ELECTRON TRANSPORT SYSTEMS OF RHIZOBIUM GROWN IN NODULES AND IN LABORATORY MEDIUM

The electon transport systems of Rhizobium japonicum were studied,comparing cells harvested from effective nodules with thosefrom artificial culture. Participation of the cytochrome systemwas confirmed in both forms of cells. Absorption peaks of thecytochromes of cultured cells were a, b, c type, resemblingthose of Bacillus subtilis, yeast and mammalian tissue. Cytochromea could not be detected in the absorption spectrum of symbioticcells, although the CO binding difference spectrum showed apeak at about 438 mµ, which can be attributed to a componenta₃ or a₁. CO difference spectrum also showed a shoulder at about416 mµ. Cells cultivated under the insufficient supply of oxygen showedthe cytochrome absorption spectrum closely resembled that ofsymbiotic cells. Diaphorase activity was lower in symbioticcells. These results are considered to be due to the insufficientsupply of oxygen within nodule tissue. Succinate oxidation bythe symbiotic cell paniculate was shown to be carbon monooxideresistant. NADH₂ oxidation by the supernatant fraction of symbioticcells was accelerated by flavin mononucleotide, 2, 6-dichiorophenolindophenol, methylene blue and vitamin K₃. ¹Present address: Faculty of Agriculture, Tôhoku University,Sendai. ²Present address: Central Agricultural Experiment Station, Kitamoto.     

Growth, metabolism and growth efficiency of a euphausiid crustacean Euphausia pacifica in the southern Japan Sea, as influenced by temperature

Growth (assessed from intermolt period and molt increment) andmetabolism (oxygen consumption) of the post-larva of Euphausiapacifica from the southern Japan Sea were determined at sevengraded temperatures ranging from 1 to 25°C. The intermoltperiod shortened progressively as temperature increased from1 to 20°C, but an effect of temperature on molt-to-moltgrowth increment was not seen. Oxygen consumption rates wereaccelerated by the increase in temperature up to 20°C. Beyond20°C, E.pacifica exhibited reduced oxygen consumption anddied within 1 day without molting. After removing the effectof body size, the relationships between growth rate and temperature,and between oxygen consumption rate and temperature, were established.The carbon budget was calculated as a function of temperature.Because of differential effects of temperature on growth andmetabolism, the net growth efficiency [K₂ growthx100/(growth+metabolism)]changed with temperature. The optimum temperature at which E.pacificaattained the maximum K₂ was 11.4°C, which was derived fromcalculation of cumulative carbon invested in growth and metabolismin this animal. In an alternative method, the optimum temperaturewas obtained mathematically by solving a set of differentialequations. The biological and ecological significance of theoptimum temperature which leads to the maximum K₂ is discussed.     

Changes in Respiratory Potential of Dormant and Non-dormant Galium aparine L. (cleavers) Seeds during Dry Storage

Hilton, J. R. and Thomas, J. A. 1987. Changes in respiratorypotential of dormant and non-dormant Galium aparine L. (cleavers)seeds during dry storage.—J. exp. Bot. 38: 1484–1490. Pre-germinative rates of O₂ uptake of two collections of Galiumaparine L. seed were compared throughout a 9 month period ofdry storage at 4 °C, 23 °C and at ambient (frost-protected)temperatures. Uptake of O₂ by the dormant seeds was generallyhigher than that of the less dormant seeds except when freshly-harvested.Moreover, changes in the O₂ consumption of seeds stored at ambienttemperatures could be associated with periods of germinationand seedling emergence in the field. The results are discussedin relation to changes in respiratory metabolism during dormancybreakage.     

Changes in Photosynthetic Characteristics Induced by Transferring Air-Grown Cells of Chlorococcum littorale to High-CO2 Conditions

When air-grown cells of Chlorococcum littorale was enrichedwith CO₂, growth was enhanced after a lag period of one to twodays at 20% CO₂, and 3 to 6 days at 40% CO₂. Changes in therate of photosynthesis measured as oxygen evolution and CO₂fixation, were similar to those observed for growth. Duringthe initial inhibition of photosynthesis in 40% CO₂, the activityof PSII was suppressed. In contrast, PSI activity was greatlyenhanced. Air-grown cells of C. littorale possessed comparatively highcarbonic anhydrase (CA) activity which was localized insidethe cells and on the cell surface. Under high CO₂ concentrationsextracellular CA activity was greatly suppressed and intracellularactivity almost completely abolished. Phosphoenol pyruvate carboxylaseactivity was also suppressed in high CO₂-grown cells. Ribulose-l,5-bisphosphatecarboxylase activity was higher in high-CO₂ grown cells thanin air-grown cells. The above results indicated that the lagphase induced by 40% CO₂ was due to suppression of PSII activity. ¹Part of this work was reported in the International PhotosynthesisCongress, Nagoya, 1992.     

Carotenoid photobleaching induced by photosystem II action in the membrane fragments of the blue-green alga Anabaena variabilis : Action of O2

Carotenoid photobleaching induced by photosystem II action wasstudied using membrane fragments of the blue-green alga Anabaenavariabilis. Special attention was paid to the action of O₂. Carotenoid photobleaching elicited by carbonyl cyanide m-chlorophenylhydrazone(CCCP) depended on O₂. However, the addition of H₂O₂, sodiumsilicotungstate or potassium ferricyanide (Ferri), an electronacceptor for reaction center II action, removed the O₂-dependency.These results indicate that O₂ acts as the electron acceptorfor this reaction. When both CGCP and Ferri were present, a short illumination(0.25 sec) caused a rapid photobleaching followed by a slowrecovery in the subsequent dark period. The spectrum of theabsorption decrease in the light was identical with that ofthe absorption increase in the subsequent dark, indicating thata reversible process is involved in the carotenoid photobleaching.The size in the dark recovery relative to the light bleachingbecame larger under anaerobic conditions and smaller under higherpartial pressure of O₂. The reuslts were interpreted as indicatingthat O₂ does not function in the primary process including areversible bleaching step, but is involved in the slow and irreversiblebleaching process. (Received April 3, 1978; )     

Inhibition of growth and photosynthesis of freshwater phytoplankton by ultraviolet A (UVA) radiation and subsequent recovery from stress

The effect of ultraviolet A (UVA) on growth and photosyntheticrate was studied in diatoms (Melosira spp.) of the phytoplanktonof a eutrophic lake and a cultured green alga Chloretla ellipsoidea.The cells were incubated under photosynthetically active radiation(PAR) (–UVA) or PAR + UVA conditions (+UVA). Growth ofC.ellipsoidea was retarded under +UVA, as shown by an increasein the lag period, but the rate of exponential growth was almostthe same in + and –UVA conditions. The photosyntheticrate was depressed markedly by UVA in Chlorella cells grownunder –UVA. In contrast, cells grown in +UVA showed onlyslight inhibition by UVA and after exposure to UVA for 6 daysthere was no inhibition. During the growth experiment, the cellularchlorophyll a content was higher in +UVA than +UVA grown cells.A similar effect was observed in diatoms from the eutrophicLake Suwa. In vivo fluorescence with (F_a) and without 3-(3,4-dichloropheny)-l,l-dimethylurea (DCMU) (F_b) and the photosynthetic rate were measured forC.ellipsoidea and the diatoms for 5 h under + and –UVAconditions. Soon after C.ellipsoidea had been subjected to +UVA,F_b and F_a / F_b decreased quickly and reached minima after 40min and 1 h, respectively. The suppressed in vivo fluorescenceresumed and full recovery was achieved after 4 h. This suggeststhat reactivation of the photosystem is acquired under prolongedexposure to UVA. A similar shift of F_a + F_b, but no change inFb, was found in diatoms by exposure to UVA. Changes in photosyntheticoxygen evolution by C.ellipsoidea under +UVA were similar tochanges in F_a + F_b. Degradation of chlorophyll a extracted inmethanol was enhanced by UVA. The rate of degradation by UVAwas independent of temperature from 15 to 34°C, suggestinga photochemical reaction. The results indicate that C.ellipsoideaand Melosira spp. acclimatize to prolonged UVA exposure by reactivationof the photosystem and enhanced cellular chlorophyll a synthesis.The ecological importance of these results to phytoplanktonproductivity in natural aquatic environments is discussed.     

Studies on the Photoreceptors for the Promotion and Inhibition of Flowering in Dark-Grown Seedlings of Pharbitis nil Choisy

Three-day-old etiolated seedlings of Pharbitis nil were exposedto red light for 10 min and sprayed with N⁶-benzyladenine beforetransfer to a 48-h inductive dark period, after which they weregrown under continuous white light. A second red irradiationpromoted flowering when given at the 5 and 24th hour of theinductive dark period but inhibited flowering at the 10 and15th hour. Far-red light inhibited flowering when given at anytime during the first 24 h of the dark period. Red/far-red reversibilitywas clearly observed at the 0, 5, 10 and 24th hour, but notat the 15th hour when both red and far-red lights completelyinhibited flowering. The action spectrum for the inhibition of flowering at the 15thhour of the inductive dark period had a sharply defined peakat 660 nm and closely resembled the absorption spectrum of theP_R form of phytochrome. The photoreceptors involved in thesephotoreactions are discussed. (Received June 10, 1983; Accepted July 6, 1983)     

Properties of the Cytochrome c Oxidase Activity of Cytochrome b561 from Photoanaerobically Grown Rhodopseudomonas sphaeroides

Cytochrome b₅₆₁ from Rhodopseudomonas sphaeroides had cytochromec (c2) oxidase activity and a pH optimum at 6.0 for this activity.The activity was affected by the ionic strength of the reactionmixture. The apparent K_m and maximal velocity (V_max) valuesin the absence of addea salts were 14 µM and 120 nmoloxidized per min per mg protein for horse heart cytochrome c.Reduced horse heart cytochrome c was reoxidized in first-orderkinetics by this cytochrome b₅₆₁. The specific activity was0.7 s^–1 per mg protein at 20°C at the concentrationof 30 µMM cytochrome c. Activity was inhibited by KCN and NaN₃, but not by antimycin.The addition of a low concentration of KCN to the cytochromeb₅₆₁ produced a change in the absorption spectrum, evidencethat KCN interacts with the heme moiety of cytochrome b₅₆₁.Results of this and preceeding studies show that the cytochromeoxidase (cytochrome "o") described earlier (Sasaki et al. 1970)is cytochrome b₅₆₁. (Received May 16, 1983; Accepted September 8, 1983)     

Mineral nutrition, of cultured chlorophyllous cells of tobacco VII. Effects of the NH4/NO3 ratio and of Cl in the media on the growth and ionic balance of cells

NG, a strain of cultured tobacco cells of Nicotiana glutinosahad high growth rates and carboxylate contents (C—A) of100 to 130 meq/100 g of dry cells on media containing 42 meqNO₃/liter as the sole N source. (C—A) is the amount ofinorganic cations minus inorganic anions in meq per 100 g ofdry cells. NG, cultured on media containing NH₄ 10+NO₃ 42 in meq/liter,had lower growth rates and lower (C—A) values as comparedwith NG on media containing NO₃ as the sole N source. NG, cultured on media containing NH₄ 30+NO₃ 42 in meq/liter,had high growth rates and (A—C) values of 22 to 53 meq/100gof dry cells. In this case, the (A—C) content may correspondto organic cations, basic organic N compounds such as free asprotein-bound basic amino acids. The easily absorbed Cl mayhave been required maintain good growth conditions such as ionicbalance and a favorable pH in the cells. Thus cultured cells of Nicotiana glutinosa may have physiologicaladaptability against variations in a relatively wide range of|C—A| contents [|C—A| being the absolute valuesof (C—A)]. (Received May 15, 1980; )     

CO2 chemosensitivity in Helix aspersa: three potassium currents mediate pH-sensitive neuronal spike timing

Elevated levels of carbon dioxide increase lung ventilation in Helix aspersa. The hypercapnic response originates from a discrete respiratory chemosensory region in the dorsal subesophageal ganglia that contains CO₂-sensitive neurons. We tested the hypothesis that pH-dependent inhibition of potassium channels in neurons in this region mediated the chemosensory response to CO₂. Cells isolated from the dorsal subesophageal ganglia retained CO₂ chemosensitivity and exhibited membrane depolarization and/or an increase in input resistance during an acid challenge. Isolated somata expressed two voltage-dependent potassium channels, an A-type and a delayed-rectifier-type channel (I_KA and I_KDR). Both conductances were inhibited during hypercapnia. The pattern of voltage dependence indicated that I_KA was affected by extracellular or intracellular pH, but the activity of I_KDR was modulated by extracellular pH only. Application of inhibitors of either channel mimicked many of the effects of acidification in isolated cells and neurons in situ. We also detected evidence of a pH-sensitive calcium-activated potassium channel (I_KCa) in neurons in situ. The results of these studies support the hypothesis that I_KA initiates the chemosensory response, and I_KDR and I_KCa prolong the period of activation of CO₂-sensitive neurons. Thus multiple potassium channels are inhibited by acidosis, and the combined effect of pH-dependent inhibition of these channels enhances neuronal excitability and mediates CO₂ chemosensory responses in H. aspersa. We did not find a single "chemosensory channel," and the chemosensitive channels that we did find were not unique in any way that we could detect. The protein "machinery" of CO₂ chemosensitivity is probably widespread among neurons, and the selection process whereby a neuron acts or does not act as a respiratory CO₂ chemosensor probably depends on the resting membrane potential and synaptic connectivity. carbon dioxide     

Stomatal Characteristics of Four Native Herbs Following Exposure to Elevated CO2

Contrasting effects on the stomatal index (SI), stomatal density,epidermal cell size and number were observed in four chalk grasslandherbs (Sanguisorba minor Scop., Lotus corniculatus L., Anthyllisvulneraria L. and Plantago media L.) following exposure to elevatedcarbon dioxide concentrations (CO₂) in controlled environmentgrowth cabinets. SI of S. minor increased for both leaf surfaces,whilst in A. vulneraria and P. media SI decreased on one surfaceonly. In L. corniculatus , no differences in SI were observedas epidermal cell density changed in parallel with stomataldensity. In L. corniculatus and S. minor stomatal density increasedon both surfaces, whereas in P. media it decreased; in A. vulnerariastomatal density decreased on the abaxial leaf surface alonefollowing exposure to elevated CO₂. In the latter three species,SI changed because stomatal density did not change in parallelwith epidermal cell density. The results suggest elevated CO₂is either directly or indirectly affecting cell differentiationand thus stomatal initiation in the meristem. In S. minor and P. media leaf growth increased in elevated CO₂,because of increased cell expansion of epidermal cells, whereasin L. corniculatus, epidermal cell size decreased and greaterleaf growth was because of an increase in epidermal cell divisions.In A. vulneraria, leaf size did not change, but increased cellexpansion on the adaxial surface suggests CO₂ affects leaf surfacesdifferently, either directly or indirectly at the cell differentiationstage or as the leaf grows. These results suggest component species of a plant communitymay differ in their response to elevated CO₂. Predicting theeffect of environmental change is therefore difficult.Copyright1994, 1999 Academic Press Elevated CO₂, Sanguisorba minor (salad burnet), Lotus corniculatus (birdsfoot trefoil), Anthyllis vulneraria (kidney vetch), Plantago media (hoary plantain), stomatal index, stomatal density, epidermal cell size     

Action Spectra Confirm Two Separate Actions of Phytochrome in the Induction of Flowering in Lemna paucicostata 441

Action spectra studies have shown that in the short day plant(SDP) Lemna paucicostat441 there are at least two actions ofphytochrome in the induction of flowering. At the beginningof the dark period far-red light inhibited flowering, and theaction spectrum corresponded to the absorption spectrum of P_FR,while at the middle of the inductive dark period both red andfar-red light were inhibitory. The action spectrum for the redlight corresponded to that of PR absorption, but there was activityin the region beyond 720 nm which exactly coincided with theabsorption by P_FR observed at the beginning of the dark period,indicating that at the middle of the dark period there was absorptionby both P_R and P_FR. The difference in quantum efficiency betweenthe red and far-red light effects was about 60-fold. These resultsare consistent with there being a stable pool of P_FR necessaryfor the induction of flowering and another pool of phytochromein a different cellular environment which participates in thenight-break reaction as P_R. ¹ Present address: School of Applied Biology, Faculty of Science,Lancashire Polytechnic, Preston PR1 2TQ, U.K. ² 2 Present address: Division of Environmental Biology, NationalInstitute for Environmental Studies, Yatabemachi, Tsukuba, Ibaraki305, Japan. ³ Present address: Division of Plant Biological Regulation,The Riken Institute for Frontier Research Program, Hirosawa,Wako-shi, 351-01, Japan. (Received December 13, 1986; Accepted July 17, 1987)     

Oxygen consumption in the marine bacterium Pseudomonas nautica predicted from ETS activity and bisubstrate enzyme kinetics

The respiratory O₂ consumption in aerobic bacterial cultureshas been modeled from the time profiles of the in vitro activityof the respiratory electron transfer system (ETS), the bacterialprotein and the concentration of the carbon source in the cultures.The model was based on the concept of bisubstrate kinetic controlof the ETS throughout the exponential, steady-state and senescentphases of the cultures. In the exponential phase, the measuredrates of O₂ consumption and the in vitro ETS activity were closelycoupled, but in the senescent phase, they were uncoupled. Thein vitro ETS activity remained high even after the culture'scarbon source was exhausted, while the O₂ consumption fell tolow levels. Based on the hypothesis that this uncoupling wascaused by limitation of the intracellular ETS substrates (NADHand NADPH), a semi-empirical model incorporating a bisubstrateenzyme kinetics algorithm was formulated and fitted to the observationsof the experiments. The model predicted the rate of O₂ consumptionthroughout the different phases of the cultures with an r² >0.92 (n = 9, P < 0.001) using physiologically realistic Michaelisand dissociation constants. These results suggest that planktonrespiration in the field could be assessed more accurately thanbefore by measuring the intracellular ETS substrates (NADH andNADPH), in addition to ETS activity, in plankton.     

Ferricyanide-induced absorbance change of carotenoid in chromatophores of Rhodopseudomonas spheroides correlated to the oxidation of bacteriochlorophyll 885

Addition of high concentrations (e.g., 1–100 mM) of ferricyanideto a chromatophorc suspension of Rhodopseudomonas spheroidescaused a change in the absorption spectrum of carotenoid (spheroidene),which was completely reversed by adding reducing reagents suchas ferrocyanide and ascorbate. The spectral change is representedby a shift in the absorption spectrum of carotenoid by 2 to2.5 nm towards the longer wavelength side. The presence of piericidinA, o-phenanthroline or Cl-CCP in the reaction mixture did notaffect the ferricyanide-induced absorbance change. Triton X-100markedly suppressed the magnitude of the change. The additionof ferricyanide also caused simultaneous absorbance changeswith maxima at 590 and 885 nm. These are ascribed to oxidationof the (bulk) bacteriochlorophyll, BChl 885. There was no absorptionchange at other peaks of bacteriochlorophyll in the infraredregion (i.e., 800 and 855 nm). Therefore, the ferricyanide-inducedabsorbance change of carotenoid did not represent an oxidation-reductionreaction of carotenoid but was intimately correlated with oxidationof BChl 885 in the chromatophores, as judged from similaritiesobserved with respect to the time course patterns, midpointpotential (545–555 mv) in the ferriferrocyanide reactionsystem, as well as behavior towards various reagents and inhibitorsadded. A similar change of carotenoid (i.e., 2–2.5 nmshift of absorption spectrum) was caused by addition of MgCl₂to the chromatophores, but this did not induce any change inthe absorption spectrum of bacteriochlorophyll. The nature ofthe spectral change of carotenoid in chromatophores is discussed. (Received April 16, 1970; )     

Decreased affinity for oxygen of cytochrome-c oxidase in Leigh syndrome caused by SURF1 mutations

Mutations in the gene SURF1 prevent synthesis of cytochrome-c oxidase (COX)-specific assembly protein and result in a fatal neurological disorder, Leigh syndrome. Because this severe COX deficiency presents with barely detectable changes of cellular respiratory rates under normoxic conditions, we analyzed the respiratory response to low oxygen in cultured fibroblasts harboring SURF1 mutations with high-resolution respirometry. The oxygen kinetics was quantified by the partial pressure of oxygen (PO₂) at half-maximal respiration rate (P₅₀) in intact coupled cells and in digitonin-permeabilized uncoupled cells. In both cases, the P₅₀ in patients was elevated 2.1- and 3.3-fold, respectively, indicating decreased affinity of COX for oxygen. These results suggest that at physiologically low intracellular PO₂, the depressed oxygen affinity may lead in vivo to limitations of respiration, resulting in impaired energy provision in Leigh syndrome patients. oxygen kinetics; mitochondrial disease     

Factors Affecting the Growth of Daughter Bulbs in the Tulip

The relative growth rate (R_w) of daughter bulbs of the tulipcultivar Rose Copland was remarkably constant during the springperiod of growth in four seasons at two sites. The Q₁₀ of R_wwas 2.2, and the long period of constant R_w is attributed tocompensation of a fall with age by increasing temperatures inthe spring. Final bulb weight differed among four cultivarsbecause of differences in R_w and in initial daughter bulb weights. Partial defoliation reduced R_w roughly in proportion to theleaf area removed, and removal of mother bulb scales resultedin reduced leaf area, fewer daughter bulbs, and a lower daughterbulb R_w. Heat-treatment of mother bulbs before planting (blindstoken)killed the flower within the bulb, inactivated the apical dominanceexerted by the flower, and resulted in a higher initial daughter-bulbweight at the start of the spring period of exponential growth.The R_w of heat-treated and control daughter bulbs were not different,neither were the leaf areas, so it is assumed that final daughter-bulbweights were higher following treatment because of increasedsink strength. The economic implications of these findings arediscussed.     

Asparagusic acid, a new plant growth inhibitor in etiolated young asparagus shoots

Yellow prisms of asparagusic acid, with a molecular formulaof C₄H₆O₂S₂ were isolated from etiolated asparagus tissues (Asparagusofficinalis L.). This acid inhibits growth in lettuce and otherseedlings when applied in concentrations of 6.67x10^–7Mto 6.67xl0^–7M. The extent of activity was very similarto that of abscisic acid. ¹ A well known shift reagent in the NMR spectrum (1). (Received April 12, 1972; )     

Respiratory rates in the cladoceran Ceriodaphnia dubia in Lake Rotongaio, a monomictic lake

The experimentally measured oxygen consumption rate by the cladoceran,Ceriodaphnia dubia, showed a linear increase between 5 and 20°C.Oxygen consumption rates of C. dubia were estimated in situfrom respiratory electron transport system (ETS) activity inLake Rotongaio during summer stratification and winter mixing.Oxygen consumption was 0.002 µl O₂ animal^–1 h^–1in the hypolimnion and 0.076 µl O₂ animal^–1 h^–1in the epilimnion during stratification. Implications of respiredoxygen for metabolic carbon requirements are discussed.     

Increases in Cellular Levels of Cytochrome cd1 in Roseobacter denitrificans upon Irradiation with Green Light during Aerobic Growth

The cellular level of cytochrome cd₁, the nitrite reductaseof the aerobic photosynthetic bacterium Roseobacter denitrificans,increased considerably when the cells were grown aerobicallyunder white light. The action spectrum for the increase, determinedboth spectroscopically and immunologically, revealed that greenlight at 561 nm was most effective, while blue light between400 and 500 nm was fairly effective. Red and far-red light (650–900nm) absorbed by the bacterio-chlorophyll had no effect, eventhough bacteriochlorophyll and carotenoids were formed normallyduring the growth of cells. Diphenylamine, an inhibitor of thebiosynthesis of carotenoids abolished the increase in levelsof the cytochrome, a result that suggests that a carotenoid(s)was responsible for this phenomenon. The bulk carotenoids seem,however, to be unlikely the candidates for the photoreceptorsbecause they did not accumulate in the light-grown cells. Attemptsto detect archaerhodopsin, 11-cis and all-trans retinal by immunologicalor HPLC analysis were unsuccessful. Although we failed to identifythe photoreceptor, it is clear that R. denitrificans has a green-lightsignal-transduction system that controls the expression of cytochromecd₁. (Received April 19, 1993; Accepted July 12, 1993)     

A carrier-mediated mechanism for pyridoxine uptake by human intestinal epithelial Caco-2 cells: regulation by a PKA-mediated pathway

Vitamin B₆ is essential for cellular functions and growth due to its involvement in important metabolic reactions. Humans and other mammals cannot synthesize vitamin B₆ and thus must obtain this micronutrient from exogenous sources via intestinal absorption. The intestine, therefore, plays a central role in maintaining and regulating normal vitamin B₆ homeostasis. Due to the water-soluble nature of vitamin B₆ and the demonstration that transport of other water-soluble vitamins in intestinal epithelial cells involves specialized carrier-mediated mechanisms, we hypothesized that transport of vitamin B₆ in these cells is also carrier mediated in nature. To test this hypothesis, we examined pyridoxine transport in a model system for human enterocytes, the human-derived intestinal epithelial Caco-2 cells. The results showed pyridoxine uptake to be 1) linear with time for up to 10 min of incubation and to occur with minimal metabolic alteration in the transported substrate, 2) temperature and energy dependent but Na⁺ independent, 3) pH dependent with higher uptake at acidic compared with alkaline pHs, 4) saturable as a function of concentration (at buffer pH 5.5 but not 7.4) with an apparent Michaelis-Menten constant (K_m) of 11.99 ± 1.41 µM and a maximal velocity (V_max) of 67.63 ± 3.87 pmol · mg protein^-1 · 3 min^-1, 5) inhibited by pyridoxine structural analogs (at buffer pH 5.5 but not 7.4) but not by unrelated compounds, and 6) inhibited in a competitive manner by amiloride with an apparent inhibitor constant (K_i) of 0.39 mM. We also examined the possible regulation of pyridoxine uptake by specific intracellular regulatory pathways. The results showed that whereas modulators of PKC, Ca⁺²/calmodulin (CaM), and nitric oxide (NO)-mediated pathways had no effect on pyridoxine uptake, modulators of PKA-mediated pathway were found to cause significant reduction in pyridoxine uptake. This reduction was mediated via a significant inhibition in the V_max, but not the apparent K_m, of the pyridoxine uptake process. These results demonstrate, for the first time, the involvement of a specialized carrier-mediated mechanism for pyridoxine uptake by intestinal epithelial cells. This system is pH dependent and amiloride sensitive and appears to be under the regulation of an intracellular PKA-mediated pathway. vitamin B₆; intestinal transport; transport regulation; Caco-2 cell