The visual allure of microscopy makes it an intuitively powerful research tool. Intuition, however, can easily obscure or distort the reality of the information contained in an image. Common cognitive biases, combined with institutional pressures that reward positive research results, can quickly skew a microscopy project towards upholding, rather than rigorously challenging, a hypothesis. The impact of these biases on a variety of research topics is well known. What might be less appreciated are the many forms in which bias can permeate a microscopy experiment. Even well-intentioned researchers are susceptible to bias, which must therefore be actively recognized to be mitigated. Importantly, although image quantification has increasingly become an expectation, ostensibly to confront subtle biases, it is not a guarantee against bias and cannot alone shield an experiment from cognitive distortions. Here, we provide illustrative examples of the insidiously pervasive nature of bias in microscopy experiments - from initial experimental design to image acquisition, analysis and data interpretation. We then provide suggestions that can serve as guard rails against bias.

Behavior relies on activity in structured neural circuits that are distributed across the brain, but most experiments probe neurons in a single area at a time. Using multiple Neuropixels probes, we recorded from multi-regional loops connected to the anterior lateral motor cortex (ALM), a circuit node mediating memory-guided directional licking. Neurons encoding sensory stimuli, choices, and actions were distributed across the brain. However, choice coding was concentrated in the ALM and subcortical areas receiving input from the ALM in an ALM-dependent manner. Diverse orofacial movements were encoded in the hindbrain; midbrain; and, to a lesser extent, forebrain. Choice signals were first detected in the ALM and the midbrain, followed by the thalamus and other brain areas. At movement initiation, choice-selective activity collapsed across the brain, followed by new activity patterns driving specific actions. Our experiments provide the foundation for neural circuit models of decision-making and movement initiation.

iBiology Courses provide trainees with just-in-time learning resources to become effective researchers. These courses can help scientists build core research skills, plan their research projects and careers, and learn from scientists with diverse backgrounds.

In September 2023, the two largest bioimaging networks in the Americas, Latin America Bioimaging (LABI) and BioImaging North America (BINA), came together during a 1-week meeting in Mexico. This meeting provided opportunities for participants to interact closely with decision-makers from imaging core facilities across the Americas. The meeting was held in a hybrid format and attended in-person by imaging scientists from across the Americas, including Canada, the United States, Mexico, Colombia, Peru, Argentina, Chile, Brazil and Uruguay. The aims of the meeting were to discuss progress achieved over the past year, to foster networking and collaborative efforts among members of both communities, to bring together key members of the international imaging community to promote the exchange of experience and expertise, to engage with industry partners, and to establish future directions within each individual network, as well as common goals. This meeting report summarises the discussions exchanged, the achievements shared, and the goals set during the LABIxBINA2023: Bioimaging across the Americas meeting.

Fluorescent carbon nanomaterials have broadly useful chemical and photophysical attributes that are conducive to applications in biology. In this review, we focus on materials whose photophysics allow for the use of these materials in biomedical and environmental applications, with emphasis on imaging, biosensing, and cargo delivery. The review focuses primarily on graphitic carbon nanomaterials including graphene and its derivatives, carbon nanotubes, as well as carbon dots and carbon nanohoops. Recent advances in and future prospects of these fields are discussed at depth, and where appropriate, references to reviews pertaining to older literature are provided.

When faced with starvation, the bacterium transforms itself into a dormant cell type called a "spore". Sporulation initiates with an asymmetric division event, which requires the relocation of the core divisome components FtsA and FtsZ, after which the sigma factor σ is exclusively activated in the smaller daughter cell. Compartment-specific activation of σ requires the SpoIIE phosphatase, which displays a biased localization on one side of the asymmetric division septum and associates with the structural protein DivIVA, but the mechanism by which this preferential localization is achieved is unclear. Here, we isolated a variant of DivIVA that indiscriminately activates σ in both daughter cells due to promiscuous localization of SpoIIE, which was corrected by overproduction of FtsA and FtsZ. We propose that the core components of the redeployed cell division machinery drive the asymmetric localization of DivIVA and SpoIIE to trigger the initiation of the sporulation program.

Generalist methods for cellular segmentation have good out-of-the-box performance on a variety of image types. However, existing methods struggle for images that are degraded by noise, blurred or undersampled, all of which are common in microscopy. We focused the development of Cellpose3 on addressing these cases, and here we demonstrate substantial out-of-the-box gains in segmentation and image quality for noisy, blurry or undersampled images. Unlike previous approaches, which train models to restore pixel values, we trained Cellpose3 to output images that are well-segmented by a generalist segmentation model, while maintaining perceptual similarity to the target images. Furthermore, we trained the restoration models on a large, varied collection of datasets, thus ensuring good generalization to user images. We provide these tools as “one-click” buttons inside the graphical interface of Cellpose as well as in the Cellpose API.

In most complex nervous systems there is a clear anatomical separation between the nerve cord, which contains most of the final motor outputs necessary for behaviour, and the brain. In insects, the neck connective is both a physical and information bottleneck connecting the brain and the ventral nerve cord (VNC, spinal cord analogue) and comprises diverse populations of descending (DN), ascending (AN) and sensory ascending neurons, which are crucial for sensorimotor signalling and control.Integrating three separate EM datasets, we now provide a complete connectomic description of the ascending and descending neurons of the female nervous system of Drosophila and compare them with neurons of the male nerve cord. Proofread neuronal reconstructions have been matched across hemispheres, datasets and sexes. Crucially, we have also matched 51% of DN cell types to light level data defining specific driver lines as well as classifying all ascending populations.We use these results to reveal the general architecture, tracts, neuropil innervation and connectivity of neck connective neurons. We observe connected chains of descending and ascending neurons spanning the neck, which may subserve motor sequences. We provide a complete description of sexually dimorphic DN and AN populations, with detailed analysis of circuits implicated in sex-related behaviours, including female ovipositor extrusion (DNp13), male courtship (DNa12/aSP22) and song production (AN hemilineage 08B). Our work represents the first EM-level circuit analyses spanning the entire central nervous system of an adult animal.

Inside the cell, proteins essential for signaling, morphogenesis, and migration navigate complex pathways, typically via vesicular trafficking or microtubule-driven mechanisms 1-3. However, the process by which soluble cytoskeletal monomers maneuver through the cytoplasm’s ever-changing environment to reach their destinations without using these pathways remains unknown. 4-6 Here, we show that actin cytoskeletal treadmilling leads to the formation of a semi-permeable actin-myosin barrier, creating a specialized compartment separated from the rest of the cell body that directs proteins toward the cell edge by advection, diffusion facilitated by fluid flow. Contraction at this barrier generates a molecularly non-specific fluid flow that transports actin, actin-binding proteins, adhesion proteins, and even inert proteins forward. The local curvature of the barrier specifically targets these proteins toward protruding edges of the leading edge, sites of new filament growth, effectively coordinating protein distribution with cellular dynamics. Outside this compartment, diffusion remains the primary mode of protein transport, contrasting sharply with the directed advection within. This discovery reveals a novel protein transport mechanism that redefines the front of the cell as a pseudo-organelle, actively orchestrating protein mobilization for cellular front activities such as protrusion and adhesion. By elucidating a new model of protein dynamics at the cellular front, this work contributes a critical piece to the puzzle of how cells adapt their internal structures for targeted and rapid response to extracellular cues. The findings challenge the current understanding of intracellular transport, suggesting that cells possess highly specialized and previously unrecognized organizational strategies for managing protein distribution efficiently, providing a new framework for understanding the cellular architecture’s role in rapid response and adaptation to environmental changes.

Vision provides animals with detailed information about their surroundings, conveying diverse features such as color, form, and movement across the visual scene. Computing these parallel spatial features requires a large and diverse network of neurons, such that in animals as distant as flies and humans, visual regions comprise half the brain’s volume. These visual brain regions often reveal remarkable structure-function relationships, with neurons organized along spatial maps with shapes that directly relate to their roles in visual processing. To unravel the stunning diversity of a complex visual system, a careful mapping of the neural architecture matched to tools for targeted exploration of that circuitry is essential. Here, we report a new connectome of the right optic lobe from a male Drosophila central nervous system FIB-SEM volume and a comprehensive inventory of the fly’s visual neurons. We developed a computational framework to quantify the anatomy of visual neurons, establishing a basis for interpreting how their shapes relate to spatial vision. By integrating this analysis with connectivity information, neurotransmitter identity, and expert curation, we classified the 53,000 neurons into 727 types, about half of which are systematically described and named for the first time. Finally, we share an extensive collection of split-GAL4 lines matched to our neuron type catalog. Together, this comprehensive set of tools and data unlock new possibilities for systematic investigations of vision in Drosophila, a foundation for a deeper understanding of sensory processing.