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 non-specific 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.