In situ Turgor Stability in Grape Mesocarp Cells and Its Relation to Cell Dimensions and Microcapillary Tip Size and Geometry

Abstract

The role of three tip fabrication methods and of tip size in cell pressure probe measurements was investigated. The stability over time of turgor pressure (P) in mesocarp cells of intact grape berries was determined for microcapillary tips of contrasting sizes and methods of production (micro-beveler, micro-grinder, and breaking) at contrasting depths below the berry epidermis. SEM images showed that breaking, and in some cases micro-grinding methods produced tips with irregular, fractured surfaces, compared to relatively smooth surfaces produced with micro-beveling. Finely pointed glass microcapillaries were made with reproducibly small tip dimensions (i.d.<5 μm) by using a jet-stream micro-beveler and a mass flow meter to continuously monitor pressurized (pneumatic) air flow from the tip during the beveling process. Optically determined tip i.d. was highly correlated (r²=0.99) to pneumatic tip conductance. Overall cell dimensions ranged from about 10 μm near the epidermis to over 100 μm at a depth of 2 mm. At shallow depths (<250 μm), P was relatively instable (declined at −0.035 MPa/min) when measured using large tips (i.d.>5 μm) and significantly more stable (−0.009 MPa/min) using small tips (i.d.<5 μm). At deeper depths (>250 μm), both tip sizes gave relatively stable P values (−0.007 and −0.004 MPa/min for large and small tips, respectively). For tips of comparable size, micro-beveling gave improved P stability compared to breaking, and an improved ability to measure cells closer to the epidermis compared to micro-grinding. Literature values for acceptability of P values as stable in higher plant cells range from −0.001 to −0.05 MPa/min. These data suggest that P stability might be influenced by both tip size/shape and cell dimensions, and that improved tip fabrication techniques should improve the stability and reproducibility of P measurements, particularly for small cells.