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Organization and function of neural circuits

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Organization and function of neural circuits

We have used genetic and viral strategies to decipher the organizational principles of the fly and mouse olfactory systems (Fig. 3-1), as well as the input–output architecture of norepinephrine, dopamine, and serotonin systems at the scale of the entire mouse brain (Fig. 3-2). We are also combining single-cell transcriptomics, anatomical tracing tools with activity TRAPing (see Section 4), as well as behavioral analyses, to interrogate the functional organization of a variety of neural circuits. 

Some of the recent studies are highlighted below:

*We have shown that dorsal raphe serotonin neurons comprise parallel subsystems with biased inputs from diverse brain regions as well as distinct projection patterns, physiological response properties, and behavioral functions (Fig. 3-3; Movie 3-1).

*We have discovered that cerebellar granule cells, comprising by far the most numerous neurons in the mammalian brain, encode expectation of reward and share dynamics with premotor cortical layer 5 projection neurons in a motor sequence planning task as a result of learning (Movie 3-2).

*By combining single-nucleus RNA-sequencing, spatial transcriptomics, and whole-brain axon mapping, we identified the unit of organization and evolution of cerebellar nuclei,which communicate the results of cerebellar computation to the rest of the brain (Fig. 3-4).

*Combining single-cell sequencing, projection mapping, and in vivo Ca2+ imaging inbehaving mice revealed differential encoding of task variables across transcriptomicallydefined prefrontal cortical projection neuron classes (Fig. 3-5).

*Applying activity TRAPing at different times after fear learning revealed temporalevolution of ensembles of prefrontal cortical neurons that promote remote memoryretrieval (Fig. 4-3).

*TRAPing dehydration-activated neurons in the hypothalamic thirst centers allowed us to examine the relationship between homeostatic need and motivated behavior; our study provided neural basis for a decades-old drive reduction theory of motivation (Fig. 3-6).

Selected publications

Olfactory circuits:

Marin EC, Jefferis GSXE, Komiyama T, Zhu H & Luo L (2002) Representation of the glomerular olfactory map in the Drosophila brain. Cell 109: 243-255; PMID: 12007410.

Jefferis GSXE*, Potter CJ*, Chan AM, Marin EC, Rohlfing T, Maurer CR & Luo L (2007) Comprehensive maps of fly higher olfactory centres: spatially segregated fruit and pheromone representation. Cell 128:1187-1203.

Chou Y-H*, Spletter ML*, Yaksi E*, Leong JCS, Wilson RI# & Luo L# (2010) Diversity and wiring variability of olfactory local interneurons in the Drosophila antennal lobe. Nat Neurosci 13: 439-449; PMID: 20139975; PMCID: PMC2847188.

Miyamichi K, Amat F, Moussavi F, Wang C, WIchersham I, Wall NR, Taniguchi H, Tasic B, Huang ZJ, He Z, Callaway EM, Horowitz MA & Luo L (2011) Cortical representations of olfactory input by trans-synaptic tracing. Nature 472:191-196.

Miyamichi K*, Shlomai-Fuchs Y*, Shu M, Weissbourd BC, Luo L# & Mizrahi A# (2013) Dissecting local circuits: parvalbumin interneurons underlie broad feedback control of olfactory bulb output. Neuron 80:1232-45.

Neuromodulatory circuits:

Weissbourd B, Ren J, DeLoach KE, Guenthner CJ, Miyamichi K & Luo L (2014) Presynaptic partners of dorsal raphe serotonergic and GABAergic neurons. Neuron 83:645-62.

Schwarz LA*, Miyamichi K*, Gao XJ, Beier KT, Weissbourd B, DeLoach KE, Ren J, Ibanes S, Malenka RC, Kremer EJ & Luo L (2015). Viral-genetic tracing of the input-output organization of a central noradrenaline circuit. Nature 524:88-92.

Beier KT, Steinberg EE, DeLoach KE, Xie S, Miyamichi K, Schwarz L, Gao XJ, Kremer EJ, Malenka RC# & Luo L# (2015). Circuit architecture of VTA dopamine neurons revealed by systematic input-output mapping. Cell 162:622-634.

Beier KT, Kim CK, Hoerbelt P, Hung LW, Heifets BD, DeLoach KE, Mosca TJ, Neuner S, Deisseroth K, Luo L♯, Malenka RC♯ (2017) Rabies screen reveals GPe control of cocaine-triggered plasticity. Nature 549:345-350; PMID: 28902833. 

Ren J, Friedmann D, Xiong J, Liu CD, Ferguson B, Weerakkody T, DeLoach K, Ran C, Pun A, Sun Y, Weissbourd B, Neve RL, Huguenard J, Horowitz MA & Luo L (2018) Anatomically defined and functionally distinct dorsal raphe serotonin syb-systems. Cell 175:472-487.

Ren J*, Isakova A*, Friedmann D*, Zeng J*, Grutzner SM, Pun A, Zhao GQ, Kolluru SS, Wang R, Lin R, Li P, Li A, Raymond JL, Luo Q, Luo M, Quake SR#, Luo L# (2019) Single-cell transcriptomes and whole-brain projections of serotonin neurons in the mouse dorsal and median raphe nuclei. eLife 8:e49424.

Hypothalamic circuits:

Allen WE*, DeNardo LA*, Chen MZ*, Liu CD, Loh KM, Fenno LE, Ramakrishnan C, Deisseroth K# & Luo L# (2017) Thirst-associated median preoptic neurons encode an aversive motivational drive. Science 357:1149-1155.

Allen WE, Chen MZ, Pichamoorthy N, Tien RH, Pachitariu M, Luo L#, Deisseroth K#. (2019) Thirst regulates motivated behavior through modulation of brainwide neural population dynamics. Science 364:253.

Cortical-Cerebellar circuits:

DeNardo LA*, Berns DS*, DeLoach K & Luo L (2015) Connectivity of mouse somatosensory and prefrontal cortex examined with trans-synaptic tracing. Nat Neurosci 18:1687-1697.

Wagner MJ*#, Kim TH*, Savall J, Schnitzer MJ# & Luo L# (2017) Cerebellar granule cells encode the expectation of reward. Nature 544:96-100.

DeNardo LA, Liu CD, Allen WE, Adams EL, Friedmann D, Fu L, Guenthner CJ, Tessier-Lavigne M, Luo L (2019) Temporal evolution of cortical ensembles promoting remote memory retrieval. Nat Neurosci 22:460-469.

Wagner MJ*#, Kim TH*, Kadmon J, Nguyen ND, Ganguli S, Schnitzer MJ#, Luo L# (2019) Shared cortex-cerebellum dynamics in the execution and learning of a motor task. Cell 177:669-682.

Kebschull JM, Richman EB, Ringach N, Friedmann D, Albarran E, Kollulu S, Jones RC, Allen WE, Wang Y, Cho SW, Zhou H, Ding JB, Chang HW, Deisseroth K, Quake SR#, Luo L# (2020). Cerebellar nuclei evolved by repeatedly duplicating a conserved cell type set. Science 370:eabd5059.

Lui JH*#, Nguyen ND*, Grutzner SM, Darmanis S, Peixoto D, Wagner MJ, Allen WE, Kebschull JM, Richman EB, Ren J, Newsome WT, Quake SR# & Luo L# (2021). Differential encoding in prefrontal cortex projection neuron classes across cognitive tasks. Cell 184:489-506.

Shuster SA*, Wagner MJ*, Pan-Doh N, Ren J, Grutzner SM, Beier KT, Kim TH, Schnitzer MJ, Luo L (2021) The relationship between birth timing, circuit wiring, and physiological response properties of cerebellar granule cells. Proc Natl Acad Sci USA 118:e2101826118. 

* co-first authors; # co-corresponding authors

Fig. 3-1 – Organization of the fly and mouse olfactory systems. (A) Left, dendrites of three Drosophila olfactory projection neurons (PNs) that target to the DL1, DM5, or VA1v glomerulus in the antennal lobe (AL), respectively. Right, branching patterns of axons of three DL1 PNs (top row), three DM5 PNs (middle row), and three VA1v PNs (bottom row) in the mushroom body (MB) and lateral horn (LH). PNs of the same type exhibit stereotyped axon branching patterns in LH, a high olfactory center mediating innate behavior, but less organized pattern in MB, a high olfactory center mediating learned behavior. After Marin, Jefferis et al. (2002) Cell 109:243-255; see also Jefferis, Potter et al. (2007) Cell 128:1187-1203. (B) Summary of rabies virus–mediated retrograde trans-synaptic tracing on a standard olfactory bulb (OB) model. Green ovals are glomeruli corresponding to mitral cells labeled by rabies virus injection into cortical amygdala (three injections combined); red and magenta ovals are glomeruli corresponding to mitral cells labeled by two separate rabies virus injections into piriform cortex. Cortical amygdala receives biased input from dorsal OB, whereas inputs to piriform cortex are distributed throughout the OB. D, dorsal; V, ventral; A, anterior; P, posterior. Adapted from Miyamichi et al. (2011) Nature 472:191-196.

Fig. 3-2 – Input–output organization of monoamine neuromodulatory systems in the mouse. Monoamine neurons in region B collectively receive inputs from regions A1–Am and send broad output to regions C1–Cn. (A) Biased input–segregated output architecture. This architecture applies to midbrain dopamine neurons and dorsal raphe serotonin neurons. Arrows of different thickness represent different input strengths. (B) Integration-and-broadcast architecture. Neuronal populations in region B that project to a specific output region also send output to other output regions, with the possibility of a quantitative bias; these populations also receive similar inputs. Locus coeruleus norepinephrine neurons approximate this architecture. For more details, see Schwarz, Miyamichi et al. (2015) Nature 524:88-92; Beier et al. (2015) Cell 162:622-634; Ren et al. (2018) Cell 175:472-487.

Fig. 3-3 – Dorsal raphe (DR) serotonin neurons comprise parallel subsystems with distinct anatomy, physiology, and behavioral function. Schematic summary of two DR serotonin subsystems. DR serotonin neurons that project to frontal cortex (blue) are activated by reward, inhibited by punishment, and promote active coping in forced swim test. DR serotonin neurons that project to amygdala (green) are activated by both reward and punishment, and promote anxiety (e.g., as assayed in an elevated plus maze shown here). Adapted from Ren et al. (2018) Cell 175:472-487.

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Movie 3-1 – Fly-through of aligned axonal projections from Trh+ and Vglut3+ serotonin subpopulations in the dorsal raphe. Left, individual slice Z projections at 125 µm depth; right, heatmap of axon density. Serotonin neurons expressing thyrotrophin release hormone (Trh) project axons (cyan) preferentially to subcortical regions, in particular the hypothalamus. Serotonin neurons that express vesicular glutamate transporter 3 (Vglut3) project axons (red) preferentially to the cortex and olfactory bulbs. Axon projections are extracted using TrailMap (see Section 4). For more details, see Ren et al. (2019) eLife 8:e49424.

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 Movie 3-2 – Example dual-site simultaneous two-photon imaging of premotor layer 5 pyramidal neurons (left) and cerebellar granule cells (right) during a forelimb motor sequence planning task. The movie is 4x temporally down-sampled from the 30-Hz acquisition rate. Analysis of imaging data like these revealed that cerebellar granule cells and premotor layer 5 neurons show similar task encoding of movement planning, execution, as well as different aspects of reward in expert mice, and their activities exhibit high correlation. These shared dynamics co-emerge over the course of learning as behavioral performance improves. See Wagner et al. (2019) Cell 117:669-682 for more details. For detailed analysis of reward encoding by cerebellar granule cells, see Wagner et al. (2017) Nature 544:96-100.

Fig. 3-4 – Evolution of the cerebellar nuclei. Comparative single-cell transcriptomics of mouse, chicken, and human (top left; neurons are color-coded by type), spatial transcriptomic analyses in mouse and chicken (top right; neurons are color-coded by type in raw and processed data) and CNS-wide projection mapping in mouse (bottom left; axons in red in a 3D mouse brain) revealed the unit of cerebellar nuclei organization and evolution. This unit (bottom right; red box) comprises three inhibitory and two excitatory neuron classes (each colored circle indicates a neuron class). Extant cerebellar nuclei likely derived from duplication and divergence of this unit, with more divergent gene expression in excitatory neurons (changed color hues), along with projection target shifts. Adapted from Kebschull et al. (2020) Science 370:eabd5059.

Fig. 3-5 – Differential encoding of task variables by prefrontal cortex (PFC) projection neuron classes. Single-cell RNA-sequencing of PFC neurons identified distinct transcriptomic clusters with complex yet stereotyped axon projection specificity. Linking this static, cell type classification data to dynamic in vivo Ca2+ imaging in freely moving mice, comparison of periaqueductal gray–projecting (comprising a single transcriptomic type) and contralateral PFC–projecting (comprising three transcriptomic types) neurons revealed a balance between redundancy and specificity in task signal encoding. Both populations redundantly encode all task signals assayed, yet the former more specifically encodes choice, and the latter more specifically encodes reward context. Adapted from Lui et al. (2021) Cell 184:489-506.

Fig. 3-6 – Activity of dehydration-activated median preoptic nucleus (MnPO) neurons encodes an aversive thirst drive. (A) Experimental setup. ChR2 is selectively expressed in hypothalamic MnPO neurons that were previously activated under a water deprivation condition (thirst-TRAP) or under a homecage condition when mice could drink water ad libitum (homecage-TRAP; see Section 4 for the TRAP method). Mice have been trained to press a lever to obtain water for drinking (B), or to turn off optogenetic stimulation of MnPO neurons (C). (B) Optogenetic activation (within the period highlighted in dark yellow) of thirst-TRAPed MnPO neurons causes intense lever pressing in water-satiated mice. Each reinforcement represents lever presses that produce one unit of water for drinking. (C) In this experiment, photostimulation is constantly on unless the mouse presses the lever, which results in a 20-second break. Each reinforcement represents lever presses that produce a 20-sec break of optogenetic stimulation. After training, mice voluntarily press the lever to turn off optogenetic activation of thirst-TRAP MnPO neurons during the entire 30-minute session, indicating that activation of dehydration-activated MnPO neurons is aversive and mice are motivated to reduce this thirst drive. Homecage-TRAP serves as a negative control in both experiments. Adapted from Allen et al. (2017) Science 357:1149-1155.