Long-term memory formation requires the coordinated regulation of gene expression. Until recently nucleosome remodeling, one of the major epigenetic mechanisms for controlling gene expression, had been largely unexplored in the field of neuroscience. Nucleosome remodeling is carried out by chromatin remodeling complexes (CRCs) that interact with DNA and histones to physically alter chromatin structure and ultimately regulate gene expression. Human exome sequencing and gene wide association studies have linked mutations in CRC subunits to intellectual disability disorders, autism spectrum disorder and schizophrenia. However, how mutations in CRC subunits were related to human cognitive disorders was unknown. There appears to be both developmental and adult specific roles for the neuron specific CRC nBAF (neuronal Brg1/hBrm Associated Factor). nBAF regulates gene expression required for dendritic arborization during development, and in the adult, contributes to long-term potentiation, a form of synaptic plasticity, and long-term memory. We propose that the nBAF complex is a novel epigenetic mechanism for regulating transcription required for long-lasting forms of synaptic plasticity and memory processes and that impaired nBAF function may result in human cognitive disorders.

This unit is designed to provide sufficient instruction for the setup and execution of tests for object location and object recognition in adult mice. This task is ideally suited for the study of a variety of mouse models that examine disease mechanisms and novel therapeutic targets. By altering several key parameters, the experimenter can investigate short-term or long-term memory and look for either memory impairments or enhancements. Object location and object recognition memory tasks rely on a rodent's innate preference for novelty, and can be conducted sequentially in the same cohort of animals. These two tasks avoid the inherent stress induced with other common measures of rodent memory such as fear conditioning and the Morris water maze. This protocol covers detailed instructions on conducting both tasks, as well as key points concerning data collection, analysis, and interpretation.

Designer receptors exclusively activated by designer drug (DREADDs) are a novel tool with the potential to bidirectionally drive cellular, circuit, and ultimately, behavioral changes. We used DREADDs to evaluate memory formation in a hippocampus-dependent task in mice and effects on synaptic physiology in the dorsal hippocampus. We expressed neuron-specific (hSyn promoter) DREADDs that were either excitatory (HM3D) or inhibitory (HM4D) in the dorsal hippocampus. As predicted, hSyn-HM3D was able to transform a subthreshold learning event into long-term memory (LTM), and hSyn-HM4D completely impaired LTM formation. Surprisingly, the opposite was observed during experiments examining the effects on hippocampal long-term potentiation (LTP). hSyn-HM3D impaired LTP and hSyn-HM4D facilitated LTP. Follow-up experiments indicated that the hSyn-HM3D-mediated depression of fEPSP appears to be driven by presynaptic activation of inhibitory currents, whereas the hSyn-HM4D-mediated increase of fEPSP is induced by a reduction in GABAA receptor function. To determine whether these observations were promoter specific, we next examined the effects of using the CaMKIIα promoter that limits expression to forebrain excitatory neurons. CaMKIIα-HM3D in the dorsal hippocampus led to the transformation of a subthreshold learning event into LTM, whereas CaMKIIα-HM4D blocked LTM formation. Consistent with these findings, baseline synaptic transmission and LTP was increased in CaMKIIα-HM3D hippocampal slices, whereas slices from CaMKIIα-HM4D mice produced expected decreases in baseline synaptic transmission and LTP. Together, these experiments further demonstrate DREADDs as being a robust and reliable means of modulating neuronal function to manipulate long-term changes in behavior, while providing evidence for specific dissociations between LTM and LTP.

Significance statement

This study evaluates the efficacy of designer receptors exclusively activated by designer drug (DREADDs) as a means of bidirectionally modulating the hippocampus in not only a hippocampus-dependent task but also in hippocampal synaptic plasticity. This is the first study to evaluate the effects of DREADD-mediated inhibition and excitation in hippocampal long-term potentiation. More specifically, this study evaluates the effect of promoter-specific expression of DREADD viruses in a heterogenic cell population, which revealed surprising effects of different promoters. With chemogenetics becoming a more ubiquitous tool throughout studies investigating circuit-specific function, these data are of broad interest to the neuroscientific community because we have shown that promoter-specific effects can drastically alter synaptic function within a specific region, without parallel changes at the level of behavior.

Rhythmic oscillations of physiological processes depend on integrating the circadian clock and diurnal environment. DNA methylation is epigenetically responsive to daily rhythms, as a subset of CpG dinucleotides in brain exhibit diurnal rhythmic methylation. Here, we show a major genetic effect on rhythmic methylation in a mouse Snord116 deletion model of the imprinted disorder Prader-Willi syndrome (PWS). More than 23,000 diurnally rhythmic CpGs are identified in wild-type cortex, with nearly all lost or phase-shifted in PWS. Circadian dysregulation of a second imprinted Snord cluster at the Temple/Kagami-Ogata syndrome locus is observed at the level of methylation, transcription, and chromatin, providing mechanistic evidence of cross-talk. Genes identified by diurnal epigenetic changes in PWS mice overlapped rhythmic and PWS-specific genes in human brain and are enriched for PWS-relevant phenotypes and pathways. These results support the proposed evolutionary relationship between imprinting and sleep, and suggest possible chronotherapy in the treatment of PWS and related disorders.

Transcriptional dysregulation is an early feature of Huntington disease (HD). We observed gene-specific changes in histone H3 lysine 4 trimethylation (H3K4me3) at transcriptionally repressed promoters in R6/2 mouse and human HD brain. Genome-wide analysis showed a chromatin signature for this mark. Reducing the levels of the H3K4 demethylase SMCX/Jarid1c in primary neurons reversed down-regulation of key neuronal genes caused by mutant Huntingtin expression. Finally, reduction of SMCX/Jarid1c in primary neurons from BACHD mice or the single Jarid1 in a Drosophila HD model was protective. Therefore, targeting this epigenetic signature may be an effective strategy to ameliorate the consequences of HD.

Transcriptional dysregulation is an early feature of Huntington disease (HD). We observed gene-specific changes in histone H3 lysine 4 trimethylation (H3K4me3) at transcriptionally repressed promoters in R6/2 mouse and human HD brain. Genome-wide analysis showed a chromatin signature for this mark. Reducing the levels of the H3K4 demethylase SMCX/Jarid1c in primary neurons reversed down-regulation of key neuronal genes caused by mutant Huntingtin expression. Finally, reduction of SMCX/Jarid1c in primary neurons from BACHD mice or the single Jarid1 in a Drosophila HD model was protective. Therefore, targeting this epigenetic signature may be an effective strategy to ameliorate the consequences of HD.

Aging is accompanied by impairments in both circadian rhythmicity and long-term memory. Although it is clear that memory performance is affected by circadian cycling, it is unknown whether age-related disruption of the circadian clock causes impaired hippocampal memory. Here, we show that the repressive histone deacetylase HDAC3 restricts long-term memory, synaptic plasticity, and experience-induced expression of the circadian gene Per1 in the aging hippocampus without affecting rhythmic circadian activity patterns. We also demonstrate that hippocampal Per1 is critical for long-term memory formation. Together, our data challenge the traditional idea that alterations in the core circadian clock drive circadian-related changes in memory formation and instead argue for a more autonomous role for circadian clock gene function in hippocampal cells to gate the likelihood of long-term memory formation.

Recent exome sequencing studies have implicated polymorphic Brg1-associated factor (BAF) complexes (mammalian SWI/SNF chromatin remodeling complexes) in several human intellectual disabilities and cognitive disorders. However, it is currently unknown how mutations in BAF complexes result in impaired cognitive function. Postmitotic neurons express a neuron-specific assembly, nBAF, characterized by the neuron-specific subunit BAF53b. Mice harboring selective genetic manipulations of BAF53b have severe defects in long-term memory and long-lasting forms of hippocampal synaptic plasticity. We rescued memory impairments in BAF53b mutant mice by reintroducing BAF53b in the adult hippocampus, which suggests a role for BAF53b beyond neuronal development. The defects in BAF53b mutant mice appeared to derive from alterations in gene expression that produce abnormal postsynaptic components, such as spine structure and function, and ultimately lead to deficits in synaptic plasticity. Our results provide new insight into the role of dominant mutations in subunits of BAF complexes in human intellectual and cognitive disorders.

Three-dimensional chromosomal conformations regulate transcription by moving enhancers and regulatory elements into spatial proximity with target genes. Here we describe activity-regulated long-range loopings bypassing up to 0.5 Mb of linear genome to modulate NMDA glutamate receptor GRIN2B expression in human and mouse prefrontal cortex. Distal intronic and 3' intergenic loop formations competed with repressor elements to access promoter-proximal sequences, and facilitated expression via a "cargo" of AP-1 and NRF-1 transcription factors and TALE-based transcriptional activators. Neuronal deletion or overexpression of Kmt2a/Mll1 H3K4- and Kmt1e/Setdb1 H3K9-methyltransferase was associated with higher-order chromatin changes at distal regulatory Grin2b sequences and impairments in working memory. Genetic polymorphisms and isogenic deletions of loop-bound sequences conferred liability for cognitive performance and decreased GRIN2B expression. Dynamic regulation of chromosomal conformations emerges as a novel layer for transcriptional mechanisms impacting neuronal signaling and cognition.