Virtual elimination of the interference of unstirred water layers on intestinal sugar transport kinetics by use of the tissue accumulation method at appropriate shaking rates
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The intestinal transport of sugars and amino acids seems to follow Michaelis-Menten kinetics, but the presence of unstirred water layers at the outer face of the brush border membrane may distort kinetic measurements. According to current theory, the capacity parameter,J mc max would not be affected, but theK t would be increased to a higher value,K t ′ , in proportion to the thickness of the unstirred water layer,d.
We reasoned that by increasing the shaking rate in the tissue accumulation method,d might drop to such small values thatK t ′ would fall to a constant level practically equal to the “true”K t .
We measuredd-galactose influx into rings of everted hamster intestine as a function of both the substrate concentration and the shaking rate. Our results show that as the circular stirring rate increases from 0.38–6.2 Hz,J mc max remains constant, as expected, butK t ′ first drops, then levels off to reach a plateau between 2 and 6.2 Hz. We conclude that the averageK t values in this frequency range (K t =7.4 mM) represent the true transportK t . Furthermore, all previous kinetic work performed in our laboratory has been carried out under identical conditions, including shaking rates of 4 Hz. The validity of our preceding results is thus upheld.
Key wordsUnstirred water layers d-galactose transport Transport kinetics Intestine
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- 1.Alvarado F (1976) Sodium-driven transport. A re-evaluation of the sodium-gradient hypothesis. In: Robinson JWL (ed) Intestinal ion transport Medical and Technical Publ. Co. Lancaster, p 117Google Scholar
- 2.Alvarado F, Mahmood A (1974) Cotransport of organic solutes and sodium ions in the small intestine: a general model. Amino acid transport. Biochemistry 13:2882–2890Google Scholar
- 3.Alvarado F, Lherminier M (1979) Unstirred water layers and the evaluation of the kinetic parameters of active transport in the small intestine in vitro. J Physiol (Lond) 289:45PGoogle Scholar
- 4.Christensen HN (1975) Biological Transport, 2nd ed chapter 4. W. A. Benjamin, Reading (Mass.)Google Scholar
- 5.Cleland WW (1967) The statistical analysis of enzyme kinetic data. Adv Enzymol 29:1–32Google Scholar
- 6.Crane RK, Mandelstam P (1960) The active transport of sugars by various preparations of hamster intestine. Biochim Biophys Acta 45:460–476Google Scholar
- 7.Dainty J (1963) Water relations of plant cells. Adv Botan Res 1:279–326Google Scholar
- 8.Dugas MC, Crane RK (1973) Effects of mucosal unstirred layers ond-glucose influx and electrical parameters across in vitro everted hamster jejunum. Fed Proc 32:423Google Scholar
- 9.Dugas MC, Ramaswamy K, Crane RK (1975) An analysis of thed-glucose influx kinetics of in vitro hamster jejunum, based on considerations of the mass-transfer coefficient. Biochim Biophys Acta 382:576–589Google Scholar
- 10.Fisher RB, Gardner MLG (1974) A kinetic approach to the study of absorption of solutes by isolated perfused small intestine. J Physiol (Lond) 241:211–234Google Scholar
- 11.Kotik A, Janacek K (1975) Cell membrane transport, principles and techniques. 2nd ed. Plenum, New YorkGoogle Scholar
- 12.Lherminier M (1979) Etude du contransport de sodium et de solutés organiques par la bordure en brosse intestinale. Thèse de doctorat de specialité, OrsayGoogle Scholar
- 13.Miller DM (1972) The effect of unstirred layers on the measurement of transport rates in individual cells. Biochim Biophys Acta 266:85–90Google Scholar
- 14.Modigliani R, Bernier JJ (1971) Absorption of glucose, sodium and water by the human jejunum studied by intestinal perfusion with a proximal occluding ballon and at variable flow rates. Gut 12:184–193Google Scholar
- 15.Okita GT, Kabara JJ, Richardson F, Le Roy GV (1957) Assaying compounds containing3H and14C. Nucleonics 15: 111–113Google Scholar
- 16.Rey F, Drillet F, Schmitz J, Rey J (1974) Influence of flow rate on the kinetics of the intestinal absorption of glucose and lysine in children. Gastroenterology 66:79–85Google Scholar
- 17.Schultz SG, Curran PF (1970) Coupled transport of sodium and organic solutes. Physiol Rev 50:637–718Google Scholar
- 18.Thomson ABR, Dietschy JM (1977) Derivation of the equations that describe the effects of unstirred water layers on the kinetic parameters of active transport processes in the intestine. J Theor Biol 64:277–294Google Scholar
- 19.Winne D (1973) Unstirred layer, source of biased Michaclis constant in membrane transport. Biochim Biophys Acta 298:27–31Google Scholar
- 20.Winne D (1978) The permeability coefficient of the wall of a villous membrane. J Math Biol 6:95–108Google Scholar
- 21.Winne D, Kopf S, Ulmer M-L (1979) Role of unstirred layer in intestinal absorption of phenylalanine in vivo. Biochim. Biophys Acta 550:120–130Google Scholar