The value of an integrated approach for understanding the neocortex by combining functional characterization of single neuron activity with the underlying circuit architecture has been understood since the dawn of modern neuroscience. However, in practice, anatomical connectivity and physiology have been studied mostly separately. Following in the footsteps of previous studies that have combined physiology and anatomy in the same tissue, here we present a unique functional connectomics dataset that contains calcium imaging of an estimated 75,000 neurons from primary visual cortex (VISp) and three higher visual areas (VISrl, VISal and VISlm), that were recorded while a mouse viewed natural movies and parametric stimuli. The functional data were co-registered with electron microscopy (EM) data of the same volume which were automatically segmented, reconstructing more than 200,000 cells (neuronal and non-neuronal) and 524 million synapses. Subsequent proofreading of some neurons in this volume yielded reconstructions that include complete dendritic trees as well the local and inter-areal axonal projections. The largest proofread excitatory axon reached a length of 19 mm and formed 1,893 synapses, while the largest inhibitory axon formed 10,081 synapses. Here we release this dataset as an open access resource to the scientific community including a set of analysis tools that allows easy data access, both programmatically and through a web user interface.

In the preceding section various strategies to interdict postreperfusion inflammatory injury of the myocardium were proposed; effectively the strategies were aimed at specific targets such as stimuli which control cell motility, mechanisms of alteration of cell phenotype, and the induction of cell adhesion and proliferation. It is of interest to see how similar these targets would be if one were to attempt a cell biological approach to vascular injury which results in subintimal hyperplasia. In the latter, cells in the media adopt a phenotype which allows cell migration in the direction of a gradient (presumably chemotactic) which directs them to the subintima which is the site of injury. This motility is associated with the expression in these cells of non-muscle myosin and beta actin, both of which are implicated in the motility of leucocytes. Another similarity between acute inflammation and subintimal hyperplasia relates to the role of cellular adhesion as an important determinant of cell motility. As reviewed above, leucocyte motility involves cell adhesion mediated by a class of molecules termed leucocyte integrins (beta 2 integrins) which vary in their alpha subunits and share CD18 as a common beta subunit (CD11a/CD18 = LFA-1, CD11b/CD18 = Mac-1). The activation and inactivation of these integrins is associated with a high and low affinity state, and motility is effected by the cycling of high and low affinity states as well as by cytoskeleton mediated redistribution of adhesion molecules on the leucocyte membrane. In similar fashion, the migration of medial cells and macrophages to the subintima is associated with specific adhesion to extracellular matrix. This adhesion is mediated by similar classes of integrins containing varying alpha subunits and common beta subunits (in this case beta 1 or beta 3) that have been also shown to undergo activation and inactivation cycles yielding high and low affinity states. The similarities between these integrin mediated adhesion events in leucocytes and in the vascular cells is further emphasised by the fact that leucocytes also contain beta 1 integrins; in fact monocytes and T-lymphocytes express a beta 1 integrin (VLA-4) which supports transmigration out of the vascular space via its interaction with VCAM-1 as an alternative to beta 2 integrin-ICAM-1 adhesion. Substantial evidence suggests that integrin mediated adhesion also functions as a transducer of cell secretion of matrix proteins, growth factors, and cytokines from smooth muscle cells, tissue macrophages, transmigrated leucocytes, and endothelial cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Axon initial segments (AISs) generate action potentials and regulate the polarized distribution of proteins, lipids, and organelles in neurons. While the mechanisms of AIS Na+ and K+ channel clustering are understood, the molecular mechanisms that stabilize the AIS and control neuronal polarity remain obscure. Here, we use proximity biotinylation and mass spectrometry to identify the AIS proteome. We target the biotin-ligase BirA* to the AIS by generating fusion proteins of BirA* with NF186, Ndel1, and Trim46; these chimeras map the molecular organization of AIS intracellular membrane, cytosolic, and microtubule compartments. Our experiments reveal a diverse set of biotinylated proteins not previously reported at the AIS. We show many are located at the AIS, interact with known AIS proteins, and their loss disrupts AIS structure and function. Our results provide conceptual insights and a resource for AIS molecular organization, the mechanisms of AIS stability, and polarized trafficking in neurons.