Isolation, visualization, characterization, and osmotic reactivity of crayfish BLMV

Michele G. Wheatly, Jennifer R. Weil, Phyllis B. Douglas

Research output: Contribution to journalArticlepeer-review

11 Scopus citations

Abstract

Procedures were developed to isolate basolateral membrane vesicles (BLMV) from gill, hepatopancreas, and antennal gland of intermolt freshwater crayfish, Procambarus clarkii. Individual procedures involved a discontinuous sucrose gradient (gill), a 65% sucrose cushion (hepatopancreas), or differential centrifugation (antennal gland). BLMV were visualized, characterized (37°C), and tested for osmotic reactivity with a view to using them for Ca2+ uptake studies. Mean diameters of BLMV were 159 nm (gill), 363 nm (hepatopancreas), and 226 nm (antennal gland). Enrichments of basolateral membranes and mitochondria in BLMV were, respectively, 18- and 1.7-fold for gill, 9- and 0.4-fold for hepatopancreas, and 10- and 1-fold for antennal gland. Apical contamination was negligible in BLMV. Percentages of resealing of vesicles as inside out, right side out, or leaky/sheets were 17:27:56% (gill), 14:26:60% (hepatopancreas), and 21:39: 40% (antennal gland). Vesicles exhibited osmotic reactivity, as indicated by a linear relationship between vesicular 45Ca2+ uptake and osmolality. Nonspecific 45Ca2+ binding was 20% in gill, 39% in hepatopancreas, and 31% in antennal gland. Data were compared with published values for marine crustaceans.

Original languageEnglish (US)
Pages (from-to)R725-R734
JournalAmerican Journal of Physiology - Regulatory Integrative and Comparative Physiology
Volume274
Issue number3 43-3
DOIs
StatePublished - Mar 1998
Externally publishedYes

Keywords

  • Antennal gland
  • Basolateral membrane vesicles
  • Freshwater crayfish
  • Gill
  • Hepatopancreas

ASJC Scopus subject areas

  • Physiology
  • Physiology (medical)

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