REGULATORY VOLUME DECREASE IN CULTURED KIDNEY-CELLS (A6) - ROLE OF AMINO-ACIDS

Volume regulation was studied in A6 epithelia grown on permeable suppo rts by measuring cell thickness (T-c) while simultaneously recording s hort circuit current (I-sc) and transepithelial conductance (G(t)). Lo wering the tonicity of the basolateral solution (pi(b)) from 250 or 21 5 to 140 mOsm/kg elicited a rapid rise in T-c followed by a regulation of the cell volume towards control. This decrease in T-c displays the characteristics of the regulatory volume decrease (RVD). Upon restori ng the isoosmotic conditions, T-c decreased rapidly below its control value. A post RVD regulatory volume increase (RVI) as described for ot her cell types was not observed. The subsequent reduction of the basol ateral osmolality increased T-c to the level recorded at the end of th e first hypoosmotic pulse. Because cell content was not altered during the isoosmotic period the second hypoosmotic challenge was isotonic w ith the cell and did therefore not evoke an RVD. How ever, the cell di d not lose its ability to volume regulate since an RVD could be elicit ed by further reduction of pi(b) from 140 to 100 mOsm/kg. The possibil ity of an involvement of amino acids in the RVD was tested. The amount of amino acids in the cell as well as excreted in the bath was determ ined by amino acid analysis. Millimolar concentrations of threonine, s erine, alanine, glutamate, glycine and aspartate were found in the cel l extract. The cellular amino acid concentration was 28.8 +/- 0.4 mM. The amounts of glycine, aspartate and glutamate excreted from the cell during the hypotonic treatment were significantly larger than in cont rol conditions. The excretion of these amino acids during hypotonicity decreased the cellular amino acid concentration by 8.4 +/- 0.2 mM. Th is quantity cannot completely account for the RVD during the first hyp otonic challenge. The addition of glycine, aspartate and glutamate to the bathing solutions, although used at concentrations higher than int racellularly, did not reduce RVD. On the contrary, this maneuver incre ased the amplitude of the RVD following both hypoosmotic pulses. This result suggests a stimulatory role of the amino acids on the processes responsible for the RVD.