Alanine-stimulated exocytosis in Aplysia enterocytes: effect of Na+ transport and requirement for actin filaments.
We used the Aplysia californica intestinal epithelium to investigate the effect of alanine-stimulated Na+ absorption on apical membrane exocytosis and whether stimulated exocytosis requires intact actin filaments. The fluid-phase marker fluorescein dextran was used to determine rates of apical membrane exocytosis. L-alanine significantly increased apical exocytosis by approximately 30% compared to controls, and there is a modest, positive correlation between alanine-stimulated exocytosis and short-circuit current (ISC). Thus, apical exocytosis is modulated to some extent by the magnitude of Na+ and alanine entry across the apical membrane. Apical exocytosis is also responsive to virtually any increase in Na+ and alanine entry because increments in alanine-stimulated ISC as small as 1 microA/cm2 stimulated exocytosis. We used D-alanine to determine which parameter (sensitivity to transport vs. magnitude of transport) was most important in activation of apical exocytosis. D-alanine-stimulated ISC was one-sixth that of L-alanine, but stimulated exocytosis was only 29% less than that of L-alanine. Therefore, the apical exocytic system is more responsive to small increases in transport than to the magnitude of transport. Latrunculin A (Lat-A) disrupts the actin cytoskeleton and reduced constitutive apical exocytosis by approximately 65% and completely abolished alanine-stimulated exocytosis. Hence, constitutive exocytosis and alanine-stimulated exocytosis require actin filaments for recruitment of vesicles to the apical membrane. During nutrient absorption, actin filament-regulated apical exocytosis may represent a negative feedback system that modulates apical membrane tension.
Keeton
R A
RA
Department of Biology, University of Central Arkansas, Conway, AR 72035-5003, USA.
Runge
S W
SW
Moran
W M
WM
D000199:Actins / Q000378:metabolism; D000409:Alanine / Q000494:pharmacology*; D000818:Animals; D001048:Aplysia / Q000502:physiology*; D003911:Dextrans; D020895:Enterocytes / Q000187:drug effects* / Q000502:physiology*; D005089:Exocytosis / Q000187:drug effects; D005452:Fluoresceins; D005456:Fluorescent Dyes; D017136:Ion Transport / Q000187:drug effects; D012964:Sodium / Q000378:metabolism
D000199:Actins; D003911:Dextrans; D005452:Fluoresceins; D005456:Fluorescent Dyes; C052526:fluorescein-dextran; D012964:Sodium; D000409:Alanine