Intracellular labeling of cat spinal neurons using a tetramethylrhodamine-dextran amine conjugate.

Abstract:

: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% tetramethylrhodamine-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.

journal_name

Brain Res Bull

journal_title

Brain research bulletin

authors

Carr PA,Noga BR,Nance DM,Jordan LM

doi

10.1016/0361-9230(94)90017-5

subject

Has Abstract

pub_date

1994-01-01 00:00:00

pages

447-51

issue

5

eissn

0361-9230

issn

1873-2747

pii

0361-9230(94)90017-5

journal_volume

34

pub_type

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