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We have compared the effects of the auxin, indole-3-acetic acid (IAA) with that of other weak acids on the plasma-membrane potential of oat (Avena sativa L.) coleoptile cells. Cells treated with 1 μM IAA at pH 6 depolarize 20-25 mV in 10-12 min, but they then repolarize, until by 20-25 min their potentials are about 25 mV more negative than the initial value. Similar concentrations of benzoic and butyric acids cause the initial depolarization, but not the subsequent hyperpolarization. The hyperpolarization is therefore specific to IAA. All the weak acids, including IAA, evoke a rapid hyperpolarization when their concentrations are raised to 10 mM. This result indicates that at high concentrations, the uptake of undissociated weak acids activates electrogenic proton pumping, most likely by lowering cytoplasmic pH. In contrast, the hyperpolarization observed with concentrations of IAA four orders of magnitude lower appears to be a specific hormonal effect. This specific, auxin-induced hyperpolarization occurs at the same time as the initiation of net proton secretion and supports the hypothesis that auxin initiates extension growth by increasing proton pumping.

Equations have been developed to describe the diffusional movement of a weak acid such as the auxin indoleacetic acid through a long file of vacuolated cells, where cellular accumulation is driven by the pH gradients across the cell membranes. If the permeability to the auxin anion is greater at one end of the cell than at the other, diffusional movement takes the form of polar transport, which exhibits: a nearly constant velocity either for the front or for a pulse of radioactive auxin, the capacity to move auxin against an external gradient of concentration, and a polar ratio that increases exponentially with the length of the section. The determinants of velocity include both diffusion through the vacuole and permeation steps at the cell membranes. Except Since the early quantitative descriptions of the transport of the endogenous plant growth hormone auxin* (1-3) its cellular basis has been elusive. Among its characteristics are (i) a polarity, in which auxin moves more effectively through tissues in one direction than in the other, with the polar ratio increasing approximately exponentially with distance (4-6); (ii) the ability to move auxin through a tissue against an external concentration gradient (3); and (iii) a velocity, evidenced by a nearly constant rate of travel of 10 or more mm hr-' of either the front (2, 4, 7-10) or a pulse of radioactively labeled auxin introduced at the apical end of a section (11). Various models involving differential secretion from the basal ends of the cells can account for the first two of these features, but there has been no convincing explanation for the third.The chemosmotic polar diffusion hypothesis draws ideas and observations from several sources and postulates that polar transport involves both steps of membrane permeation and diffusion through the cell (12). Uptake of auxin is pH dependent and appears to be driven by the pH difference between the inside of the cell and the acidic wall space. With a difference of 2 pH units, and with only the undissociated acid permeant, auxin can accumulate passively to an internal concentration more than 50 times greater than the external. If the auxin anion is also permeant, this accumulation will be reduced. The suggestion that the polarity of transport is caused by a greater permeability to the anion at the basal than at the apical end of each cell (13, 14) is a central feature of this hypothesis.Because neither cytoplasmic streaming (15) nor a lateral sheath of cytoplasm around the vacuole (16) is necessary to support transport, the auxin seems to cross the tonoplast and diffuse through the vacuole in traversing the length of each cell. The proposed path of auxin movement (Fig. 1) The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.

The temporal relations between early responses to indoleacetic acid (IAA), proton secretion, hyperpolarization of the membrane potential, and growth change during the incubation of segments of oat (Avena sativa L.) coleoptiles in a low salt medium. When IAA is added after pretreatment of several hours, proton secretion increases after a latency of 7 minutes and reaches its maximum 10 to 15 minutes later. This timing coincides with both the increase in growth of the segments and the hyperpolarization of the membrane potential of parenchyma cells, consistent with the hypothesis that the change in membrane voltage reflects the activity of an electrogenic proton pump. The extent of IAA-induced hyperpolarization is substantially reduced by elevating [KClI1, most likely because this increases the passive conductance of the membrane. Neither growth nor proton secretion is affected by high [KCJIL (30 millimolar), indicating that neither process is limited by the magnitude of the membrane potential. These results are consistent with the acid growth hypothesis. Following short incubation times, however, IAA-induced hyperpolarization and growth are detected within 10 minutes, while acidification of the medium is delayed for more than 40 minutes. This result is seemingly in conflict with the acid growth hypothesis, but in freshly cut tissue, the pH of the external medium may not reflect the pH of the epidermal cell walls. The temporal coincidence of auxin-induced growth and hyperpolarization suggests that in freshly isolated segments the hyperpolarization is a more sensitive indication of proton secretion than is acidification of the external aqueous environment.through secondary effects on essential transport processes and metabolism, may be more important in the stimulation ofgrowth by auxin than is wall acidification.Since protons are secreted from the cell against both a considerable concentration and electrical gradient, this process requires energy presumably furnished by H+-translocating ATPases in the plasma membrane. Electrophysiological measurements have shown that both IAA and FC3 cause the plasma membrane to hyperpolarize, indicating that the increased proton secretion is electrogenic (1,9). Although a careful comparison has been lacking, the hyperpolarization should be coincident or even prior to measurable acidification of the external medium. The time required to acidify the inner portion of the wall where loosening occurs (26) is not known; however, the onset ofhyperpolarization should be a sensitive indication of the time at which proton secretion and wall acidification actually begins.The purpose of these experiments was to determine the temporal relations between the induction of membrane hyperpolarization, proton secretion, and growth by IAA. Furthermore, by adjusting the membrane potential and the auxin-induced hyperpolarization by varying [KCl]0, we examined the suggestion that a highly negative membrane potential per se is important for the stimulation of growth by auxin. MATERIALS AND METHODS...