1. Preincubation of the immature rat uterus under physiological conditions was found to increase its subsequent ability to transport alpha-aminoisobutyric acid, l-proline, l-alanine and 1-aminocyclopentanecarboxylic acid. Uptakes of l-valine, l-phenylalanine and l-leucine were not affected. With alpha-aminoisobutyric acid, a doubling of the uptake could be obtained after a 3-5h preincubation period. Uteri from oestradiol-primed rats gave increases similar to those found in tissues from untreated animals. In both cases the preincubation increased the V(max.) of alpha-aminoisobutyric acid uptake but did not affect the K(m). 2. The conditions during the preincubation period determined the increase in subsequent uptake of alpha-aminoisobutyric acid. No increase in uptake was found if the preincubation was carried out at 1 degrees C, in the presence of cyanide or dinitrophenol, under anaerobiosis or with a concentration of puromycin that depressed incorporation of l-leucine into protein by 95%. The puromycin was also shown to prevent the increase in V(max.) normally found after the preincubation period. In addition, no increase was found if Na(+) was omitted from the preincubation medium. Other inorganic ions had smaller effects. 3. The uptake of alpha-aminoisobutyric acid by uteri before and after a preincubation period showed the same general patterns of sensitivity to competitive inhibitors, K(+), pH, temperature and 2,4-dinitrophenol. 4. The results suggest that the preincubation leads to an increase in a protein component of the ;A system' for amino acid transport in the uterus, and that metabolic energy is required for the reactions involved.
Radioreceptor and high-performance liquid chromatographic (HPLC) assays for nitrendipine were developed and applied to the analysis of serum samples. The HPLC assay required both extraction and concentration of the serum samples, whereas the radioreceptor assay involved only direct dilution of the serum. The HPLC assay, in contrast to the radioreceptor assay, can be used for detection and quantitative analysis of biologically inactive metabolites. The radioreceptor assay is based on the competition between nitrendipine in serum and [3H]nitrendipine for high-affinity binding sites on cardiac membranes. Forty-two serum samples were obtained from five volunteers, and the HPLC and radioreceptor assay results were compared. A correlation coefficient of 0.98 was found between the results of the two assays within the nitrendipine serum concentration range of 1 to 20 ng/ml. No significant levels of active metabolites or any other interferences were found. The radioreceptor assay provides a specific and sensitive alternative to HPLC; it is rapid and inexpensive and with minor modifications may be applicable to most presently available Ca2+ channel antagonists.
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