Axonal Dynamics of Excitatory and Inhibitory Neurons in Somatosensory Cortex

Electrophysiology-delivery of fluorescent viral vectors-and two-photon microscopy were used to demonstrate the rapidity of axonal restructuring of both excitatory and inhibitory neurons in rodent cortical layer II/III following alterations in sensory experience

Plasticity of Horizontal Connections at a Functional Border in Adult Rat Somatosensory Cortex

Horizontal connections in superficial cortical layers integrate information across sensory maps by connecting related functional columns. It has been hypothesized that these connections mediate cortical reorganization via synaptic plasticity. However, it is not known if the horizontal connections from discontinuous cortical regions can undergo plasticity in the adult. Here we located the border between two discontinuous cortical representations in vivo and used either pairing or low-frequency stimulation to induce synaptic plasticity in the horizontal connections surrounding this border in vitro. Individual neurons revealed significant and diverse forms of synaptic plasticity for horizontal connections within a continuous representation and discontinuous representations. Interestingly, both enhancement and depression were observed following both plasticity paradigms. Furthermore, plasticity was not restricted by the border's presence. Depolarization in the absence of synaptic stimulation also produced synaptic plasticity, but with different characteristics. These experiments suggest that plasticity of horizontal connections may mediate functional reorganization

Axons of inhibitory interneurons located within deprived rows following 2 d of whisker plucking.

<p>(A) Left, axons located within deprived rows before plucking. Right, same area after 2 d of plucking. Axons that are stable over the two imaging sessions are color coded blue; those pruned after the first session colored red; and newly sprouted axons shown in yellow. (B–D) Quantification of changes in axonal length for 2 (B), 14 (C), and 30 (D) d of whisker plucking. As in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1000395#pbio-1000395-g008" target="_blank">Figure 8</a>, the data are normalized relative to the highest density of retracted axons for each pair of time points. Retracted (left) and added (right) axons are plotted in 50 micron bins, with the injection site, shown by the asterisk, at coordinate (0,0,0). Distances are expressed in µm from injection site. The average locations of barrel columns for animals in each condition are marked on each graph. Deprived barrel column labels are in white, non-deprived barrel labels are in green.</p

Methods for longitudinal imaging of cortical circuitry.

<p>(A) Timeline of experimental procedures. The barrel cortex is mapped electrophysiologically in order to guide placement of injections of genetically modified virus. After allowing several weeks for expression of genes carried by the virus, images of fluorescent label are taken to establish baseline connectivity. The cortex is reimaged at various time points following initiation of whisker plucking. (B) Surface view of horizontally projecting axons, imaged with 2-photon microscopy. (C) Cortical representation of mouse whiskers, with barrels arranged in rows (A to E) and arcs (1 to 6). (D) Whisker plucking induced remapping of barrel cortex topography in adult animals. The movement of the representation of C row whiskers into the deprived cortex following plucking of D and E rows is plotted as a function of the number of days of plucking. The original C/D row border corresponds to zero on the <i>y</i>-axis. The horizontal line at 300 micrometers indicates the original boundary between the D and E rows. (E) A reconstruction of the axonal plexus of excitatory neurons labeled by AAV-eYFP injection into C3 barrel column, shown in surface view. The axonal reconstruction is superimposed on the whisker map. The virus injection site was at the center of the C3 barrel column.</p

Bouton dynamics of selected axons in superficial cortical layers of whisker barrel columns of deprived and non-deprived animals.

<p>Graphs depict the number of boutons on axons before and after whisker plucking. Each bar represents a single identified axon, imaged before (left set) and the same axons imaged after whisker plucking (right set). Blue: stable boutons, red: eliminated boutons, green: added boutons. (A–C) Excitatory neurons. (A) Bouton turnover in whisker barrel columns in animals that underwent normal experience. (B) Bouton turnover during whisker plucking for axons projecting into non-deprived barrel columns and (C) projecting into deprived barrel columns (D–F) Inhibitory interneurons. (D) Bouton turnover in whisker barrel columns of animals that underwent normal experience. (E) Axonal boutons of neurons located in non-deprived barrel columns before and after whisker plucking. (F) Boutons of neurons located in deprived barrel columns.</p

Axons of inhibitory interneurons located in the non-deprived rows following whisker plucking.

<p>(A) Top, the whisker barrel map. Rectangular box depicts the region of axonal reconstructions shown on the right. Middle, axons located within non-deprived rows before whisker plucking. Right, same area after 2 d of plucking. In the reconstruction, the axons that persisted over both sessions are shown in blue, axons retracted from the first to second imaging session in red, and new axons in yellow. Scale bar  = 50 µm. (B–D) Changes in axonal length for 2 (B), 14 (C), and 30 (D) d of whisker plucking. Left, the distribution of axonal length that was lost between each baseline and post-plucking time point. Right, the distribution of axonal length that was gained between the baseline and post-plucking time points. The data for each pair of time points were obtained by averaging over several mice. Here, the magnitude of the axonal changes for each data pair is normalized with respect to the maximum length of axon that was retracted within any bin. The maximum length of retracted axon in each data pair is therefore 1.0 (in arbitrary units of length), and the length of added axon is measured with respect to that value. The dimensions of the bins are 50 µm×50 µm. The average locations of barrel columns for animals in each condition are marked on each map, with deprived barrel columns indicated in white and non-deprived barrel columns indicated in green.</p

Change in axonal density plotted as a function of days of deprivation.

<p>Expressed as a ratio of axonal length at the indicated duration of deprivation to that measured before deprivation. The green line depicts axons of inhibitory interneurons whose somata are located in deprived barrel columns and are projecting into the non-deprived barrel columns. The yellow line depicts axons of excitatory neurons in the non-deprived barrel columns that are projecting into the deprived barrel columns.</p

Changes in axonal density following varying durations of whisker plucking.

<p>(A–D) For 2 (A), 14 (B), 30 (C), and 60 (D) d. Left column, first imaging session. Right column, second imaging session. The axonal length in each bin was averaged over multiple animals and then normalized. The normalization was performed by dividing all the bins in each baseline condition by the value of the highest density bin maximum in that condition. The bins in the corresponding post-plucking time point were divided by the same number. The maximum mean axonal length for any bin in each baseline graph is therefore 1.0 (expressed in arbitrary units, A.U.), and the length in the corresponding post-plucking graph is expressed relative to that standard. The bins in the histogram are 50 µm×50 µm; the injection site is located at coordinate (0,0,0). Average location of barrel column for animals in each condition is indicated on each graph. White text depicts deprived barrel columns. Green text depicts non-deprived barrel columns.</p