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CRISPR-Cas9-Mediated Intersectional Knockout of Glycogen Synthase Kinase 3? in D2 Receptor-Expressing Medial Prefrontal Cortex Neurons Reveals Contributions to Emotional Regulation.

ABSTRACT: Glycogen synthase kinase 3? (GSK3?) activity is regulated by dopamine D2 receptor signaling and can be inhibited by psychoactive drugs in a D2 receptor-dependent manner. However, GSK3? is ubiquitously expressed in the brain, and D2 receptor-expressing cells are distributed as a mosaic in multiple cortical regions. This complicates the interrogation of GSK3? functions in cortical D2 cells in a circuit-defined manner using conventional animal models. We used a CRISPR-Cas9-mediated intersectional approach to achieve targeted deletion of GSK3? in D2-expressing neurons of the adult medial prefrontal cortex (mPFC). Isolation and analysis of ribosome-associated RNA specifically from mPFC D2 neurons lacking GSK3? demonstrated large-scale translatome alterations. Deletion of GSK3? in mPFC D2 neurons revealed its contribution to anxiety-related, cognitive, and social behaviors. Our results underscore the viability of an intersectional knockout approach to study functions of a ubiquitous gene in a network-defined fashion while uncovering the contribution of GSK3? expressed in mPFC D2 neurons in the regulation of behavioral dimensions related to mood and emotions. This advances our understanding of GSK3? action at a brain circuit level and can potentially lead to the development of circuit selective therapeutics.

SUBMITTER: Khlghatyan J

PROVIDER: S-EPMC7307679 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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CRISPR-Cas9-Mediated Intersectional Knockout of Glycogen Synthase Kinase 3β in D2 Receptor-Expressing Medial Prefrontal Cortex Neurons Reveals Contributions to Emotional Regulation.

Khlghatyan Jivan J Beaulieu Jean-Martin JM

The CRISPR journal 20200601 3

Glycogen synthase kinase 3β (GSK3β) activity is regulated by dopamine D2 receptor signaling and can be inhibited by psychoactive drugs in a D2 receptor-dependent manner. However, GSK3β is ubiquitously expressed in the brain, and D2 receptor-expressing cells are distributed as a mosaic in multiple cortical regions. This complicates the interrogation of GSK3β functions in cortical D2 cells in a circuit-defined manner using conventional animal models. We used a CRISPR-Cas9-mediated intersectional a ...[more]

PMID: 32584144

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Project description:OBJECTIVES:Implicit regulation of emotions involves medial-prefrontal cortex (mPFC) regions exerting regulatory control over limbic structures. Diminished regulation relates to aberrant mPFC functionality and psychopathology. Establishing means of modulating mPFC functionality could benefit research on emotion and its dysregulation. Here, we tested the capacity of transcranial direct current stimulation (tDCS) targeting mPFC to modulate subjective emotional states by facilitating implicit emotion regulation. MATERIALS AND METHODS:Stimulation was applied concurrently with functional magnetic resonance imaging to validate its neurobehavioral effect. Sixteen participants were each scanned twice, counterbalancing active and sham tDCS application, while undergoing negative mood induction (clips featuring negative vs. neutral contents). Effects of stimulation on emotional experience were assessed using subjective and neural measures. RESULTS:Subjectively, active stimulation led to significant reduction in reported intensity of experienced emotions to negatively valenced (p?=?0.005) clips but not to neutral clips (p?>?0.99). Active stimulation further mitigated a rise in stress levels from pre- to post-induction (sham: p?=?0.004; active: p?=?0.15). Neurally, stimulation increased activation in mPFC regions associated with implicit emotion regulation (ventromedial-prefrontal cortex; subgenual anterior-cingulate cortex, sgACC), and in ventral striatum, a core limbic structure (all ps?<?0.05). Stimulation also altered functional connectivity (assessed using whole-brain psycho-physiological interaction) between these regions, and with additional limbic regions. Stimulation-induced sgACC activation correlated with reported emotion intensity and depressive symptoms (rs?>?0.64, ps?<?0.018), suggesting individual differences in stimulation responsivity. CONCLUSIONS:Results of this study indicate the potential capacity of tDCS to facilitate brain activation in mPFC regions underlying implicit regulation of emotion and accordingly modulate subjective emotional experiences.

Project description:Decreased expression of dopamine D2 receptors (D2R), dysfunction of inhibitory neurotransmission and impairments in the structure and connectivity of neurons in the medial prefrontal cortex (mPFC) are involved in the pathogenesis of schizophrenia and major depression, but the relationship between these changes remains unclear. The polysialylated form of the neural cell adhesion molecule (PSA-NCAM), a plasticity-related molecule, may serve as a link. This molecule is expressed in cortical interneurons and dopamine, via D2R, modulates its expression in parallel to that of proteins related to synapses and inhibitory neurotransmission, suggesting that D2R-targeted antipsychotics/antidepressants may act by affecting the plasticity of mPFC inhibitory circuits. To understand the role of PSA-NCAM in this plasticity, rats were chronically treated with a D2R agonist (PPHT) after cortical PSA depletion. PPHT-induced increases in GAD67 and synaptophysin (SYN) neuropil expression were blocked when PSA was previously removed, indicating a role for PSA-NCAM in this plasticity. The number of PSA-NCAM expressing interneuron somata also increased after PPHT treatment, but the percentages of these cells belonging to different interneuronal subpopulations did not change. Cortical pyramidal neurons did not express PSA-NCAM, but puncta co-expressing this molecule and parvalbumin could be found surrounding their somata. PPHT treatment increased the number of PSA-NCAM and parvalbumin expressing perisomatic puncta, but decreased the percentage of parvalbumin puncta that co-expressed SYN. PSA depletion did not block these effects on the perisomatic region, but increased further the number of parvalbumin expressing puncta and increased the percentage of puncta co-expressing SYN and parvalbumin, suggesting that the polysialylation of NCAM may regulate perisomatic inhibition of mPFC principal neurons. Summarizing, the present results indicate that dopamine acting on D2R influences structural plasticity of mPFC interneurons and point to PSA-NCAM as a key player in this remodeling.

Dataset Information

CRISPR-Cas9-Mediated Intersectional Knockout of Glycogen Synthase Kinase 3? in D2 Receptor-Expressing Medial Prefrontal Cortex Neurons Reveals Contributions to Emotional Regulation.

Publications

CRISPR-Cas9-Mediated Intersectional Knockout of Glycogen Synthase Kinase 3β in D2 Receptor-Expressing Medial Prefrontal Cortex Neurons Reveals Contributions to Emotional Regulation.

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