Luminal L-alanine stimulates exocytosis at the K+-conductive apical membrane of Aplysia enterocytes.
In Aplysia intestine, stimulation of Na+ absorption with luminal alanine increases apical membrane K+ conductance (GK,a), which presumably regulates enterocyte volume during stimulated Na+ absorption. However, the mechanism responsible for the sustained increase in plasma membrane K+ conductance is not known for any nutrient-absorbing epithelium. In the present study, we have begun to test the hypothesis that the alanine-induced increase in GK,a in Aplysia enterocytes results from exocytic insertion of K+ channels into the apical membrane. We used the fluid-phase marker horseradish peroxidase to assess the effect of alanine on apical membrane exocytosis and conventional microelectrode techniques to assess the effect of alanine on fractional capacitance of the apical membrane (fCa). Luminal alanine significantly increased apical membrane exocytosis from 1.04 +/- 0.30 to 1.39 +/- 0.38 ng. min-1. cm-2. To measure fCa, we modeled the Aplysia enterocyte as a double resistance-capacitance (RC) electric circuit arranged in series. Several criteria were tested to confirm application of the model to the enterocytes, and all satisfied the model. When added to the luminal surface, alanine significantly increased fCa from 0.27 +/- 0. 02 to 0.33 +/- 0.04 (n = 10) after 4 min. There are two possible explanations for our findings: 1) the increase in exocytosis, which adds membrane to the apical plasma membrane, prevents plasma membrane fracture, and 2) the increase in exocytosis delivers K+ channels to the apical membrane by exocytic insertion. After the alanine-induced depolarization of apical membrane potential (Va), there is a strong correlation (r = 0.96) between repolarization of Va, which reflects the increase in GK,a, and increase in fCa. This correlation supports the exocytic insertion hypothesis for activation of GK,a.
Denton
J
J
Department of Biology, University of Central Arkansas, Conway, Arkansas 72035-0001, USA.
Boahene
D
D
Moran
W M
WM
D000409:Alanine / Q000494:pharmacology*; D000818:Animals; D001048:Aplysia; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D005089:Exocytosis / Q000187:drug effects* / Q000502:physiology; D006735:Horseradish Peroxidase / Q000493:pharmacokinetics; D007413:Intestinal Mucosa / Q000166:cytology / Q000502:physiology*; D007700:Kinetics; D008353:Mannitol / Q000493:pharmacokinetics; D008564:Membrane Potentials / Q000502:physiology; D008954:Models, Biological; D011188:Potassium / Q000378:metabolism; D015221:Potassium Channels / Q000187:drug effects / Q000502:physiology*; D012044:Regression Analysis; D013997:Time Factors
D015221:Potassium Channels; D008353:Mannitol; D006735:Horseradish Peroxidase; D000409:Alanine; D011188:Potassium