pH regulation in anoxic rice coleoptiles at pH 3.5: biochemical pHstats and net H(+) influx in the absence and presence of NO(3)(−)

During anoxia, cytoplasmic pH regulation is crucial. Mechanisms of pH regulation were studied in the coleoptile of rice exposed to anoxia and pH 3.5, resulting in H(+) influx. Germinating rice seedlings survived a combination of anoxia and exposure to pH 3.5 for at least 4 d, although development was retarded and net K(+) efflux was continuous. Further experiments used excised coleoptile tips (7–10 mm) in anoxia at pH 6.5 or 3.5, either without or with 0.2 mM NO(3)(−), which distinguished two processes involved in pH regulation. Net H(+) influx (μmol g(−1) fresh weight h(−1)) for coleoptiles with NO(3)(−) was ∼1.55 over the first 24 h, being about twice that in the absence of NO(3)(−), but then decreased to 0.5–0.9 as net NO(3)(−) uptake declined from ∼1.3 to 0.5, indicating reduced uptake via H(+)–NO(3)(−) symports. NO(3)(−) reduction presumably functioned as a biochemical pHstat. A second biochemical pHstat consisted of malate and succinate, and their concentrations decreased substantially with time after exposure to pH 3.5. In anoxic coleoptiles, K(+) balancing the organic anions was effluxed to the medium as organic anions declined, and this efflux rate was independent of NO(3)(−) supply. Thus, biochemical pHstats and reduced net H(+) influx across the plasma membrane are important features contributing to pH regulation in anoxia-tolerant rice coleoptiles at pH 3.5.