Deciphering how neural circuits are anatomically organized with regard to input and output is instrumental in understanding how the brain processes information. For example, locus coeruleus norepinephrine (LC-NE) neurons receive input from and send output to broad regions of the brain and spinal cord, and regulate diverse functions including arousal, attention, mood, and sensory gating1–8. However, it is unclear how LC-NE neurons divide up their brain-wide projection patterns and whether different LC-NE neurons receive differential input. Here, we developed a set of viral-genetic tools to quantitatively analyze the input–output relationship of neural circuits, and applied these tools to dissect the LC-NE circuit in mice. Rabies virus-based input mapping indicated that LC-NE neurons receive convergent synaptic input from many regions previously identified as sending axons to the LC and suggested novel presynaptic partners, including cerebellar Purkinje cells. The TRIO (tracing the relationship between input and output) method enables trans-synaptic input tracing from specific subsets of neurons based on their projection and cell type. We found that LC-NE neurons projecting to diverse output regions receive mostly similar input. Projection-based viral labeling revealed extensive output divergence: LC-NE neurons projecting to one output region also project to all brain regions we examined. Thus, the LC-NE circuit overall integrates information from, and broadcasts to, many brain regions, consistent with its primary role in regulating brain states. At the same time, we uncovered several levels of specificity in certain LC-NE sub-circuits. These viral-genetic tools for mapping output architecture and input–output relationship are applicable to other neuronal circuits and organisms. More broadly, our viral-genetic approaches provide an efficient intersectional means to target neuronal populations based on cell type and projection pattern.

The mammalian basal forebrain (BF) has important roles in controlling sleep and wakefulness, but the underlying neural circuit remains poorly understood. We examined the BF circuit by recording and optogenetically perturbing the activity of four genetically defined cell types across sleep-wake cycles and by comprehensively mapping their synaptic connections. Recordings from channelrhodopsin-2 (ChR2)-tagged neurons revealed that three BF cell types, cholinergic, glutamatergic and parvalbumin-positive (PV+) GABAergic neurons, were more active during wakefulness and rapid eye movement (REM) sleep (wake/REM active) than during non-REM (NREM) sleep, and activation of each cell type rapidly induced wakefulness. By contrast, activation of somatostatin-positive (SOM+) GABAergic neurons promoted NREM sleep, although only some of them were NREM active. Synaptically, the wake-promoting neurons were organized hierarchically by glutamatergic→cholinergic→PV+ neuron excitatory connections, and they all received inhibition from SOM+ neurons. Together, these findings reveal the basic organization of the BF circuit for sleep-wake control.

Genomic imprinting results in preferential expression of the paternal, or maternal allele of certain genes. We have performed a genome-wide characterization of imprinting in the mouse embryonic and adult brain. This approach uncovered parent-of-origin allelic effects in over 1300 loci. We identified parental bias in the expression of individual genes and of specific transcript isoforms, with differences between brain regions. Many imprinted genes are expressed in neural systems associated with feeding and motivated behaviors, and parental biases preferentially target genetic pathways governing metabolism and cell adhesion. We observed a preferential maternal contribution to gene expression in the developing brain and a major paternal contribution in the adult brain. Thus, parental expression bias emerges as a major mode of epigenetic regulation in the brain.Parent-of-origin effects influence gene expression and trait inheritance in offspring. Genomic imprinting is a form of epigenetic regulation that results in the preferential expression of the paternally or maternally inherited allele of certain genes (1). Currently, fewer than 100 imprinted genes have been identified, and the evolutionary pressures that underlie imprinting are debated (2,3). Clinical and experimental data suggest roles for imprinting in regulating brain development and function (4). In humans, Prader-Willi Syndrome (PWS) and Angelman Syndrome (AS) result from a deletion of the paternal or maternal copy of 15q11-13, respectively. PWS is associated with hyperphagia, stubbornness and compulsive traits (5), whereas AS is associated with absent speech, happy affect and inappropriate laughter (6). Further, studies of parthenogenetic (PG)-and androgenetic (AG)-chimeras in the mouse have suggested preferential maternal contribution to the development of the cortex, but preferential paternal contribution to the hypothalamus (7,8). Such biased roles have yet to be clearly demonstrated. Moreover, despite tantalizing reports, our understanding of the neural systems governed by imprinted genes, and of the scope and features of imprinted loci expressed in the brain is very limited.6 Address correspondance to Dr. Catherine Dulac: dulac@fas.harvard.edu or Dr. Christopher Gregg: cgregg@mcb.harvard.edu. 7 These authors contributed equally to this study. HHMI Author Manuscript HHMI Author Manuscript HHMI Author ManuscriptImprinting refers to functional differences between the maternal and paternal chromosomes or alleles (9), and is also used more strictly to define complete allele-specific silencing (10). Known imprinted genes have been shown to display all-or-none and biased allelic expression according to the gene and tissue considered (11,12). We report here a genomewide analysis of parental allelic effects involving complete silencing or parental biases in gene expression in the murine embryonic day 15 (E15) brain, and in the adult male and female cortex (medial prefrontal cortex (mPFC)) and hypothalamus (preoptic area (POA)). Together with a compa...