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In Vivo Loose-Patch-Juxtacellular Labeling of Cerebellar Neurons in Mice

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Extracellular Recording Approaches

Part of the book series: Neuromethods ((NM,volume 134))

Abstract

Extracellular recording techniques provide a critical means for measuring neuronal function in vivo. For many experiments, metal electrodes yield spike data that is clean enough for resolving spike waveforms to identify neurons. However, even though some neurons can be distinguished based on their spiking features, it is still not possible to definitively identify most neurons only by their activity. To circumvent this problem, Pinault (J Neurosci Methods 65:113–136, 1996) developed the juxtacellular recording-labeling method to anatomically identify individually recorded neurons. His method utilized glass electrodes to isolate and record single units in vivo. The use of pulled capillary pipettes for recording was key for the success of the method since it allowed him to load the electrode with a tracer that could be delivered across the cell membrane and into the cell. Tracing is achieved by placing the electrode in very close proximity to a cell and then, by delivering pulses of current, pores in the membrane transiently open to allow the tracer to enter. Later studies expanded on the precision of the method by making a loose patch onto the cell before filling it. However, while the juxtacellular method has opened new avenues for relating structure to function at single-cell resolution, the approach remains a challenge to execute because there are several critical steps that are difficult to perform. Here, we provide a step-by-step description for how to perform loose-patch-juxtacellular labeling in vivo in mice. Using the cerebellum as a model system, we outline how to fill Purkinje cells with Neurobiotin. The procedure can be adapted to labeling neurons in any part of the brain, and we discuss its value for unambiguously identifying cells in mutant mice.

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Acknowledgments

This work was supported by funds from Baylor College of Medicine (BCM) and Texas Children’s Hospital. R.V.S. received support from the Bachmann-Strauss Dystonia and Parkinson Foundation, Inc., the Caroline Wiess Law Fund for Research in Molecular Medicine, BCM IDDRC U54HD083092, the National Center for Research Resources C06RR029965, and the National Institutes of Neurological Disorders and Stroke (NINDS) R01NS089664. A.M.B. received support from NINDS F31NS101891, and J.J.W. received support from NINDS F31NS092264. The BCM IDDRC Neuropathology Sub-Core performed a portion of the tissue staining (the BCM IDDRC Neurovisualization Core is supported by U54HD083092). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Center For Research Resources or the National Institutes of Health (NIH).

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Authors and Affiliations

  1. Department of Pathology & Immunology, Jan and Dan Duncan Neurological Research Institute of Texas Children’s Hospital, Houston, TX, USA

    Amanda M. Brown, Joshua J. White, Joy Zhou, Taylor Jefferson, Tao Lin & Roy V. Sillitoe

  2. Department of Neuroscience, Jan and Dan Duncan Neurological Research Institute of Texas Children’s Hospital, Houston, TX, USA

    Amanda M. Brown, Joshua J. White, Joy Zhou, Taylor Jefferson, Tao Lin & Roy V. Sillitoe

  3. Baylor College of Medicine, Jan and Dan Duncan Neurological Research Institute of Texas Children’s Hospital, Houston, TX, USA

    Amanda M. Brown, Joshua J. White, Joy Zhou, Taylor Jefferson, Tao Lin & Roy V. Sillitoe

  4. Program in Developmental Biology, Jan and Dan Duncan Neurological Research Institute of Texas Children’s Hospital, Houston, TX, USA

    Roy V. Sillitoe

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  1. Amanda M. Brown
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  2. Joshua J. White
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  3. Joy Zhou
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  4. Taylor Jefferson
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  5. Tao Lin
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  6. Roy V. Sillitoe
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Correspondence to Roy V. Sillitoe .

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  1. Department of Pathology & Immunology and Neuroscience, Baylor College of Medicine, Houston, Texas, USA

    Roy V. Sillitoe

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Brown, A.M., White, J.J., Zhou, J., Jefferson, T., Lin, T., Sillitoe, R.V. (2018). In Vivo Loose-Patch-Juxtacellular Labeling of Cerebellar Neurons in Mice. In: Sillitoe, R. (eds) Extracellular Recording Approaches. Neuromethods, vol 134. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7549-5_1

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  • DOI: https://doi.org/10.1007/978-1-4939-7549-5_1

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