Understanding the cell-type composition and spatial organization of brain regions is crucial for interpreting brain computation and function. In the thalamus, the anterior thalamic nuclei (ATN) are involved in a wide variety of functions, yet the cell-type composition of the ATN remains unmapped at a single-cell and spatial resolution. Combining single-cell RNA sequencing, spatial transcriptomics, and multiplexed fluorescent in situ hybridization, we identify three discrete excitatory cell-type clusters that correspond to the known nuclei of the ATN and uncover marker genes, molecular pathways, and putative functions of these cell types. We further illustrate graded spatial variation along the dorsomedial-ventrolateral axis for all individual nuclei of the ATN and additionally demonstrate that the anteroventral nucleus exhibits spatially covarying protein products and long-range inputs. Collectively, our study reveals discrete and continuous cell-type organizational principles of the ATN, which will help to guide and interpret experiments on ATN computation and function.

Mammals taste many compounds yet use a sensory palette consisting of only five basic taste modalities: sweet, bitter, sour, salty and umami (the taste of monosodium glutamate). Although this repertoire may seem modest, it provides animals with critical information about the nature and quality of food. Sour taste detection functions as an important sensory input to warn against the ingestion of acidic (for example, spoiled or unripe) food sources. We have used a combination of bioinformatics, genetic and functional studies to identify PKD2L1, a polycystic-kidney-disease-like ion channel, as a candidate mammalian sour taste sensor. In the tongue, PKD2L1 is expressed in a subset of taste receptor cells distinct from those responsible for sweet, bitter and umami taste. To examine the role of PKD2L1-expressing taste cells in vivo, we engineered mice with targeted genetic ablations of selected populations of taste receptor cells. Animals lacking PKD2L1-expressing cells are completely devoid of taste responses to sour stimuli. Notably, responses to all other tastants remained unaffected, proving that the segregation of taste qualities even extends to ionic stimuli. Our results now establish independent cellular substrates for four of the five basic taste modalities, and support a comprehensive labelled-line mode of taste coding at the periphery. Notably, PKD2L1 is also expressed in specific neurons surrounding the central canal of the spinal cord. Here we demonstrate that these PKD2L1-expressing neurons send projections to the central canal, and selectively trigger action potentials in response to decreases in extracellular pH. We propose that these cells correspond to the long-sought components of the cerebrospinal fluid chemosensory system. Taken together, our results suggest a common basis for acid sensing in disparate physiological settings.

Salt taste in mammals can trigger two divergent behavioural responses. In general, concentrated saline solutions elicit robust behavioural aversion, whereas low concentrations of NaCl are typically attractive, particularly after sodium depletion. Notably, the attractive salt pathway is selectively responsive to sodium and inhibited by amiloride, whereas the aversive one functions as a non-selective detector for a wide range of salts. Because amiloride is a potent inhibitor of the epithelial sodium channel (ENaC), ENaC has been proposed to function as a component of the salt-taste-receptor system. Previously, we showed that four of the five basic taste qualities-sweet, sour, bitter and umami-are mediated by separate taste-receptor cells (TRCs) each tuned to a single taste modality, and wired to elicit stereotypical behavioural responses. Here we show that sodium sensing is also mediated by a dedicated population of TRCs. These taste cells express the epithelial sodium channel ENaC, and mediate behavioural attraction to NaCl. We genetically engineered mice lacking ENaCalpha in TRCs, and produced animals exhibiting a complete loss of salt attraction and sodium taste responses. Together, these studies substantiate independent cellular substrates for all five basic taste qualities, and validate the essential role of ENaC for sodium taste in mice.

Microscopy core facilities are increasingly utilized research resources, but they are generally only available to users within the host institution. Such localized access misses an opportunity to facilitate research across a broader user base. Here, we present the model of an open-access microscopy facility, using the Advanced Imaging Center (AIC) at Howard Hughes Medical Institute Janelia Research Campus as an example. The AIC has pioneered a model whereby advanced microscopy technologies and expertise are made accessible to researchers on a global scale. We detail our experiences in addressing the considerable challenges associated with this model for those who may be interested in launching an open-access imaging facility. Importantly, we focus on how this model can empower researchers, particularly those from resource-constrained settings. This article is protected by copyright. All rights reserved.

DNA sequencing-by-synthesis (SBS) technology, using a polymerase or ligase enzyme as its core biochemistry, has already been incorporated in several second-generation DNA sequencing systems with significant performance. Notwithstanding the substantial success of these SBS platforms, challenges continue to limit the ability to reduce the cost of sequencing a human genome to $100,000 or less. Achieving dramatically reduced cost with enhanced throughput and quality will require the seamless integration of scientific and technological effort across disciplines within biochemistry, chemistry, physics and engineering. The challenges include sample preparation, surface chemistry, fluorescent labels, optimizing the enzyme-substrate system, optics, instrumentation, understanding tradeoffs of throughput versus accuracy, and read-length/phasing limitations. By framing these challenges in a manner accessible to a broad community of scientists and engineers, we hope to solicit input from the broader research community on means of accelerating the advancement of genome sequencing technology.

Chemical fluorophores find wide use in biology to detect and visualize different phenomena. A key advantage of small-molecule dyes is the ability to construct compounds where fluorescence is activated by chemical or biochemical processes. Fluorogenic molecules, in which fluorescence is activated by enzymatic activity, light, or environmental changes, enable advanced bioassays and sophisticated imaging experiments. Here, we detail the collection of fluorophores and highlight both general strategies and unique approaches that are employed to control fluorescence using chemistry.

Bacterial infection of mucosal epithelial cells triggers cell exfoliation to limit the dissemination of infection within the tissue. Therefore, mucosal pathogens must possess strategies to counteract cell extrusion in response to infection. Chlamydia trachomatis spends most of its intracellular development in the non-infectious form. Thus, premature host cell extrusion is detrimental to the pathogen. We demonstrate that C. trachomatis alters the dynamics of focal adhesions. Live-cell microscopy showed that focal adhesions in C. trachomatis-infected cells displayed increased stability. In contrast, focal adhesions in mock-infected cells readily disassembled upon inhibition of myosin II by blebbisttin. Super-resolution microscopy revealed a reorganization of paxillin and FAK in infected cells. Ectopically expressed type III effector TarP localized to focal adhesions, leading to their stabilization and reorganization in a vinculin-dependent manner. Overall, the results indicate that C. trachomatis possesses a dedicated mechanism to regulate host cell focal adhesion dynamics.

Clonal analysis is helping us understand the dynamics of cell replacement in homeostatic adult tissues (Simons and Clevers, 2011). Such an analysis, however, has not yet been achieved for continuously growing adult tissues, but is essential if we wish to understand the architecture of adult organs. The retinas of lower vertebrates grow throughout life from retinal stem cells (RSCs) and retinal progenitor cells (RPCs) at the rim of the retina, called the ciliary marginal zone (CMZ). Here, we show that RSCs reside in a niche at the extreme periphery of the CMZ and divide asymmetrically along a radial (peripheral to central) axis, leaving one daughter in the peripheral RSC niche and the other more central where it becomes an RPC. We also show that RPCs of the CMZ have clonal sizes and compositions that are statistically similar to progenitor cells of the embryonic retina and fit the same stochastic model of proliferation. These results link embryonic and postembryonic cell behaviour, and help to explain the constancy of tissue architecture that has been generated over a lifetime.

The claustrum is a brain region that has been investigated for over 200 years, yet its precise function remains unknown. In the final posthumously released article of Francis Crick, written with Christof Koch, the claustrum was suggested to be critically linked to consciousness. Though the claustrum remained relatively obscure throughout the last half century, it has enjoyed a renewed interest in the last 15 years since Crick and Koch's article. During this time, the claustrum, like many other brain regions, has been studied with the myriad of modern systems neuroscience tools that have been made available by the intersection of genetic and viral technologies. This has uncovered new information about its anatomical connectivity and physiological properties and begun to reveal aspects of its function. From these studies, one clear consensus has emerged which supports Crick and Koch's primary interest in the claustrum: the claustrum has widespread extensive connectivity with the entire cerebral cortex, suggesting a prominent role in 'higher order processes'.

Thermosensation is an indispensable sensory modality. Here, we study temperature coding in Drosophila, and show that temperature is represented by a spatial map of activity in the brain. First, we identify TRP channels that function in the fly antenna to mediate the detection of cold stimuli. Next, we identify the hot-sensing neurons and show that hot and cold antennal receptors project onto distinct, but adjacent glomeruli in the Proximal-Antennal-Protocerebrum (PAP) forming a thermotopic map in the brain. We use two-photon imaging to reveal the functional segregation of hot and cold responses in the PAP, and show that silencing the hot- or cold-sensing neurons produces animals with distinct and discrete deficits in their behavioral responses to thermal stimuli. Together, these results demonstrate that dedicated populations of cells orchestrate behavioral responses to different temperature stimuli, and reveal a labeled-line logic for the coding of temperature information in the brain.