engleski

Incubation in tissue culture media allows isolated rabbit proximal tubules to regain in- vivo-like transport function: response of HCO3- absorption to norepinephrine

Using a new stop-flow perfusion technique with microspectrofluorometric determination of luminal fluid pH, we have studied which substrates or incubation conditions allow isolated rabbit proximal tubules to attain in- vivo-like rates of HCO3- absorption (JHco3) and maximal responses of JHCO3 to norepinephrine (NE). Essentially three incubation media were tested: plasma-like HCO3--Ringer solution containing 5 mmol/l D-glucose (G-Ringer sol.), the same solution also containing 10 mmol/l lactate and 5 mmol/l L-alanine, (LAG-Ringer sol.), and two tissue culture media (DMEM and RPMI 1640). Compared to G-Ringer sol., application of LAG-Ringer sol. in the bath and/or lumen, or application of DMEM or RPMJ 1640 in the bath either slightly in-creased or decreased ~HCO3 with borderline significance. However, RPML 1640 plus 1 mmol/l pyruvate stimulated JHCO3 by 55%. While NE (10-5 mol/l), if applied in G-Ringer sol., had no effect, in the presence of LAG-Ringer sol. it increased JHC03 by =40%, and in the presence of DMEM or RPMJ 1640 it increased JHCO3 by ~100%. This stimulation by NE followed Michaelis—Menten kinetics with an EC50 value of 0.25 I.tmol/l and was probably mediated by cx1-adrenergic receptors. Additional cell pH measurements suggest that NE stimulates the basolateral Na+- HCO3- cotransporter which then becomes susceptible to inhibition by cAMP. We conclude that incubation in tissue culture media allows isolated proximal tubules to maintain a better functional state than the commonly used solutions with unphysiologically high substrate concentrations.

Isolated proximal tubule ; Metabolic substrates ; Norepinephrine ; Tissue culture media ; Tubular bicarbonate absorption

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