Plasma membranes of guard cells in epidermal peels of Vicia faba and C
ommelina communis can be made accessible to a patch-clamp pipet by rem
oving a small portion (1-3 mu m in diameter) of the guard cell wall us
ing a microbeam of ultraviolet light generated by a nitrogen laser. Us
ing this laser microsurgical technique, we have measured channel activ
ity across plasma membranes of V. faba guard cells in both cell-attach
ed and isolated patch configurations. Measurements made in the inside-
out patch configuration revealed two distinct K+-selective channels. M
ajor advantages of the laser microsurgical technique include the avoid
ance of enzymatic protoplast isolation, the ability to study cell type
s that have been difficult to isolate as protoplasts or for which enzy
matic isolation protocols result in protoplasts not amenable to patch-
clamp studies, the maintenance of positional information in single-cha
nnel measurements, reduced disruption of cell-wall-mediated signaling
pathways, and the ability to investigate intercellular signaling throu
gh studies of cells remaining situated within tissue.