 and Nab2 transcripts in Neuro2A cells and cause similar bidirectional expression changes we observed in D1-MSNs and D2-MSNs of mouse repeated cocaine exposure model. Contrastingly, our Opto-CRISPR-p300 activation system induced the Egr3 and Nab2 transcripts and caused opposite bidirectional transcription regulations. Our study sheds light on the expression patterns of Nab2 and Egr3 in specific NAc MSNs in cocaine action and uses CRISPR tools to further mimic these expression patterns.SIGNIFICANCE STATEMENT Substance use disorder is a major societal issue. The lack of medication to treat cocaine addiction desperately calls for a treatment development based on precise understanding of molecular mechanisms underlying cocaine addiction. In this study, we show that Egr3 and Nab2 are bidirectionally regulated in mouse NAc D1-MSNs and D2-MSNs after repeated exposure to cocaine. Furthermore, histone lysine demethylations enzymes with putative EGR3 binding sites showed bidirectional regulation in D1- and D2-MSNs after repeated exposure to cocaine. Using Cre- and light-inducible CRISPR tools, we show that we can mimic this bidirectional regulation of Egr3 and Nab2 in Neuro2a cells.

Rostral Intralaminar Thalamus Engagement in Cognition and Behavior.

The thalamic rostral intralaminar nuclei (rILN) are a contiguous band of neurons that include the central medial, paracentral, and central lateral nuclei. The rILN differ from both thalamic relay nuclei, such as the lateral geniculate nucleus, and caudal intralaminar nuclei, such as the parafascicular nucleus, in afferent and efferent connectivity as well as physiological and synaptic properties. rILN activity is associated with a range of neural functions and behaviors, including arousal, pain, executive function, and action control. Here, we review this evidence supporting a role for the rILN in integrating arousal, executive and motor feedback information. In light of rILN projections out to the striatum, amygdala, and sensory as well as executive cortices, we propose that such a function enables the rILN to modulate cognitive and motor resources to meet task-dependent behavioral engagement demands.

Axo-axonic synapses: Diversity in neural circuit function.

The chemical synapse is the principal form of contact between neurons of the central nervous system. These synapses are typically configured as presynaptic axon terminations onto postsynaptic dendrites or somata, giving rise to axo-dendritic and axo-somatic synapses, respectively. Beyond these common synapse configurations are less-studied, non-canonical synapse types that are prevalent throughout the brain and significantly contribute to neural circuit function. Among these are the axo-axonic synapses, which consist of an axon terminating on another axon or axon terminal. Here, we review evidence for axo-axonic synapse contributions to neural signaling in the mammalian nervous system and survey functional neural circuit motifs enabled by these synapses. We also detail how recent advances in microscopy, transgenics, and biological sensors may be used to identify and functionally assay axo-axonic synapses.

Activation of the Rostral Intralaminar Thalamus Drives Reinforcement through Striatal Dopamine Release.

Glutamatergic projections of the thalamic rostral intralaminar nuclei of the thalamus (rILN) innervate the dorsal striatum (DS) and are implicated in dopamine (DA)-dependent incubation of drug seeking. However, the mechanism by which rILN signaling modulates reward seeking and striatal DA release is unknown. We find that activation of rILN inputs to the DS drives cholinergic interneuron burst-firing behavior and DA D2 receptor-dependent post-burst pauses in cholinergic interneuron firing. In vivo, optogenetic activation of this pathway drives reinforcement in a DA D1 receptor-dependent manner, and chemogenetic suppression of the rILN reduces dopaminergic nigrostriatal terminal activity as measured by fiber photometry. Altogether, these data provide evidence that the rILN activates striatal cholinergic interneurons to enhance the pursuit of reward through local striatal DA release and introduce an additional level of complexity in our understanding of striatal DA signaling.

Estradiol shifts interactions between the infralimbic cortex and central amygdala to enhance fear extinction memory in female rats.

There is growing evidence that estradiol (E2) enhances fear extinction memory consolidation. However, it is unclear how E2 influences the nodes of the fear extinction network to enhance extinction memory. This study begins to delineate the neural circuits underlying the influence of E2 on fear extinction acquisition and consolidation in female rats. After fear conditioning (day 1), naturally cycling female rats underwent extinction learning (day 2) in a low-E2 state, receiving a systemic administration of either E2 or vehicle prior to extinction training. Extinction memory recall was then tested 24 hr later (day 3). We measured immediate early gene c-fos expression within the extinction network during fear extinction learning and extinction recall. During extinction learning, E2 treatment increased centrolateral amygdala c-fos activity and reduced lateral amygdala activity relative to vehicle. During extinction recall, E2-treated rats exhibited reduced c-fos expression in the centromedial amygdala. There were no group differences in c-fos expression within the medial prefrontal cortex or dorsal hippocampus. Examining c-fos ratios with the infralimbic cortex (IL) revealed that, despite the lack of group differences within the IL, E2 treatment induced greater IL activity relative to both prelimbic cortex and central amygdala (CeA) activity during extinction memory recall. Only the relationship between IL and CeA activity positively correlated with extinction retention. In conclusion, E2 appears to modify interactions between the IL and the CeA in females, shifting from stronger amygdalar modulation of fear during extinction learning to stronger IL control during extinction recall. © 2016 Wiley Periodicals, Inc.

Protocol for studying extinction of conditioned fear in naturally cycling female rats.

Extinction of conditioned fear has been extensively studied in male rodents. Recently, there have been an increasing number of studies indicating that neural mechanisms for certain behavioral tasks and response behaviors are different in females and males. Using females in research studies can represent a challenge because of the variation of gonadal hormones during their estrous cycle. This protocol describes well-established procedures that are useful in investigating the role of estrogen in fear extinction memory consolidation in female rats. Phase of the estrous cycle and exogenous estrogen administration prior to extinction training can influence extinction recall 24 hr later. The vaginal swabbing technique for estrous phase identification described here aids the examination and manipulation of naturally cycling gonadal hormones. The use of this basic rodent model may further delineate the mechanisms by which estrogen can modulate fear extinction memory in females.