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.
Singh
S K
SK
Department of Medicine, Yale University School of Medicine, New Haven, CT 06510, USA.
Binder
H J
HJ
Geibel
J P
JP
Boron
W F
WF
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