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Functional organization of locomotor interneurons in the ventral lumbar spinal cord of the newborn rat.

Antri M, Mellen N, Cazalets JR - PLoS ONE (2011)

Bottom Line: Moreover, L1 segment lesioning induced the most important changes in the locomotor activity in comparison with lesions at the T13 or L2 segments.However, no lesions led to selective disruption of either flexor or extensor output.In addition, this study found no evidence of functional parcellation of locomotor interneurons into flexor and extensor pools at the dorsal-ventral midline of the lumbar spinal cord of the rat.

View Article: PubMed Central - PubMed

Affiliation: Université de Bordeaux, Centre National de la Recherche Scientifique, Institut des Neurosciences Cognitives et Intégratives d'Aquitaine, Unité Mixte de Recherche 5287, Bordeaux, France. myriam.antri@gmail.com

ABSTRACT
Although the mammalian locomotor CPG has been localized to the lumbar spinal cord, the functional-anatomical organization of flexor and extensor interneurons has not been characterized. Here, we tested the hypothesis that flexor and extensor interneuronal networks for walking are physically segregated in the lumbar spinal cord. For this purpose, we performed optical recordings and lesion experiments from a horizontally sectioned lumbar spinal cord isolated from neonate rats. This ventral hemi spinal cord preparation produces well-organized fictive locomotion when superfused with 5-HT/NMDA. The dorsal surface of the preparation was visualized using the Ca(2+) indicator fluo-4 AM, while simultaneously monitoring motor output at ventral roots L2 and L5. Using calcium imaging, we provided a general mapping view of the interneurons that maintained a stable phase relationship with motor output. We showed that the dorsal surface of L1 segment contains a higher density of locomotor rhythmic cells than the other segments. Moreover, L1 segment lesioning induced the most important changes in the locomotor activity in comparison with lesions at the T13 or L2 segments. However, no lesions led to selective disruption of either flexor or extensor output. In addition, this study found no evidence of functional parcellation of locomotor interneurons into flexor and extensor pools at the dorsal-ventral midline of the lumbar spinal cord of the rat.

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Ca2+ imaging experiments.A, Photomicrographs showing Fluo-4 AM labeling in the in vitro ventral SC preparation at the T13-L1 level (A1, magnification ×4; scale bar = 200 µm). Right photomicrographs were taken from areas delineated by grey squares 1 and 2 (A2, A3, magnification ×10; Scale bar = 100 µm). B, Extracellular recordings of left L2 VR and its rectified and low-pass filtered trace illustrating a typical rhythmic activity. Grey bars are aligned with r L2 VR bursts and reveal three cells examples tending to have Ca2+ peaks in phase (cell1), out of phase (cell2) or mixed (cell3) with lL2 VR activity. C, Circular plots illustrating the Ca2+ peak phases (black circles) for each cell in relation to the timing of L2 VRs bursts. The mean VR bursts over a 60 s recording bout range from spans phases 0 to 0.4 in the circular plot and is illustrated in grey. Vectors show the mean phase values and r values. Vector orientation indicates preferred phase of firing; vector length is proportional to coupling strength. D, Dot diagram shows distribution of all cells with phase relationship with motor output from T12 to L5. Cells were illustrated in phase (red dots), out of phase (black dots) and mixed (grey dots). E, Density plot of in phase and out of phase cells from T12 to L5 segments (n = 4 experiments; cc: central canal; scale bar = 100 µm).
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pone-0020529-g002: Ca2+ imaging experiments.A, Photomicrographs showing Fluo-4 AM labeling in the in vitro ventral SC preparation at the T13-L1 level (A1, magnification ×4; scale bar = 200 µm). Right photomicrographs were taken from areas delineated by grey squares 1 and 2 (A2, A3, magnification ×10; Scale bar = 100 µm). B, Extracellular recordings of left L2 VR and its rectified and low-pass filtered trace illustrating a typical rhythmic activity. Grey bars are aligned with r L2 VR bursts and reveal three cells examples tending to have Ca2+ peaks in phase (cell1), out of phase (cell2) or mixed (cell3) with lL2 VR activity. C, Circular plots illustrating the Ca2+ peak phases (black circles) for each cell in relation to the timing of L2 VRs bursts. The mean VR bursts over a 60 s recording bout range from spans phases 0 to 0.4 in the circular plot and is illustrated in grey. Vectors show the mean phase values and r values. Vector orientation indicates preferred phase of firing; vector length is proportional to coupling strength. D, Dot diagram shows distribution of all cells with phase relationship with motor output from T12 to L5. Cells were illustrated in phase (red dots), out of phase (black dots) and mixed (grey dots). E, Density plot of in phase and out of phase cells from T12 to L5 segments (n = 4 experiments; cc: central canal; scale bar = 100 µm).

Mentions: We then performed series of Ca2+ imaging experiments (n = 4). Successive optical recordings were performed to characterize locomotor cells in the region between segments T12 and L5, during fictive locomotion induced by NMDA/5-HT application (Figure 2A). Cells were classified in phase (cell 1, Figure 2B, C; r>0.4), out of phase (cell 2, Figure 2B, C; r>0.4) or mixed cells (cell 3, Figure 2B, C; 0.2>r>0.4).


Functional organization of locomotor interneurons in the ventral lumbar spinal cord of the newborn rat.

Antri M, Mellen N, Cazalets JR - PLoS ONE (2011)

Ca2+ imaging experiments.A, Photomicrographs showing Fluo-4 AM labeling in the in vitro ventral SC preparation at the T13-L1 level (A1, magnification ×4; scale bar = 200 µm). Right photomicrographs were taken from areas delineated by grey squares 1 and 2 (A2, A3, magnification ×10; Scale bar = 100 µm). B, Extracellular recordings of left L2 VR and its rectified and low-pass filtered trace illustrating a typical rhythmic activity. Grey bars are aligned with r L2 VR bursts and reveal three cells examples tending to have Ca2+ peaks in phase (cell1), out of phase (cell2) or mixed (cell3) with lL2 VR activity. C, Circular plots illustrating the Ca2+ peak phases (black circles) for each cell in relation to the timing of L2 VRs bursts. The mean VR bursts over a 60 s recording bout range from spans phases 0 to 0.4 in the circular plot and is illustrated in grey. Vectors show the mean phase values and r values. Vector orientation indicates preferred phase of firing; vector length is proportional to coupling strength. D, Dot diagram shows distribution of all cells with phase relationship with motor output from T12 to L5. Cells were illustrated in phase (red dots), out of phase (black dots) and mixed (grey dots). E, Density plot of in phase and out of phase cells from T12 to L5 segments (n = 4 experiments; cc: central canal; scale bar = 100 µm).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3117791&req=5

pone-0020529-g002: Ca2+ imaging experiments.A, Photomicrographs showing Fluo-4 AM labeling in the in vitro ventral SC preparation at the T13-L1 level (A1, magnification ×4; scale bar = 200 µm). Right photomicrographs were taken from areas delineated by grey squares 1 and 2 (A2, A3, magnification ×10; Scale bar = 100 µm). B, Extracellular recordings of left L2 VR and its rectified and low-pass filtered trace illustrating a typical rhythmic activity. Grey bars are aligned with r L2 VR bursts and reveal three cells examples tending to have Ca2+ peaks in phase (cell1), out of phase (cell2) or mixed (cell3) with lL2 VR activity. C, Circular plots illustrating the Ca2+ peak phases (black circles) for each cell in relation to the timing of L2 VRs bursts. The mean VR bursts over a 60 s recording bout range from spans phases 0 to 0.4 in the circular plot and is illustrated in grey. Vectors show the mean phase values and r values. Vector orientation indicates preferred phase of firing; vector length is proportional to coupling strength. D, Dot diagram shows distribution of all cells with phase relationship with motor output from T12 to L5. Cells were illustrated in phase (red dots), out of phase (black dots) and mixed (grey dots). E, Density plot of in phase and out of phase cells from T12 to L5 segments (n = 4 experiments; cc: central canal; scale bar = 100 µm).
Mentions: We then performed series of Ca2+ imaging experiments (n = 4). Successive optical recordings were performed to characterize locomotor cells in the region between segments T12 and L5, during fictive locomotion induced by NMDA/5-HT application (Figure 2A). Cells were classified in phase (cell 1, Figure 2B, C; r>0.4), out of phase (cell 2, Figure 2B, C; r>0.4) or mixed cells (cell 3, Figure 2B, C; 0.2>r>0.4).

Bottom Line: Moreover, L1 segment lesioning induced the most important changes in the locomotor activity in comparison with lesions at the T13 or L2 segments.However, no lesions led to selective disruption of either flexor or extensor output.In addition, this study found no evidence of functional parcellation of locomotor interneurons into flexor and extensor pools at the dorsal-ventral midline of the lumbar spinal cord of the rat.

View Article: PubMed Central - PubMed

Affiliation: Université de Bordeaux, Centre National de la Recherche Scientifique, Institut des Neurosciences Cognitives et Intégratives d'Aquitaine, Unité Mixte de Recherche 5287, Bordeaux, France. myriam.antri@gmail.com

ABSTRACT
Although the mammalian locomotor CPG has been localized to the lumbar spinal cord, the functional-anatomical organization of flexor and extensor interneurons has not been characterized. Here, we tested the hypothesis that flexor and extensor interneuronal networks for walking are physically segregated in the lumbar spinal cord. For this purpose, we performed optical recordings and lesion experiments from a horizontally sectioned lumbar spinal cord isolated from neonate rats. This ventral hemi spinal cord preparation produces well-organized fictive locomotion when superfused with 5-HT/NMDA. The dorsal surface of the preparation was visualized using the Ca(2+) indicator fluo-4 AM, while simultaneously monitoring motor output at ventral roots L2 and L5. Using calcium imaging, we provided a general mapping view of the interneurons that maintained a stable phase relationship with motor output. We showed that the dorsal surface of L1 segment contains a higher density of locomotor rhythmic cells than the other segments. Moreover, L1 segment lesioning induced the most important changes in the locomotor activity in comparison with lesions at the T13 or L2 segments. However, no lesions led to selective disruption of either flexor or extensor output. In addition, this study found no evidence of functional parcellation of locomotor interneurons into flexor and extensor pools at the dorsal-ventral midline of the lumbar spinal cord of the rat.

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