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Related Experiment Videos

Isolation and separation of toad bladder epithelial cells.

M Rossier, B C Rossier, J Pfeiffer

    The Journal of Membrane Biology
    |July 16, 1979
    PubMed
    Summary

    This study isolated five cell types from the toad bladder epithelium, revealing distinct metabolic activities and reaggregation capabilities. These findings advance our understanding of amphibian urinary bladder cell function and differentiation.

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    Area of Science:

    • Cell Biology
    • Amphibian Physiology
    • Epithelial Cell Research

    Background:

    • The urinary bladder epithelium of Bufo marinus comprises five distinct cell types: granular (Gr), mitochondria-rich (MR), goblet (G), microfilament-rich (MFR), and undifferentiated (Un) cells.
    • Understanding the functional and metabolic differences between these cell types is crucial for comprehending bladder physiology.

    Purpose of the Study:

    • To dissociate and fractionate the five cell types of the Bufo marinus urinary bladder epithelium.
    • To characterize the metabolic activity of the isolated cell fractions.
    • To investigate the reaggregation potential of these cells.

    Main Methods:

    • Epithelial dissociation using collagenase and EGTA.
    • Cell fractionation via isopycnic centrifugation in dense serum albumin solutions.

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  • Metabolic assessment through oxygen consumption measurements (in the presence/absence of ouabain, acetazolamide, and dinitrophenol).
  • Cell viability assessment using Trypan blue exclusion.
  • Cell reaggregation studies with calcium supplementation.

    • Main Results:

    • Four distinct cell fractions were obtained, enriched with specific cell types (MR/MFR, Gr, G/Un).
    • Metabolic activity, measured by oxygen consumption, was significantly higher in lighter fractions (MR/MFR-rich) compared to heavier fractions.
    • Ouabain-sensitive oxygen consumption varied by fraction (12-35%), and acetazolamide-sensitive oxygen consumption was higher in lighter fractions (up to 20%).
    • Dinitrophenol consistently increased oxygen consumption across all fractions (20-50%).
    • Isolated cells demonstrated the ability to reaggregate and form junctional complexes when calcium was added.

    Conclusions:

    • Isopycnic centrifugation effectively separated distinct cell populations from the Bufo marinus bladder epithelium.
    • Mitochondria-rich and microfilament-rich cells exhibit higher metabolic rates, suggesting specialized functions.
    • The observed reaggregation capability indicates inherent cellular mechanisms for tissue organization.