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Distal astrocytic processes have a complex morphology, reminiscent of branchlets and leaflets. Astrocytic branchlets are rod-like processes containing mitochondria and endoplasmic reticulum, capable of generating inositol-3-phosphate (IP3)-dependent Ca2+ signals. Leaflets are small and flat processes that protrude from branchlets and fill the space between synapses. Here we use three-dimensional (3D) reconstructions from serial section electron microscopy (EM) of rat CA1 hippocampal neuropil to determine the astrocytic coverage of dendritic spines, shafts and axonal boutons. The distance to the maximum of the astrocyte volume fraction (VF) correlated with the size of the spine when calculated from the center of mass of the postsynaptic density (PSD) or from the edge of the PSD, but not from the spine surface. This suggests that the astrocytic coverage of small and larger spines is similar in hippocampal neuropil. Diffusion simulations showed that such synaptic microenvironment favors glutamate spillover and extrasynaptic receptor activation at smaller spines. We used complexity and entropy measures to characterize astrocytic branchlets and leaflets. The 2D projections of astrocytic branchlets had smaller spatial complexity and entropy than leaflets, consistent with the higher structural complexity and less organized distribution of leaflets. The VF of astrocytic leaflets was highest around dendritic spines, lower around axonal boutons and lowest around dendritic shafts. In contrast, the VF of astrocytic branchlets was similarly low around these three neuronal compartments. Taken together, these results suggest that astrocytic leaflets preferentially contact synapses as opposed to the dendritic shaft, an arrangement that might favor neurotransmitter spillover and extrasynaptic receptor activation along dendritic shafts.

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a) (b) (b) Figure 1: a) bounding boxes fitting; b) empty space skipping and early ray termination; c) tri-linear (left) & tri-cubic (right) interpolation modes side-by-side comparison (MANIX test dataset).

We describe some observations on the practical implementation of the median cut color quantization algorithm, suitably modified for accurate color rendering. The RGB color space is successively divided in such a way that colors with visual significance, even if relatively small in population, are given representatives in the colormap. Appropriately modified, median cut quantization is nearly as good as our best octree quantization. As with octree quantization, error-diffusion dithering is useful for reducing posterization in large regions with a slow variation in color.

The brain extracellular matrix (ECM) is a proteoglycan complex that occupies the extracellular space between brain cells and regulates brain development, brain wiring, and synaptic plasticity. However, the action of the ECM on synaptic plasticity remains controversial. Here, we employed serial section electron microscopy to show that enzymatic attenuation of ECM with chondroitinase ABC (ChABC) triggers the appearance of new glutamatergic synapses onto thin dendritic spines of CA1 pyramidal neurons. The appearance of new synapses increased the ratio of the field excitatory postsynaptic potential (fEPSP) to presynaptic fiber volley (PrV), suggesting that these new synapses are formed on existing axonal fibers. However, both the mean miniature excitatory postsynaptic current (mEPSC) amplitude and AMPA/NMDA ratio were decreased, suggesting that ECM attenuation increased the proportion of ‘unpotentiated’ synapses. A higher proportion of unpotentiated synapses would be expected to promote long-term potentiation (LTP). Surprisingly, theta-burst induced LTP was suppressed by ChABC treatment. The suppression of LTP was accompanied by decreased excitability of CA1 pyramidal neurons due to the upregulation of small conductance Ca2+-activated K+ (SK) channels. A pharmacological blockade of SK channels restored cell excitability and, expectedly, enhanced LTP above the level of control. This enhancement of LTP was abolished by a blockade of Rho-associated protein kinase (ROCK), which is involved in the maturation of dendritic spines. Thus, ECM attenuation enables the appearance of new synapses in the hippocampus, which is compensated for by a reduction in the excitability of postsynaptic neurons, thereby preventing network overexcitation at the expense of synaptic plasticity.

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