Single cell ganglioside catabolism in primary cerebellar neurons and glia

David C. Essaka, Jillian Prendergast, Richard B. Keithley, Ole Hindsgaul, Monica M. Palcic, Ronald Lee Schnaar, Norman J. Dovichi

Research output: Contribution to journalArticle

Abstract

Cell-to-cell heterogeneity in ganglioside catabolism was determined by profiling fluorescent tetramethylrhodamine-labeled GM1 (TMR-GM1) breakdown in individual primary neurons and glia from the rat cerebellum. Cells isolated from 5 to 6 day old rat cerebella were cultured for 7 days, and then incubated for 14 h with TMR-GM1. Intact cells were recovered from cultures by mild proteolysis, paraformaldehyde fixed, and subjected to single cell analysis. Individual cells were captured in a capillary, lysed, and the released single-cell contents analyzed by capillary electrophoresis with quantitative laser-induced fluorescent detection of metabolites. Non-neuronal cells on average took up much more exogenous TMR-GM1 than neuronal cells, and catabolized it more extensively. After 14 h of incubation, non-neuronal cells retained only 14% of the TMR products as GM1 and GM2, compared to >50% for neurons. On average, non-neuronal cells contained 74% of TMR-labeled product as TMR-ceramide, compared to only 42% for neurons. Non-neuronal cells retained seven times as much TMRGM3 (7%) compared to neuronal cells (1%). To confirm the observed single cell metabolomics, we lysed and compared TMR-GM1 catabolic profiles from mixed neuron/glial cell cultures and from cultures depleted of non-neuronal cells by treatment with the antimitotic agent cytosine arabinoside. The lysed culture catabolic profiles were consistent with the average profiles of single neurons and glia. We conclude that the ultrasensitive analytic methods described accurately reflect single cell ganglioside catabolism in different cell populations from the brain.

Original languageEnglish (US)
Pages (from-to)1308-1314
Number of pages7
JournalNeurochemical Research
Volume37
Issue number6
DOIs
StatePublished - Jun 2012

Fingerprint

Gangliosides
Neuroglia
Neurons
Cell culture
Rats
Proteolysis
Antimitotic Agents
Capillary electrophoresis
Ceramides
Cytarabine
Metabolites
Brain
Cells
Cerebellum
tetramethylrhodamine
Lasers
Single-Cell Analysis
Metabolomics
Capillary Electrophoresis

Keywords

  • Capillary electrophoresis
  • Ganglioside
  • Glia
  • Neuron
  • Single cell analysis

ASJC Scopus subject areas

  • Cellular and Molecular Neuroscience
  • Biochemistry

Cite this

Essaka, D. C., Prendergast, J., Keithley, R. B., Hindsgaul, O., Palcic, M. M., Schnaar, R. L., & Dovichi, N. J. (2012). Single cell ganglioside catabolism in primary cerebellar neurons and glia. Neurochemical Research, 37(6), 1308-1314. https://doi.org/10.1007/s11064-012-0733-1

Single cell ganglioside catabolism in primary cerebellar neurons and glia. / Essaka, David C.; Prendergast, Jillian; Keithley, Richard B.; Hindsgaul, Ole; Palcic, Monica M.; Schnaar, Ronald Lee; Dovichi, Norman J.

In: Neurochemical Research, Vol. 37, No. 6, 06.2012, p. 1308-1314.

Research output: Contribution to journalArticle

Essaka, DC, Prendergast, J, Keithley, RB, Hindsgaul, O, Palcic, MM, Schnaar, RL & Dovichi, NJ 2012, 'Single cell ganglioside catabolism in primary cerebellar neurons and glia', Neurochemical Research, vol. 37, no. 6, pp. 1308-1314. https://doi.org/10.1007/s11064-012-0733-1
Essaka DC, Prendergast J, Keithley RB, Hindsgaul O, Palcic MM, Schnaar RL et al. Single cell ganglioside catabolism in primary cerebellar neurons and glia. Neurochemical Research. 2012 Jun;37(6):1308-1314. https://doi.org/10.1007/s11064-012-0733-1
Essaka, David C. ; Prendergast, Jillian ; Keithley, Richard B. ; Hindsgaul, Ole ; Palcic, Monica M. ; Schnaar, Ronald Lee ; Dovichi, Norman J. / Single cell ganglioside catabolism in primary cerebellar neurons and glia. In: Neurochemical Research. 2012 ; Vol. 37, No. 6. pp. 1308-1314.
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