Creation of a novel CRISPR-generated allele to express HA epitope-tagged C1QL1 and improved methods for its detection at synapses.

C1QL1 is expressed in a subset of cells in the brain and likely has pleiotropic functions, including the regulation of neuron-to-neuron synapses. Research progress on C1QL proteins has been slowed by a dearth of available antibodies. Therefore, we created a novel knock-in mouse line in which an HA-tag is inserted into the endogenous C1ql1 locus. We examined the entire brain, identifying previously unappreciated nuclei expressing C1QL1, presumably in neurons. By total numbers, however, the large majority of C1QL1-expressing cells are of the oligodendrocyte lineage. Subcellular immunolocalization of synaptic cleft proteins is challenging, so we developed a new protocol to improve signal at synapses. Lastly, we compared various anti-HA antibodies to assist future investigations using this and likely other HA epitope-tagged alleles.

C1ql1 expression in oligodendrocyte progenitor cells promotes oligodendrocyte differentiation.

Myelinating oligodendrocytes arise from the stepwise differentiation of oligodendrocyte progenitor cells (OPCs). Approximately 5% of all adult brain cells are OPCs. Why would a mature brain need such a large number of OPCs? New myelination is possibly required for higher-order functions such as cognition and learning. Additionally, this pool of OPCs represents a source of new oligodendrocytes to replace those lost during injury, inflammation, or in diseases such as multiple sclerosis (MS). How OPCs are instructed to differentiate into oligodendrocytes is poorly understood, and for reasons presently unclear, resident pools of OPCs are progressively less utilized in MS. The complement component 1, q subcomponent-like (C1QL) protein family has been studied for their functions at neuron-neuron synapses, but we show that OPCs express C1ql1. We created OPC-specific conditional knockout mice and show that C1QL1 deficiency reduces the differentiation of OPCs into oligodendrocytes and reduces myelin production during both development and recovery from cuprizone-induced demyelination. In vivo over-expression of C1QL1 causes the opposite phenotype: increased oligodendrocyte density and myelination during recovery from demyelination. We further used primary cultured OPCs to show that C1QL1 levels can bidirectionally regulate the extent of OPC differentiation in vitro. Our results suggest that C1QL1 may initiate a previously unrecognized signaling pathway to promote differentiation of OPCs into oligodendrocytes. This study has relevance for possible novel therapies for demyelinating diseases and may illuminate a previously undescribed mechanism to regulate the function of myelination in cognition and learning.

Frontal-Sensory Cortical Projections Become Dispensable for Attentional Performance Upon a Reduction of Task Demand in Mice.

Top-down attention is a dynamic cognitive process that facilitates the detection of the task-relevant stimuli from our complex sensory environment. A neural mechanism capable of deployment under specific task-demand conditions would be crucial to efficiently control attentional processes and improve promote goal-directed attention performance during fluctuating attentional demand. Previous studies have shown that frontal top-down neurons projecting from the anterior cingulate area (ACA) to the visual cortex (VIS; ACAVIS) are required for visual attentional behavior during the 5-choice serial reaction time task (5CSRTT) in mice. However, it is unknown whether the contribution of these projecting neurons is dependent on the extent of task demand. Here, we first examined how behavior outcomes depend on the number of locations for mice to pay attention and touch for successful performance, and found that the 2-choice serial reaction time task (2CSRTT) is less task demanding than the 5CSRTT. We then employed optogenetics to demonstrate that suppression ACAVIS projections immediately before stimulus presentation has no effect during the 2CSRTT in contrast to the impaired performance during the 5CSRTT. These results suggest that ACAVIS projections are necessary when task demand is high, but once a task demand is lowered, ACAVIS neuron activity becomes dispensable to adjust attentional performance. These findings support a model that the frontal-sensory ACAVIS projection regulates visual attention behavior during specific high task demand conditions, pointing to a flexible circuit-based mechanism for promoting attentional behavior.

Chemogenetic suppression of anterior cingulate cortical neurons projecting to the visual cortex disrupts attentional behavior in mice.

AIM: Attention is a goal-directed cognitive process that facilitates the detection of task-relevant sensory stimuli from dynamic environments. Anterior cingulate cortical area (ACA) is known to play a key role in attentional behavior, but the specific circuits mediating attention remain largely unknown. As ACA modulates sensory processing in the visual cortex (VIS), we aim to test a hypothesis that frontal top-down neurons projecting from ACA to VIS (ACAVIS ) contributes to visual attention behavior through chemogenetic approach. METHODS: Adult, male mice were trained to perform the 5-choice serial reaction time task (5CSRTT) using a touchscreen system. An intersectional viral approach was used to selectively express inhibitory designer receptors exclusively activated by designer drugs (iDREADD) or a static fluorophore (mCherry) in ACAVIS neurons. Mice received counterbalanced injections (i.p.) of the iDREADD ligand (clozapine-N-oxide; CNO) or vehicle (saline) prior to 5CSRTT testing. Finally, mice underwent progressive ratio testing and open field testing following CNO or saline administration. RESULTS: Chemogenetic suppression of ACAVIS neuron activity decreased correct task performance during the 5CSRTT mainly driven by an increase in omission and a trending decrease in accuracy with no change in behavioral outcomes associated with motivation, impulsivity, or compulsivity. Breakpoint during the progressive ratio task and distance moved in the open field test were unaffected by ACAVIS neuron suppression. CNO administration itself had no effect on task performance in mCherry-expressing mice. CONCLUSION: These results identify long-range frontal-sensory ACAVIS projection neurons as a key enactor of top-down attentional behavior and may serve as a beneficial therapeutic target.

Nicotinic regulation of local and long-range input balance drives top-down attentional circuit maturation.

Cognitive function depends on frontal cortex development; however, the mechanisms driving this process are poorly understood. Here, we identify that dynamic regulation of the nicotinic cholinergic system is a key driver of attentional circuit maturation associated with top-down frontal neurons projecting to visual cortex. The top-down neurons receive robust cholinergic inputs, but their nicotinic tone decreases following adolescence by increasing expression of a nicotinic brake, Lynx1 Lynx1 shifts a balance between local and long-range inputs onto top-down frontal neurons following adolescence and promotes the establishment of attentional behavior in adulthood. This key maturational process is disrupted in a mouse model of fragile X syndrome but was rescued by a suppression of nicotinic tone through the introduction of Lynx1 in top-down projections. Nicotinic signaling may serve as a target to rebalance local/long-range balance and treat cognitive deficits in neurodevelopmental disorders.

Post-error recruitment of frontal sensory cortical projections promotes attention in mice.

The frontal cortex, especially the anterior cingulate cortex area (ACA), is essential for exerting cognitive control after errors, but the mechanisms that enable modulation of attention to improve performance after errors are poorly understood. Here we demonstrate that during a mouse visual attention task, ACA neurons projecting to the visual cortex (VIS; ACAVIS neurons) are recruited selectively by recent errors. Optogenetic manipulations of this pathway collectively support the model that rhythmic modulation of ACAVIS neurons in anticipation of visual stimuli is crucial for adjusting performance following errors. 30-Hz optogenetic stimulation of ACAVIS neurons in anesthetized mice recapitulates the increased gamma and reduced theta VIS oscillatory changes that are associated with endogenous post-error performance during behavior and subsequently increased visually evoked spiking, a hallmark feature of visual attention. This frontal sensory neural circuit links error monitoring with implementing adjustments of attention to guide behavioral adaptation, pointing to a circuit-based mechanism for promoting cognitive control.

A prefrontal-paraventricular thalamus circuit requires juvenile social experience to regulate adult sociability in mice.

Juvenile social isolation reduces sociability in adulthood, but the underlying neural circuit mechanisms are poorly understood. We found that, in male mice, 2 weeks of social isolation immediately following weaning leads to a failure to activate medial prefrontal cortex neurons projecting to the posterior paraventricular thalamus (mPFC→pPVT) during social exposure in adulthood. Chemogenetic or optogenetic suppression of mPFC→pPVT activity in adulthood was sufficient to induce sociability deficits without affecting anxiety-related behaviors or preference toward rewarding food. Juvenile isolation led to both reduced excitability of mPFC→pPVT neurons and increased inhibitory input drive from low-threshold-spiking somatostatin interneurons in adulthood, suggesting a circuit mechanism underlying sociability deficits. Chemogenetic or optogenetic stimulation of mPFC→pPVT neurons in adulthood could rescue the sociability deficits caused by juvenile isolation. Our study identifies a pair of specific medial prefrontal cortex excitatory and inhibitory neuron populations required for sociability that are profoundly affected by juvenile social experience.

Adolescent frontal top-down neurons receive heightened local drive to establish adult attentional behavior in mice.

Frontal top-down cortical neurons projecting to sensory cortical regions are well-positioned to integrate long-range inputs with local circuitry in frontal cortex to implement top-down attentional control of sensory regions. How adolescence contributes to the maturation of top-down neurons and associated local/long-range input balance, and the establishment of attentional control is poorly understood. Here we combine projection-specific electrophysiological and rabies-mediated input mapping in mice to uncover adolescence as a developmental stage when frontal top-down neurons projecting from the anterior cingulate to visual cortex are highly functionally integrated into local excitatory circuitry and have heightened activity compared to adulthood. Chemogenetic suppression of top-down neuron activity selectively during adolescence, but not later periods, produces long-lasting visual attentional behavior deficits, and results in excessive loss of local excitatory inputs in adulthood. Our study reveals an adolescent sensitive period when top-down neurons integrate local circuits with long-range connectivity to produce attentional behavior.