Nanoinfusion: an integrating tool to study elicitor perception and signal transduction in intact leaves.

•  To study elicitor effects in intact leaves of Hordeum vulgare cv. Ingrid, chitin fragments were delivered to substomatal cavities with a micropipette. Responses were monitored by a calibrated reference microelectrode and a pH-sensitive microelectrode, simply positioned below neighbouring open stomata in the air-filled space of the substomatal cavities. •  Flooding of a leaf spot of approx. 600 × 300 µm with physiological aqueous solutions caused an immediate transient polarization of the extracellular solution in the order of -25 ± 12 mV. Immediately after the pipette solution was consumed, the extracellular solution was again polarized, still before the cavities dried out. In dry cavities, the extracellular equilibrium potential was -34 ± 10 mV. On flooding, the extracellular pH rose to 5.7 ± 0.3 after approx. 12 ± 7 min and returned to a stable level of 5.2 ± 0.3 after 30 min. •  Sequential infusion on the same leaf spot, first with elicitor-free solution, then with the same solution containing 25 µmN-acetyl-chitooctaose, into the still-flooded cavities yielded an elicitor-specific pH maximum between 6 and 7 approx. 10 min after flooding. A pronounced pH maximum > 7 occurred between 40 and 240 min after flooding in wild-type plants. •  The use of elicitor nanoinfusion for the integrated development of resistance inducers in cereals, wine and poplar is discussed.