Tomographic optical imaging of cortical responses after crossing nerve transfer in mice

<div><p>To understand the neural mechanisms underlying the therapeutic effects of crossing nerve transfer for brachial plexus injuries in human patients, we investigated the cortical responses after crossing nerve transfer in mice using conventional and tomographic optical imaging. The distal cut ends of the left median and ulnar nerves were connected to the central cut ends of the right median and ulnar nerves with a sciatic nerve graft at 8 weeks of age. Eight weeks after the operation, the responses in the primary somatosensory cortex (S1) elicited by vibratory stimulation applied to the left forepaw were visualized based on activity-dependent flavoprotein fluorescence changes. In untreated mice, the cortical responses to left forepaw stimulation were mainly observed in the right S1. In mice with nerve crossing transfer, cortical responses to left forepaw stimulation were observed in the left S1 together with clear cortical responses in the right S1. We expected that the right S1 responses in the untreated mice were produced by thalamic inputs to layer IV, whereas those in the operated mice were mediated by callosal inputs from the left S1 to layer II/III of the right S1. To confirm this hypothesis, we performed tomographic imaging of flavoprotein fluorescence responses by macroconfocal microscopy. Flavoprotein fluorescence responses in layer IV were dominant compared to those in layer II/III in untreated mice. In contrast, responses in layer II/III were dominant compared to those in layer IV in operated mice. The peak latency of the cortical responses in the operated mice was longer than that in the untreated mice. These results confirmed our expectation that drastic reorganization in the cortical circuits was induced after crossing nerve transfer in mice.</p></div

Conventional imaging of cortical responses after crossing nerve transfer.

<p>(A) Schematic drawing of crossing nerve transfer. IM represents imaged area. (B) Original fluorescence image. (C) Pseudo-color image of fluorescence responses elicited by vibratory stimulation at 50 Hz for 0.5 s applied to the left forepaw. The image in (B) and the responses in (C) were recorded in the same mouse with crossing nerve transfer. (D) Fluorescence responses elicited by stimulation of the left forepaw in an untreated mouse. The contralateral right S1 shows clear responses, while the ipsilateral left S1 is only weakly activated. (E) Time course of ΔF/F₀ changes in the square windows (1–4) shown in (C) and (D). (F) Amplitudes of the cortical responses. Mean and S.E.M. are shown. (G) Bilaterality index (ratio of the inferior response amplitudes normalized by the superior response amplitudes) in the untreated and operated mice.</p

Tomographic imaging of cortical responses in operated mice.

<p>(A) Original tomographic images in the right S1 (upper panels) and pseudocolor images of fluorescence responses elicited by vibratory stimulation at 50 Hz for 0.5 s applied to the left forepaw (lower panels). (B) Time courses of the fluorescence responses measured at 50, 200, 400 and 800 μm deep from the cortical surface using a macroconfocal microscope. Data in (A) and (B) were obtained from the same mouse. (C) Amplitudes of fluorescence responses measured at each depth.</p

Comparison of cortical responses between untreated and operated mice.

<p>(A) Time courses of the fluorescence responses measured at 50, 200, 400 and 800 μm deep from the cortical surface using a macroconfocal microscope. Statistical differences were evaluated regarding the peak amplitude and peak latency between the untreated and operated mice. (B) Relative response amplitudes at 200 μm normalized to those at 400 μm. This ratio is smaller than 1.0 in the untreated mice, while it was larger than 1.0 in the operated mice. (C) Schematic drawing of the neural circuits in the operated mice.</p

Tomographic imaging of cortical responses in untreated mice.

<p>(A) Original tomographic images in the right S1 (upper panels) and pseudo-color images of fluorescence responses elicited by vibratory stimulation at 50 Hz for 0.5 s applied to the left forepaw (lower panels). The numbers in the upper panels represent the depth (μm) from the cortical surface. The two arrows in the upper panels show the position of an artery that is visible in the leftmost panel but not in the rightmost panel. The circle in the leftmost lower panel shows the circular window in which response amplitudes were measured in ΔF/F₀. (B) Time courses of the fluorescence responses measured at 50, 200, 400 and 800 μm deep from the cortical surface using a macroconfocal microscope. Data in (A) and (B) were obtained from the same mouse. (C) Amplitudes of fluorescence responses measured at each depth.</p

Effects of Preterm Birth on Intrinsic Fluctuations in Neonatal Cerebral Activity Examined Using Optical Imaging

<div><p>Medical advancements in neonatology have significantly increased the number of high-risk preterm survivors. However, recent long-term follow-up studies have suggested that preterm infants are at risk for behavioral, educational, and emotional problems. Although clear relationships have been demonstrated between preterm infants and developmental problems during childhood and adolescence, less is known about the early indications of these problems. Recently, numerous studies on resting-state functional connectivity (RSFC) have demonstrated temporal correlations of activity between spatially remote cortical regions not only in healthy adults but also in neuropathological disorders and early childhood development. In order to compare RSFC of the cerebral cortex between preterm infants at term-equivalent ages and full-term neonates without any anatomical abnormality risk during natural sleep, we used an optical topography system, which is a recently developed extension of near-infrared spectroscopy. We clarified the presence of RSFC in both preterm infants and full-term neonates and showed differences between these groups. The principal differences were that on comparison of RSFC between the bilateral temporal regions, and bilateral parietal regions, RSFC was enhanced in preterm infants compared with full-term neonates; whereas on comparison of RSFC between the left temporal and left parietal regions, RSFC was enhanced in full-term neonates compared with preterm infants. We also demonstrated a difference between the groups in developmental changes of RSFC related to postmenstrual age. Most importantly, these findings suggested that preterm infants and full-term neonates follow different developmental trajectories during the perinatal period because of differences in perinatal experiences and physiological and structural development.</p></div

Demographic Data for infants.

<p>GA: gestational age, CA: chronological age, and PMA: postmenstrual age.</p

Arrangement of measurement positions.

<p>The lowest lines of the probes correspond to the T3-Fp1-Fp2-T4 line according to International 10–20 electrode system. The vertical midline of the channels was centered in the nasion–inion line, and the channels between the 5th and 6th probe in the lowest line corresponded to T3 on the left and T4 on the right.</p

Developmental changes of functional connections.

<p>Each line showed correlation coefficients (<i>r</i> values) between <i>z</i> (<i>r</i>) values in measurement channels for each channel and PMA at the time of the scan in the preterm (A) and full-term groups (B). The displayed results fulfilled the criteria of significance (<i>p</i><0.005, uncorrected). Detailed results of the relationship between each line are shown in Supplementary <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0067432#pone.0067432.s003" target="_blank">Figures S3</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0067432#pone.0067432.s004" target="_blank">S4</a>.</p

Significant differences in functional connectivity between preterm infants at term-equivalent ages and full-term neonates.

<p>The lines represent <i>z</i> values >3.29 (corresponding to <i>p</i><0.0005, uncorrected). The red lines show the connections significantly higher in preterm infants. The blue lines show the connections significantly higher in full-term neonates.</p