Oscillatory chloride efflux at the pollen tube apex has a role in growth and cell volume regulation and is targeted by inositol 3,4,5,6-tetrakisphosphate

Oscillatory growth of pollen tubes has been correlated with oscillatory influxes of the cations Ca2+, H+, and K+. Using an ion-specific vibrating probe, a new circuit was identified that involves oscillatory efflux of the anion Cl- at the apex and steady influx along the tube starting at 12 mum distal to the tip. This spatial coupling of influx and efflux sites predicts that a vectorial flux of Cl- ion traverses the apical region. The Cl- channel blockers 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) and 5-nitro-2-(3-phenylpropylaminio)benzoic acid completely inhibited tobacco pollen tube growth at 80 and 20 muM, respectively. Cl- channel blockers also induced increases in apical cell volume. The apical 50 mum of untreated pollen tubes had a mean cell volume of 3905+/-75 mum(3). DIDS at 80 muM caused a rapid and lethal cell volume increase to 6206+/-171 mum(3), which is at the point of cell bursting at the apex. DIDS was further demonstrated to disrupt Cl- efflux from the apex, indicating that Cl- flux correlates with pollen tube growth and cell volume status. The signal encoded by inositol 3,4,5,6-tetrakisphosphate [Ins(3,4,5,6)P-4] antagonized pollen tube growth, induced cell volume increases, and disrupted Cl- efflux. [Ins(3,4,5,6)P-4] decreased the mean growth rate by 85%, increased the cell volume to 5997+/-148 muM(3), and disrupted normal Cl- efflux oscillations. These effects were specific for Ins(3,4,5,6)P-4 and were not mimicked by either Ins(1,3,4,5)P-4 or Ins(1,3,4,5,6)P-5. Growth correlation analysis demonstrated that cycles of Cl- efflux were coupled to and temporally in phase with cycles of growth. A role for Cl- flux in the dynamic cellular events during growth is assessed. Differential interference contrast microscopy and kymographic analysis of individual growth cycles revealed that vesicles can advance transiently to within 2 to 4 mum of the apex during the phase of maximally increasing Cl- efflux, which temporally overlaps the phase of cell elongation during the growth cycle. In summary, these investigations indicate that Cl- ion dynamics are an important component in the network of events that regulate pollen tube homeostasis and growth.