TY - JOUR
T1 - Intracellular labeling of cat spinal neurons using a tetramethykhodamine-dextran amine conjugate
AU - Carr, P. A.
AU - Noga, B. R.
AU - Nance, D. M.
AU - Jordan, L. M.
N1 - Copyright:
Copyright 2014 Elsevier B.V., All rights reserved.
PY - 1994
Y1 - 1994
N2 - Tetramethylrhodamine-dextran is a highly fluorescent neuroanatomical tracer that, in its 10,000 MW form, has seen widespread use as a sensitive anterograde tract-tracing label. We report here the use of a lower molecular weight tetramethylrhodamine-dextran (3000 MW; Molecular Probes, OR) as an in vivo intracellular marker of locomotor-related spinal neurons. In the paralyzed, decerebrate cat preparation, fictive locomotion was evoked by electrical stimulation of the mesencephalic locomotor region. Extracellular and intracellular potentials of rhythmically active spinal neurons were recorded using microelectrodes filled with 2% tetrarnethybrhodamine-dextran (3000 MW) in 0.9% saline (impedance 5-20 Mohm). Following impalement and electrophysiological characterization, neurons were iontophoretically injected for 2-30 min with 3-10 nA of pulsed positive current. Animals were then perfused 30 min to 7 h postinjection with a variety of paraformaldehyde-and glutaraldehyde-containing fixatives. After tissue sectioning, more than 90% of the injected neurons were recovered. Choline acetyltransferase-immunoreactivity could be demonstrated in a subpopulation of tetramethylrhodamine-dextran-labeled neurons. This technique, in addition to producing high-quality electrodes, has the advantages of rapid yet extensive filling of neuronal processes, no tissue processing prior to visualization, and compatibility with immunohistochemistry.
AB - Tetramethylrhodamine-dextran is a highly fluorescent neuroanatomical tracer that, in its 10,000 MW form, has seen widespread use as a sensitive anterograde tract-tracing label. We report here the use of a lower molecular weight tetramethylrhodamine-dextran (3000 MW; Molecular Probes, OR) as an in vivo intracellular marker of locomotor-related spinal neurons. In the paralyzed, decerebrate cat preparation, fictive locomotion was evoked by electrical stimulation of the mesencephalic locomotor region. Extracellular and intracellular potentials of rhythmically active spinal neurons were recorded using microelectrodes filled with 2% tetrarnethybrhodamine-dextran (3000 MW) in 0.9% saline (impedance 5-20 Mohm). Following impalement and electrophysiological characterization, neurons were iontophoretically injected for 2-30 min with 3-10 nA of pulsed positive current. Animals were then perfused 30 min to 7 h postinjection with a variety of paraformaldehyde-and glutaraldehyde-containing fixatives. After tissue sectioning, more than 90% of the injected neurons were recovered. Choline acetyltransferase-immunoreactivity could be demonstrated in a subpopulation of tetramethylrhodamine-dextran-labeled neurons. This technique, in addition to producing high-quality electrodes, has the advantages of rapid yet extensive filling of neuronal processes, no tissue processing prior to visualization, and compatibility with immunohistochemistry.
KW - Dextran amine conjugate
KW - Fluorescent tracer
KW - Immunohistochemistry
KW - Intracellular staining
KW - Locomotion
KW - Spinal cord
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U2 - 10.1016/0361-9230(94)90017-5
DO - 10.1016/0361-9230(94)90017-5
M3 - Article
C2 - 7521780
AN - SCOPUS:0028333855
VL - 34
SP - 447
EP - 451
JO - Brain Research Bulletin
JF - Brain Research Bulletin
SN - 0361-9230
IS - 5
ER -