An apical permeability barrier to NH3/NH4+ in isolated, perfused colonic crypts.
Fermentation of nonabsorbed nutrients in the colon generates high concentrations of NH3/NH4+ in the colonic lumen. NH3 is a small, lipophilic neutral weak base that readily permeates almost all cell membranes, whereas its conjugate weak acid NH4+ generally crosses membranes much more slowly. It is not known how colonocytes maintain intracellular pH in the unusual acid-base environment of the colon, where permeant acid-base products of fermentation exist in high concentration. To address this issue, we hand dissected and perfused single, isolated crypts from rabbit proximal colon, adapting techniques from renal-tubule microperfusion. Crypt perfusion permits control of solutions at the apical (luminal) and basolateral (serosal) surfaces of crypt cells. We assessed apical- vs. basolateral-membrane transport of NH3/NH4+ by using fluorescent dyes and digital imaging to monitor intracellular pH of microvacuolated crypt cells as well as luminal pH. We found that, although the basolateral membranes have normal NH3/NH4+ permeability properties, there is no evidence for transport of either NH3 or NH4+ across the apical borders of these crypt cells. Disaggregating luminal mucus did not increase the transport of NH3/NH4+ across the apical border. We conclude that, compared to the basolateral membrane, the apical border of crypt colonocytes has a very low permeability-area product for NH3/NH4+. This barrier may represent an important adaptation for the survival of crypt cells in the environment of the colon.
Department of Medicine, Yale University School of Medicine, New Haven, CT 06510, USA.
S K
SK
Singh
H J
HJ
Binder
J P
JP
Geibel
W F
WF
Boron
D000042:Absorption; D000641:Ammonia / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport / Q000502:physiology; D002463:Cell Membrane Permeability / Q000187:drug effects / Q000502:physiology*; D016764:Cell Polarity / Q000502:physiology*; D003106:Colon / Q000166:cytology / Q000378:metabolism*; D004210:Dissection; D004229:Dithiothreitol / Q000494:pharmacology; D005260:Female; D005452:Fluoresceins / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007091:Image Processing, Computer-Assisted; D066298:In Vitro Techniques; D010477:Perfusion; D000644:Quaternary Ammonium Compounds / Q000378:metabolism*; D011817:Rabbits
D005452:Fluoresceins; D005456:Fluorescent Dyes; D000644:Quaternary Ammonium Compounds; D000641:Ammonia; C043829:2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein; D004229:Dithiothreitol