DOI: 10.1002/jemt.1070240107. Electron microscopy of intracellularly labeled neurons in the hippocampal slice preparation

Electron microscopy of intracellularly labeled neurons in the hippocampal slice preparation

10.1002/jemt.1070240107

Crossref journal-article

Wiley

Microscopy Research and Technique (311)

Abstract

AbstractWe have assessed the properties of three intracellular markers, horseradish peroxidase, biocytin/Neurobiotin, and Lucifer Yellow, and have compared their usefulness as neuronal markers for light and electron microscopic visualization. Neurons in the acute slice preparation of rat hippocampus were filled with one of these markers, and the marker was converted to an optical and electron‐dense reaction product. Dimethylsulfoxide (DMSO) greatly facilitated penetration of recognition reagents while preserving membrane integrity. The markers were compared with respect to injection parameters, mobility and recognition, stability and visibility, and ultrastructural clarity. Horseradish peroxidase (HRP)‐labeled neurons, recognized histochemically with diaminobenzedine (DAB), were easily visualized by the density of the DAB reaction product; however, the electron density was often so great as to obscure ultrastructural details. Biocytin (BC)‐/Neurobiotin (NB)‐labeled neurons were recognized by avidin‐HRP, followed by histochemical localization of HRP with DAB. The optically dense reaction product gave complete visualization of the soma and processes at the light microscopic level. The electron density was homogeneously distributed throughout the cell, so that ultrastructural features were easily identified. Lucifer Yellow (LY), a fluorescent marker, was converted to an optical and electron‐dense reaction product via immunocytochemical staining with a rabbit anti‐LY antibody, followed by goat anti‐rabbit IgG‐HRP and DAB histochemical localization. Similar to BC/NB, the reaction product was evenly dispersed, providing good light microscopic and ultrastructural clarity. Under our experimental conditions, BC/NB and LY were superior markers that could be used routinely to label neurons, and give excellent visualization not only at the light but also at the electron microscopic level. © 1993 Wiley‐Liss, Inc.

Bibliography

Kunkel, D. D., Scharfman, H. E., Schmiege, D. L., & Schwartzkroin, P. A. (1993). Electron microscopy of intracellularly labeled neurons in the hippocampal slice preparation. Microscopy Research and Technique, 24(1), 67â84. Portico.

Authors 4

Dennis D. Kunkel (first)
Helen E. Scharfman (additional)
Donna L. Schmiege (additional)
Philip A. Schwartzkroin (additional)

References 52 Referenced 12

10.1016/0165-0270(90)90006-2
10.1002/cne.902590405
10.1007/978-3-642-87123-8_8
{'key': 'e_1_2_1_5_1', 'volume-title': 'Intracellular Staining in Mammalian Neurones', 'author': 'Brown A. G.', 'year': '1984'} / Intracellular Staining in Mammalian Neurones by Brown A. G. (1984)
10.1016/0306-4522(89)90227-3
10.1007/BF00687074
10.1523/JNEUROSCI.10-06-01892.1990
10.1113/jphysiol.1989.sp017713
10.1523/JNEUROSCI.04-12-03034.1984
10.1016/0304-3940(76)90165-8
10.1016/0165-0270(90)90083-R
10.1016/0306-4522(85)90067-3
10.1085/jgp.59.6.637
10.1016/0165-0270(88)90114-8
10.1016/0006-8993(76)90603-X
{'key': 'e_1_2_1_17_1', 'volume-title': 'Nerve and Muscle Excitation', 'author': 'Junge D.', 'year': '1981'} / Nerve and Muscle Excitation by Junge D. (1981)
10.1152/jn.1989.62.5.1052
10.1523/JNEUROSCI.10-10-03421.1990
10.1016/0006-8993(89)90281-3
10.1007/BF00340492
10.1016/0165-0270(91)90124-I
10.1007/978-1-4613-3189-6_7
10.1016/0006-8993(76)90039-1
10.1007/978-1-4757-2055-6_7
{'key': 'e_1_2_1_26_1', 'first-page': '533', 'volume-title': 'Cytochemical Methods in Neuroanatomy', 'author': 'Kitai S. T.', 'year': '1982'} / Cytochemical Methods in Neuroanatomy by Kitai S. T. (1982)
10.1016/0306-4522(82)90029-X
10.1002/cne.902310203
10.1002/syn.890020405
{'key': 'e_1_2_1_30_1', 'first-page': '285', 'volume-title': 'The Hippocampus‐New Vistas', 'author': 'Lacaille J.‐C.', 'year': '1989'} / The Hippocampus‐New Vistas by Lacaille J.‐C. (1989)
10.1016/0006-8993(74)90154-1
10.1016/0006-8993(80)90413-8
10.1126/science.6266013
10.1126/science.7112109
10.1007/978-3-642-87123-8_1
10.1016/0014-4886(90)90010-P
10.1016/0304-3940(86)90611-7
10.1177/36.5.3356898
10.1016/0306-4522(90)90324-W
10.1016/0304-3940(89)90237-1
10.1016/0306-4522(90)90100-I
{'key': 'e_1_2_1_42_1', 'first-page': '846', 'article-title': 'Antibodies to Lucifer Yellow and Cascade Blue: Light and electron microscopic observations of dye‐filled neurons in rat hippocampus', 'volume': '16', 'author': 'Schmiege D. L.', 'year': '1990', 'journal-title': 'Soc. Neurosci. Absts.'} / Soc. Neurosci. Absts. / Antibodies to Lucifer Yellow and Cascade Blue: Light and electron microscopic observations of dye‐filled neurons in rat hippocampus by Schmiege D. L. (1990)
10.1002/cne.902320206
10.1016/0306-4522(90)90043-4
10.1126/science.54936
10.1007/978-1-4757-2055-6_9
10.1016/0306-4522(86)90122-3
10.1016/0092-8674(78)90256-8
10.1038/292017a0
10.1007/978-3-642-82115-8_7
10.1016/0012-1606(82)90356-6
10.1016/B978-0-12-590350-9.50013-6
10.1016/0006-8993(86)90027-2

Dates

Type	When
Created	20 years, 6 months ago (Feb. 23, 2005, 5:36 p.m.)
Deposited	1 year, 10 months ago (Oct. 25, 2023, 2:36 a.m.)
Indexed	1 year, 10 months ago (Oct. 30, 2023, 6:59 a.m.)
Issued	32 years, 8 months ago (Jan. 1, 1993)
Published	32 years, 8 months ago (Jan. 1, 1993)
Published Online	20 years, 6 months ago (Feb. 4, 2005)
Published Print	32 years, 8 months ago (Jan. 1, 1993)

Funders 0

None

BibTeX

@article{Kunkel_1993, title={Electron microscopy of intracellularly labeled neurons in the hippocampal slice preparation}, volume={24}, ISSN={1097-0029}, url={http://dx.doi.org/10.1002/jemt.1070240107}, DOI={10.1002/jemt.1070240107}, number={1}, journal={Microscopy Research and Technique}, publisher={Wiley}, author={Kunkel, Dennis D. and Scharfman, Helen E. and Schmiege, Donna L. and Schwartzkroin, Philip A.}, year={1993}, month=jan, pages={67–84} }

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