The influence of atmospheric NH3 on the apoplastic pH of green leaves: A non-invasive approach with pH-sensitive microelectrodes

Abstract

The apoplastic pH of intact green leaves of Bromus erectus was measured non-invasively by inserting blunt microelectrodes through stomatal openings. After making electrical contact, the recorded signal was stable for hours, yielding a pH of 4.67 ± 0.10. The leaves responded to 'light-off' with an initial transient acidification and subsequent sustained alkalinization of 0.2-0.3 pH; 'light-on' caused the opposite response. Flushing the leaves with 280 nmol NH3 mol-1 air within 18 ± 6 s alkalinized the apoplast by 0.22 ± 0.07 pH, followed by a slower pH increase to reach a steady-state alkalinization of 0.53 ± 0.14 after 19 ± 7 min. This pH shift was persistent as long as the NH3 was flushed, and readily returned to its initial value after replacing the NH3 with clean air. The resultant [NH4/+] increase within the apoplast was measured with a NH4/+-selective microelectrode. In the presence of 280 nmol NH3 mol-1 air, apoplastic NH4/+ initially increased within 15 ± 10 s to 1.53 ± 0.41 mM, to reach a steady state of 1.62 ± 0.16 mM after 27 ± 7 min. An apoplastic buffer capacity of 6 mM pH-1 unit was calculated from the initial changes of pH and [NH4/+], whereas the steady-state values yielded 2.7 mM pH-1. Infiltrated leaves responded to NH4/+ with concentration-dependent depolarizations, the maxima of which yielded saturation kinetics indicating carrier-mediated NH4/+ uptake into adjacent cells, as well as a linear component indicating nonspecific transport. We infer that the initial alkalinization is due to rapid conversion of NH3 to NH4/+, whereas the slower pH increase would be caused by regulatory processes involving both membrane transport, and (mainly) NH4/+ assimilation. Possible consequences of the NH3-induced pH shift for the development of plants growing in polluted areas are discussed.