Ultra-high density electrodes improve detection, yield, and cell type identification in neuronal recordings.

To understand the neural basis of behavior, it is essential to sensitively and accurately measure neural activity at single neuron and single spike resolution. Extracellular electrophysiology delivers this, but it has biases in the neurons it detects and it imperfectly resolves their action potentials. To minimize these limitations, we developed a silicon probe with much smaller and denser recording sites than previous designs, called Neuropixels Ultra (NP Ultra). This device samples neuronal activity at ultra-high spatial density (~10 times higher than previous probes) with low noise levels, while trading off recording span. NP Ultra is effectively an implantable voltage-sensing camera that captures a planar image of a neuron's electrical field. We use a spike sorting algorithm optimized for these probes to demonstrate that the yield of visually-responsive neurons in recordings from mouse visual cortex improves up to ~3-fold. We show that NP Ultra can record from small neuronal structures including axons and dendrites. Recordings across multiple brain regions and four species revealed a subset of extracellular action potentials with unexpectedly small spatial spread and axon-like features. We share a large-scale dataset of these brain-wide recordings in mice as a resource for studies of neuronal biophysics. Finally, using ground-truth identification of three major inhibitory cortical cell types, we found that these cell types were discriminable with approximately 75% success, a significant improvement over lower-resolution recordings. NP Ultra improves spike sorting performance, detection of subcellular compartments, and cell type classification to enable more powerful dissection of neural circuit activity during behavior.

A multifaceted architectural framework of the mouse claustrum complex.

Accurate anatomical characterizations are necessary to investigate neural circuitry on a fine scale, but for the rodent claustrum complex (CLCX), this has yet to be fully accomplished. The CLCX is generally considered to comprise two major subdivisions, the claustrum (CL) and the dorsal endopiriform nucleus (DEn), but regional boundaries to these areas are debated. To address this, we conducted a multifaceted analysis of fiber- and cytoarchitecture, genetic marker expression, and connectivity using mice of both sexes, to create a comprehensive guide for identifying and delineating borders to CLCX, including an online reference atlas. Our data indicated four distinct subregions within CLCX, subdividing both CL and DEn into two. Additionally, we conducted brain-wide tracing of inputs to CLCX using a transgenic mouse line. Immunohistochemical staining against myelin basic protein (MBP), parvalbumin (PV), and calbindin (CB) revealed intricate fiber-architectural patterns enabling precise delineations of CLCX and its subregions. Myelinated fibers were abundant dorsally in CL but absent ventrally, whereas PV expressing fibers occupied the entire CL. CB staining revealed a central gap within CL, also visible anterior to the striatum. The Nr2f2, Npsr1, and Cplx3 genes expressed specifically within different subregions of the CLCX, and Rprm helped delineate the CL-insular border. Furthermore, cells in CL projecting to the retrosplenial cortex were located within the myelin sparse area. By combining own experimental data with digitally available datasets of gene expression and input connectivity, we could demonstrate that the proposed delineation scheme allows anchoring of datasets from different origins to a common reference framework.

Mice are a highly tractable model for studying the claustrum complex (CLCX). However, without a consensus on how to delineate the CLCX in rodents, comparing results between studies is challenging. It is therefore important to expand our anatomical knowledge of the CLCX, to match the level of detail needed to study its functional properties. To improve and expand upon preexisting delineation schemes, we used the combinatorial expression of several markers to create a comprehensive guide to delineate the CLCX and its subregions, including an online reference atlas. This anatomical framework will allow researchers to anchor future experimental data into a common reference space. We demonstrated the power of this new structural framework by combining our own experimental data with digitally available data on gene expression and input connectivity of the CLCX.

Temporal origin of mouse claustrum and development of its cortical projections.

The claustrum is known for its extensive connectivity with many other forebrain regions, but its elongated shape and deep location have made further study difficult. We have sought to understand when mouse claustrum neurons are born, where they are located in developing brains, and when they develop their widespread connections to the cortex. We established that a well-characterized parvalbumin plexus, which identifies the claustrum in adults, is only present from postnatal day (P) 21. A myeloarchitectonic outline of the claustrum can be derived from a triangular fiber arrangement from P15. A dense patch of Nurr1+ cells is present at its core and is already evident at birth. Bromodeoxyuridine birth dating of forebrain progenitors reveals that the majority of claustrum neurons are born during a narrow time window centered on embryonic day 12.5, which is later than the adjacent subplate and endopiriform nucleus. Retrograde tracing revealed that claustrum projections to anterior cingulate (ACA) and retrosplenial cortex (RSP) follow distinct developmental trajectories. Claustrum-ACA connectivity matures rapidly and reaches adult-like innervation density by P10, whereas claustrum-RSP innervation emerges later over a protracted time window. This work establishes the timeline of claustrum development and provides a framework for understanding how the claustrum is built and develops its unique connectivity.

Human lesions and animal studies link the claustrum to perception, salience, sleep and pain.

The claustrum is the most densely interconnected region in the human brain. Despite the accumulating data from clinical and experimental studies, the functional role of the claustrum remains unknown. Here, we systematically review claustrum lesion studies and discuss their functional implications. Claustral lesions are associated with an array of signs and symptoms, including changes in cognitive, perceptual and motor abilities; electrical activity; mental state; and sleep. The wide range of symptoms observed following claustral lesions do not provide compelling evidence to support prominent current theories of claustrum function such as multisensory integration or salience computation. Conversely, the lesions studies support the hypothesis that the claustrum regulates cortical excitability. We argue that the claustrum is connected to, or part of, multiple brain networks that perform both fundamental and higher cognitive functions. As a multifunctional node in numerous networks, this may explain the manifold effects of claustrum damage on brain and behaviour.

In search of common developmental and evolutionary origin of the claustrum and subplate.

The human claustrum, a major hub of widespread neocortical connections, is a thin, bilateral sheet of gray matter located between the insular cortex and the striatum. The subplate is a largely transient cortical structure that contains some of the earliest generated neurons of the cerebral cortex and has important developmental functions to establish intra- and extracortical connections. In human and macaque some subplate cells undergo regulated cell death, but some remain as interstitial white matter cells. In mouse and rat brains a compact layer is formed, Layer 6b, and it remains underneath the cortex, adjacent to the white matter. Whether Layer 6b in rodents is homologous to primate subplate or interstitial white matter cells is still debated. Gene expression patterns, such as those of Nurr1/Nr4a2, have suggested that the rodent subplate and the persistent subplate cells in Layer 6b and the claustrum might have similar origins. Moreover, the birthdates of the claustrum and Layer 6b are similarly precocious in mice. These observations prompted our speculations on the common developmental and evolutionary origin of the claustrum and the subplate. Here we systematically compare the currently available data on cytoarchitecture, evolutionary origin, gene expression, cell types, birthdates, neurogenesis, lineage and migration, circuit connectivity, and cell death of the neurons that contribute to the claustrum and subplate. Based on their similarities and differences we propose a partially common early evolutionary origin of the cells that become claustrum and subplate, a likely scenario that is shared in these cell populations across all amniotes.

Myosin-independent cytokinesis in Giardia utilizes flagella to coordinate force generation and direct membrane trafficking.

Devoid of all known canonical actin-binding proteins, the prevalent parasite Giardia lamblia uses an alternative mechanism for cytokinesis. Unique aspects of this mechanism can potentially be leveraged for therapeutic development. Here, live-cell imaging methods were developed for Giardia to establish division kinetics and the core division machinery. Surprisingly, Giardia cytokinesis occurred with a median time that is ∼60 times faster than mammalian cells. In contrast to cells that use a contractile ring, actin was not concentrated in the furrow and was not directly required for furrow progression. Live-cell imaging and morpholino depletion of axonemal Paralyzed Flagella 16 indicated that flagella-based forces initiated daughter cell separation and provided a source for membrane tension. Inhibition of membrane partitioning blocked furrow progression, indicating a requirement for membrane trafficking to support furrow advancement. Rab11 was found to load onto the intracytoplasmic axonemes late in mitosis and to accumulate near the ends of nascent axonemes. These developing axonemes were positioned to coordinate trafficking into the furrow and mark the center of the cell in lieu of a midbody/phragmoplast. We show that flagella motility, Rab11, and actin coordination are necessary for proper abscission. Organisms representing three of the five eukaryotic supergroups lack myosin II of the actomyosin contractile ring. These results support an emerging view that flagella play a central role in cell division among protists that lack myosin II and additionally implicate the broad use of membrane tension as a mechanism to drive abscission.