Kinetics of Determination in the Differentiation of Isolated Mesophyll Cells of Zinnia elegans to Tracheary Elements

Mechanically isolated mesophyll cells of Zinnia elegans L. cv Envy differentiate to tracheary elements when cultured in inductive medium containing 0.5 micromolar α-naphthaleneacetic acid and 0.5 micromolar benzyladenine. The cells do not differentiate when cultured in medium in which the concentration of auxin and/or cytokinin has been reduced to 0.005 micromolar. Cells require an initial 24-hour exposure to inductive cytokinin and 56-hour exposure to inductive auxin for differentiation at 72 hours of culture. Freshly isolated Zinnia cells can be maintained in medium having low concentrations of both auxin and cytokinin for only 1 day without significant loss of potential to differentiate upon transfer to inductive medium. Initial culture for up to 2 days in medium having high auxin and low cytokinin, or low auxin and high cytokinin, allows full differentiation on the third day after transfer to inductive medium and potentiates the early differentiation of some cells.     

Increase of embryogenic callus formation in cucumber by initial culture on high concentration of 2,4-dichlorophenoxyacetic acid

Cucumber (Cucumis sativus L.) leaf explants were cultured either continuously on standard medium containing 4.5 µM 2,4- dichlorophenoxyacetic acid (2,4-d) and 4.4 µM benzylaminopurine, or first cultured for various periods at different levels of 2,4-d, picloram or naphthaleneacetic acid (NAA), and then transferred to standard medium. When cultured continuously on standard medium, less than 10% of the explants formed embryogenic callus. Initial culture on picloram or NAA, or on 2,4-d at a low concentration (1.4 µM) did not result in any embryogenic callus formation. Embryogenic callus formation increased to 40% if during the initial phase of the culture (10 days), the 2,4-d concentration was raised to 14 µM. Prolonged culture on 14 µM 2,4-d resulted in less embryogenic callus formation.Abbreviations BA benzylaminopurine - 2,4-d 2,4-dichlorophenoxyacetic acid - NAA naphthaleneacetic acid     

Derepression of amino Acid-h cotransport in developing soybean embryos

The uptake of the unnatural amino acid α-aminoisobutyric acid (AIB) and glutamine by developing soybean (Glycine max Merr. cv Chippewa 64) embryos was investigated. In freshly excised embryos, the accumulation ratio (cytoplasmic concentration/external concentration) of AIB did not exceed 1.0. After an 18-hour preincubation in nitrogen-free medium the accumulation ratio of AIB exceeded 4.5 at an external AIB concentration of 10 micromolar. This indicates the derepression of an active amino acid uptake mechanism operative at low external amino acid concentration. The presence of sucrose, NH₄NO₃, or glutamine during a 21-hour preincubation prior to measuring glutamine uptake inhibited the enhancement of uptake by 43%, 51%, and 96%, respectively. The time course of the decline in free amino acids and the time course of enhancement of amino acid uptake was not consistent with enhanced uptake resulting from relief of transinhibition, but suggested instead the derepression of synthesis of new carriers. The time course of enhancement of amino acid uptake was paralleled by an increase in glutamine-induced depolarization of the membrane potential. The kinetics of glutamine uptake indicated the presence of a saturable and a nonsaturable component of uptake. The saturable component of uptake is attributed to a mechanism of amino acid-H⁺ cotransport which is derepressed by nitrogen and/or carbon starvation. At physiological concentrations of amino acids, uptake through the saturable system in freshly excised embryos is negligible. Thus, uptake through the nonsaturable system is of primary importance in the nitrogen nutrition of developing soybean embryos.     

Occurrence of an Inhibitor of Tissue-Type Plasminogen Activator in Seeds and in Vitro Cultures of Erythrina caffra Thunb

The level of an inhibitor of tissue-type plasminogen activator (t-PA) increased slowly during the early developmental stage of seeds of Erythrina caffra Thunb. Thereafter, the inhibitor increased exponentially until the seeds reached maturity. At maturity, the t-PA inhibitor levels in the cotyledons were 38 times higher than the levels at the onset of seed development. The t-PA inhibitor accumulated at a faster rate than the storage proteins, which reached a concentration 15 times higher than the protein concentration at the onset of seed development. During the imbibition and germination process, the t-PA inhibitor decreased gradually. The inhibitor kept on decreasing during the growth of the seedlings until the 10th day after imbibition, when it leveled off at 4.1% of that of the initial inhibitor concentration. The inhibitor remained at this level until the cotyledons were shed at day 22. The total protein in the cotyledons decreased at a slower rate than the inhibitor and reached a minimum concentration at day 20 of 3.6% of the initial protein concentration in the cotyledons. Callus cultures of root, shoot, leaf, and cotyledonary tissue was established and maintained on Murashige-Skoog medium supplemented with 3% sucrose, 10 micromolar benzyladenine, and 5 micromolar 2,4-dichlorophenoxyacetic acid. A shoot cell suspension culture was established on Murashige-Skoog medium supplemented with 3% sucrose, 1 micromolar benzyladenine, and 0.5 micromolar 2,4-dichlorophenoxyacetic acid (pH 5.7) and shaken at 60 revolutions per minute. The level of t-PA inhibitor in root, shoot, leaf, and cotyledonary callus was substantially lower than in the corresponding intact tissue. The t-PA inhibitor levels in the linear growth phase was higher than in the lag or stationary growth phases of the cell suspension culture. A hydrolysate of the cell walls of tomato and E. caffra Thunb, as well as polyamines and organic acids, did not increase the concentration of t-PA inhibitor in suspension cultures or intact leaf tissue of E. caffra. The t-PA inhibitor levels of suspension cultures were increased by Na₂SO₄ but not by I-cysteine in the nutrient medium.     

Polar transport of 1-naphthaleneacetic Acid determines the distribution of flower buds on explants of tobacco

Upon addition of 1-naphthaleneacetic acid (1-NAA) and benzylaminopurine, flower buds developed on explants from flower stalks of Nicotiana tabacum L. cv Samsun cultured in vitro. At low concentrations of 1-NAA, buds emerged mainly at the basal edge, whereas at high concentrations they developed on the remaining surface. The optimum concentrations for the two groups of buds were 0.45 micromolar and 2.2 micromolar, respectively, and the shapes of the concentration versus response curves were similar. The level of benzylaminopurine in the medium affected neither the shape nor the optimum concentration of these curves. The distribution of the buds over the explants was shown to be caused by polar auxin transport, leading to accumulation at the basal side. First, in the presence of the inhibitors 2,3,5-triiodobenzoic acid and 1-naphthylphthalamic acid, both groups of buds had the same optimum concentration of 1 micromolar 1-NAA. Second, after 6 hours of culture applied 1-NAA had accumulated in the basal part of the explant. In the presence of 1-naphthylphthalamic acid, no transport or accumulation of applied 1-NAA occurred.     

Development of Defined, Minimal, and Complete Media for the Growth of Hyphomicrobium neptunium

A complete synthetic medium containing 15 amino acids, a minimal synthetic medium (GAMS) containing 4 amino acids, and a supplemented minimal medium (GAMS + calcium pantothenate) have been developed for the cultivation of Hyphomicrobium neptunium ATCC 15444. Depending on the complexity of the synthetic media, generation times were approximately 2 to 3 times longer, and maximum cell densities were 0.3 to 0.9 log₁₀ lower than in ZoBell marine broth 2216. The fates of ¹⁴C-labeled amino acids in GAMS were monitored. Results suggested that H. neptunium was auxotrophic for methionine, utilized glutamic acid as a primary energy source, and readily anabolized and catabolized serine and aspartic acid. Individual amino acid concentrations above 125 mM induced prolonged lag periods, whereas only methionine was not growth limiting at a concentration as low as 2 mM.     

Cloning,Expression, and Functional Characterization of Secondary Amino Acid Transporters of Lactococcus lactis

Fourteen genes encoding putative secondary amino acid transporters were identified in the genomes of Lactococcus lactis subsp. cremoris strains MG1363 and SK11 and L. lactis subsp. lactis strains IL1403 and KF147, 12 of which were common to all four strains. Amino acid uptake in L. lactis cells overexpressing the genes revealed transporters specific for histidine, lysine, arginine, agmatine, putrescine, aromatic amino acids, acidic amino acids, serine, and branched-chain amino acids. Substrate specificities were demonstrated by inhibition profiles determined in the presence of excesses of the other amino acids. Four knockout mutants, lacking the lysine transporter LysP, the histidine transporter HisP (formerly LysQ), the acidic amino acid transporter AcaP (YlcA), or the aromatic amino acid transporter FywP (YsjA), were constructed. The LysP, HisP, and FywP deletion mutants showed drastically decreased rates of uptake of the corresponding substrates at low concentrations. The same was observed for the AcaP mutant with aspartate but not with glutamate. In rich M17 medium, the deletion of none of the transporters affected growth. In contrast, the deletion of the HisP, AcaP, and FywP transporters did affect growth in a defined medium with free amino acids as the sole amino acid source. HisP was essential at low histidine concentrations, and AcaP was essential in the absence of glutamine. FywP appeared to play a role in retaining intracellularly synthesized aromatic amino acids when these were not added to the medium. Finally, HisP, AcaP, and FywP did not play a role in the excretion of accumulated histidine, glutamate, or phenylalanine, respectively, indicating the involvement of other transporters.     

Lactate and Amino Acid Catabolism in the Cheese-Ripening Yeast Yarrowia lipolytica

The consumption of lactate and amino acids is very important for microbial development and/or aroma production during cheese ripening. A strain of Yarrowia lipolytica isolated from cheese was grown in a liquid medium containing lactate in the presence of a low (0.1×) or high (2×) concentration of amino acids. Our results show that there was a dramatic increase in the growth of Y. lipolytica in the medium containing a high amino acid concentration, but there was limited lactate consumption. Conversely, lactate was efficiently consumed in the medium containing a low concentration of amino acids after amino acid depletion was complete. These data suggest that the amino acids are used by Y. lipolytica as a main energy source, whereas lactate is consumed following amino acid depletion. Amino acid degradation was accompanied by ammonia production corresponding to a dramatic increase in the pH. The effect of adding amino acids to a Y. lipolytica culture grown on lactate was also investigated. Real-time quantitative PCR analyses were performed with specific primers for five genes involved in amino acid transport and catabolism, including an amino acid transporter gene (GAP1) and four aminotransferase genes (ARO8, ARO9, BAT1, and BAT2). The expression of three genes involved in lactate transport and catabolism was also studied. These genes included a lactate transporter gene (JEN1) and two lactate dehydrogenase genes (CYB2-1 and CYB2-2). Our data showed that GAP1, BAT2, BAT1, and ARO8 were maximally expressed after 15 to 30 min following addition of amino acids (BAT2 was the most highly expressed gene), while the maximum expression of JEN1, CYB2-1, and CYB2-2 was delayed (≥60 min).     

Precocious Germination during In Vitro Growth of Soybean Seeds

Immature Glycine max (L.) Merrill seeds were grown and matured in liquid medium at 25°C under fluorescent light. In standard medium containing minerals, 146 millimolar sucrose and 62.5 millimolar glutamine (osmolality 0.24), precocious germination seldom occurred with a starting seed size of less than 300 milligrams fresh weight. Frequency of precocious germination increased with increased starting seed size. Sucrose concentration strongly affected precocious germination while glutamine concentration had no effect. Starting with 300 to 350 milligrams fresh weight seeds, treatments which reduced the sucrose concentration or lowered the osmolality of the culture medium stimulated precocious germination, and increased the fresh weight growth but not the dry weight growth of seeds. Increasing the osmolality to 0.38 with sucrose or mannitol prevented precocious germination without reducing dry weight accumulation in seeds. In medium with initially low osmolality, precocious germination was inhibited by addition of 1 to 100 micromolar abscisic acid to the medium without a reduction in seed growth. During growth and maturation of large soybean seeds in vitro, precocious germination and other abnormal tissue growth can be prevented by high sucrose or mannitol concentrations in the medium or by addition of abscisic acid.     

Analysis of free amino acids during fermentation by Bacillus subtilis using capillary electrophoresis

A high performance capillary electrophoresis (HPCE) method was presented to identify and quantitate free amino acids during fermentation by Bacillus subtilis. Amino acids, pre-column derivatized with phenylisothicyanate, were separated and characterized by HPCE. In order to optimize separation conditions, the assay was developed by varying the β-cyclodextrin concentration and pH of the background electrolyte. A buffer system comprising 30 mM phosphate and 3 mM β-cyclodextrin at pH 7.0, voltage of 20 kV and detection wavelength of 254 nm showed the best results, with 17 out of 20 phenylthioncarbamyl amino acids in a solution adequately separated. For quantification, p-aminobenzoic acid was added as an internal standard. Analysis of free amino acids in Bacillus subtilis culture medium using this method revealed good consistency with the values obtained using conventional ninhydrin-based amino acid analyzer. Four free amino acids (aspartic acid, glutamic acid, proline, and tyrosine) concentration in an extracellular matrix during fermentation by Bacillus subtilis were mainly monitored using this method.     

Interface-mediated death of unconditioned Tetrahymena cells: effect of the medium composition

ABSTRACT We have previously shown that the cell death of Tetrahymena thermophila in low inocula cultures in a chemically-defined medium is not apoptotic. The death is caused by a cell lysis occurring at the medium-air interface and can be prevented by the addition of insulin or Pluronic F-68. Here, we report that cell death can also be caused by the medium. The specific effects of several medium constituents were tested in the presence and absence of an interface. Four of the 19 amino acids (arginine, aspartic acid, glutamic acid, and histidine in millimolar concentration) as well as Ca²⁺ (68 μM) and Mg²⁺ (2 mM) and trace metal ions (micromolar concentrations) are all sufficient to induce the interface-mediated death. The effect of the amino acids and the salt ions Ca²⁺ and Mg²⁺ can be abolished by the addition of insulin (10^-6 M) or Pluronic F-68 (0.01% w/v), whereas insulin/Pluronic F-68 only postpones the death induced by trace metal ions. On the basis of our findings, a new recipe for a chemically-defined medium has been formulated. Single cells can grow in this medium in the presence of medium-air interface without any supplements.     

Control and Kinetics of Branch Root Formation in Cultured Root Segments of Haplopappus ravenii

Branch root formation required only the presence of minerals, sucrose as a carbon source, and an auxin. The number of primordia formed was a function of auxin concentration. With naphthaleneacetic acid at 0.1 mg/l, up to 60 or more branches were formed per centimeter of Haplopappus ravenii root segment. Under our conditions, pea root segments formed only five or six branches per centimeter, but tomato and radish, like H. ravenii, formed large numbers of branches. Cytokinin inhibited branch formation, while gibberellic acid was without effect. Vitamins were not required for branch formation, although they enhanced elongation. Up to 5 days were required for the maximum number of stable branch primordia to form under the influence of naphthaleneacetic acid. If naphthaleneacetic acid was withdrawn earlier, fewer branch primordia developed. The requirement for a lengthy exposure to naphthaleneacetic acid, the kinetics of the response, and the ease with which naphthaleneacetic acid could be rinsed out of the tissue with consequent cessation of branch root formation, were similar to other hormone-regulated developmental systems. Anatomical and cytological studies were made of segments exposed for various times to auxin. The segments were mostly diarch, and branches formed obliquely to protoxylem poles. While primarily only pericycle-endodermis cells divided, both these and cortex cells responded in the first 24 hours exposure to naphthaleneacetic acid with enlarged nuclei and nucleoli, and a few cortical cells divided. Maximum nucleus and nucleolus size was reached approximately 9 hours after exposure to naphthaleneacetic acid. Branches rarely elongated more than 5 cm before their meristems died. The H. ravenii culture is maintained only by the frequent formation of new naphthaleneacetic acid-induced branches.     

Continuous Culture of Ruminal Microorganisms in Chemically Defined Medium: II. Culture Medium Studies

Ruminal ciliates have been grown in continuous culture in chemically defined media and in the absence of viable bacteria. Oligotrichic ruminal ciliates seem to require insoluble carbohydrates for growth; the holotrichic ciliates require soluble carbohydrates, but at low concentrations. Both groups of ciliates utilize amino acids as their principal nitrogen source when these are supplied in micromolar concentrations; at millimolar concentrations, amino acids are toxic, possibly from excessive ammonia formation arising from ciliate deaminase activity. Holotrichic ruminal ciliates are destroyed by overdeposition of amylopectin when glucose is present above 0.1% concentration in the medium. Ecological requirements of ruminal ciliates are also described.     

The effect of amino acid composition of serum-free medium on DNA synthesis in primary hepatocyte cultures in the presence of epidermal growth factor

Summary The presence of optimal nutritional elements in cell culture medium is very important in studies of cultured cells. For this reason, several researchers have experimented with adding or increasing the concentration of one or more amino acids to the medium they were using to determine “essential” amino acids and optimal concentrations. We studied how leaving out one amino acid at a time from Dulbecco’s modified Eagle’s medium would affect epidermal growth factor-induced DNA synthesis in primary hepatocytes of the rat. Our “modified” DMEM contained only eight amino acids: arginine, cysteine, isoleucine, leucine, lysine, phenylalanine, tryptophan, and valine. Proline was found to be an essential amino acid in normal DMEM but not in the modified DMEM, and some other amino acids reduced DNA synthesis in this medium. This study showed that perhaps no single amino acid such as proline can be called “essential,” but rather an optimal balance of amino acids is required for each major function of each cell type cultured.     

Micropropagation of juvenile and adult Sorbus domestica L.

Successful propagation of seedlings and mature trees of Sorbus domestica L. has been achieved by in vitro methods. Multiple shoot formation was obtained by placing shoot apices or nodal segments on a modified Schenck and Hildebrandt medium containing benzyladenine. Regenerated shoots were excised and induced to root on media with auxin. In the best treatments 75–85% of shoots from juvenile material rooted. Rooting capacity of shoots from mature explants was lower (30%) and was not improved by dipping the base of shoots in concentration solutions of indolebutyric or naphthaleneacetic acids. Plantlets were ultimately established in soil.Abbreviations BA benzyladenine - IAA indoleacetic acid - IBA indolebutyric acid - NAA naphthaleneacetic acid     

Growth Characteristics of Bartonella henselae in a Novel Liquid Medium: Primary Isolation, Growth-Phase-Dependent Phage Induction, and Metabolic Studies

Bartonella henselae is a zoonotic pathogen that usually causes a self-limiting infection in immunocompetent individuals but often causes potentially life-threatening infections, such as bacillary angiomatosis, in immunocompromised patients. Both diagnosis of infection and research into the molecular mechanisms of pathogenesis have been hindered by the absence of a suitable liquid growth medium. It has been difficult to isolate B. henselae directly from the blood of infected humans or animals or to grow the bacteria in liquid culture media under laboratory conditions. Therefore, we have developed a liquid growth medium that supports reproducible in vitro growth (3-h doubling time and a growth yield of approximately 5 × 10⁸ CFU/ml) and permits the isolation of B. henselae from the blood of infected cats. During the development of this medium, we observed that B. henselae did not derive carbon and energy from the catabolism of glucose, which is consistent with genome nucleotide sequence data suggesting an incomplete glycolytic pathway. Of interest, B. henselae depleted amino acids from the culture medium and accumulated ammonia in the medium, an indicator of amino acid catabolism. Analysis of the culture medium throughout the growth cycle revealed that oxygen was consumed and carbon dioxide was generated, suggesting that amino acids were catabolized in a tricarboxylic acid (TCA) cycle-dependent mechanism. Additionally, phage particles were detected in the culture supernatants of stationary-phase B. henselae, but not in mid-logarithmic-phase culture supernatants. Enzymatic assays of whole-cell lysates revealed that B. henselae has a complete TCA cycle. Taken together, these data suggest B. henselae may catabolize amino acids but not glucose to derive carbon and energy from its host. Furthermore, the newly developed culture medium should improve isolation of B. henselae and basic research into the pathogenesis of the bacterium.     

Amino acids metabolism by VO 208 hybridoma cells: some aspects of the culture process and medium composition influence

In the present study an approach has been developed in order to examine the consequence of essential and non essential amino acid supplementation on VO208 hybridoma cells behaviour. The effect of amino acid enrichment has been studied taking into account the culture process, i.e., batch or continuous culture mode and the medium composition, i.e., a home made serum-free medium or a serum containing one. A group of 4 amino acids, i.e., Ser, Pro, Gly and Arg presented atypical evolution pattern of their extracellular concentration depending on the type of the medium and on the culture mode. Some amino acids were probably involved in the limitation of the cellular proliferation. Met was one of the amino acids that appears to may have been at limiting concentration in all cases. In continuous culture mode, an enrichment of amino acids resulted in a rapid improvement of the viable cell density in both media, with or without the presence of serum. For most amino acids, supplementation during continuous culture induced an increase of the amino acid uptake rate. A comparative analysis of amino acids utilisation, depending on the culture conditions studied in the present study, has been performed in order to propose an overall picture of amino acids metabolism by VO 208 Hybridoma cell line. This revised version was published online in August 2006 with corrections to the Cover Date.     

Stoichiometry,Kinetics, and Regulation of Glucose and Amino Acid Metabolism of a Recombinant BHK Cell Line in Batch and Continuous Cultures

Batch and continuous cultures were carried out to study the stoichiometry, kinetics, and regulation of glucose and amino acid metabolism of a recombinant BHK cell line, with particular attention to the metabolism at low levels of glucose and glutamine. The apparent yields of cells on glucose and glutamine, lactate on glucose, and ammonium on glutamine were all found to change significantly at low residual concentrations of glucose (<5 mmol/L) and glutamine (<1 mmol/L) . The uptake rates of glucose and glutamine were markedly reduced at low concentrations, leading to a more effective utilization of these nutrients for energy metabolism and biosynthesis and reduced formation rates of lactate and ammonium. However, the consumption of other amino acids, especially the essential amino acids leucine, isoleucine, and valine and the nonessential amino acids serine and glutamate, was strongly enhanced at low glutamine concentration. Quantitatively, it was shown that the cellular yields and rates associated with glucose metabolism were primarily determined by the residual glucose concentration, while those associated with glutamine metabolism depended mainly on the residual glutamine. Both experimental results and analysis of the kinetic data with models showed that the glucose metabolism of BHK cells is not affected by glutamine except for a slight influence under glucose limitation and glutaminolysis not by glucose, at least not significantly under the experimental conditions. Compared to hybridoma and other cultured animal cells, the recombinant BHK cell line showed remarkable differences in terms of nutrient sensitivity, stoichiometry, and amino acid metabolism at low levels of nutrients. These cell-line-specific stoichiometry and nutrient needs should be considered when designing an optimal medium and/or feeding strategy for achieving high cell density and high productivity of BHK cells. In this work, a cell density of 1.1 × 10⁷ cells/mL was achieved in a conventional continuous culture by using a proper feed medium.     

Very Low Ethanol Concentrations Affect the Viability and Growth Recovery in Post-Stationary-Phase Staphylococcus aureus Populations

Pharmaceuticals, culture media used for in vitro diagnostics and research, human body fluids, and environments can retain very low ethanol concentrations (VLEC) (≤0.1%, vol/vol). In contrast to the well-established effects of elevated ethanol concentrations on bacteria, little is known about the consequences of exposure to VLEC. We supplemented growth media for Staphylococcus aureus strain DSM20231 with VLEC (VLEC⁺ conditions) and determined ultramorphology, growth, and viability compared to those with unsupplemented media (VLEC⁻ conditions) for prolonged culture times (up to 8 days). VLEC⁺-grown late-stationary-phase S. aureus displayed extensive alterations of cell integrity as shown by scanning electron microscopy. Surprisingly, while ethanol in the medium was completely metabolized during exponential phase, a profound delay of S. aureus post-stationary-phase recovery (>48 h) was observed. Concomitantly, under VLEC⁺ conditions, the concentration of acetate in the culture medium remained elevated while that of ammonia was reduced, contributing to an acidic culture medium and suggesting decreased amino acid catabolism. Interestingly, amino acid depletion was not uniformly affected: under VLEC⁺ conditions, glutamic acid, ornithine, and proline remained in the culture medium while the uptake of other amino acids was not affected. Supplementation with arginine, but not with other amino acids, was able to restore post-stationary-phase growth and viability. Taken together, these data demonstrate that VLEC have profound effects on the recovery of S. aureus even after ethanol depletion and delay the transition from primary to secondary metabolite catabolism. These data also suggest that the concentration of ethanol needed for bacteriostatic control of S. aureus is lower than that previously reported.     

The Proton Electrochemical Transmembrane Gradients Generated by the Transfer Cells of the Haustorium of Polytrichum formosum and Their Use in the Uptake of Amino Acids

The epidermal cells of the sporophyte haustorium of Polytrichum formosum are modified into transfer cells. These cells are located in a strategic place allowing them to control the exchanges between the two generations. Their plasmalemma creates proton gradients (Δψ and ΔpH) which increase during the development of the sporophyte. As the sporophyte grows from 2 to 4 cm long, the pH of the incubation medium of the haustoria decreases from 5.2 to 4.3, and the transmembrane potential difference (PD) hyperpolarizes form −140 to −210 millivolts. These gradients become rapidly larger than that generated by the plasmalemma of the basal cells of the sporophyte. They are used to energize the uptake of the solutes present in the apoplast of the gametophyte, particularly the amino acids. Below 20 micromolar α-aminoisobutyric acid uptake in the transfer cells is mediated by a saturable system and is optimal at acidic pH (4.0 and 4.5). It is strongly inhibited by compounds dissipating both Δψ and ΔpH (10 micromolar carbonylcyanide-m-chlorophenyl hydrazone) or only Δψ (0.1 molar KCl). The absorption of α-aminoisobutyric acid and of the other neutral amino acids tested induces an alkalinization of the medium and a depolarization of membrane potential difference which is concentration dependent. These data show that the uptake of amino acids by the transfer cells of the haustorium is a secondary translocation (proton-amino acid symport) energized by a primary translocation (proton efflux). More particularly, they show that transfer cells possess a membrane enzymic equipment particularly efficient to achieve the uptake of the solutes leaked in the apoplast from other cell types.