Na+-dependent Ca2+ uptake in isolated opercular epithelium of Fundulus heteroclitus

It is concluded that Ca²⁺ transport across the basolateral membranes of the ionocytes in killifish skin is mediated for the major part by a Na⁺/Ca²⁺-exchange mechanism that is driven by the (transmembrane) Na⁺ gradient established by Na⁺/K⁺-ATPase. The conclusion is based, firstly, on the biochemical evidence for the presence of a Na⁺/Ca²⁺-exchanger next to the Ca²⁺-ATPase in the basolateral membranes of killifish gill cells. Secondly, the transcellular Ca²⁺ uptake measured in an Ussing chamber setup was 85% and 80% reduced in freshwater (FW) and SW (SW) opercular membranes, respectively, as the Na⁺ gradient across the basolateral membrane was directly or indirectly (by ouabain) reduced. Thapsigargin or dibutyryl-cAMP/IBMX in SW opercular membranes reduced Ca²⁺ influx to 46%, comparable to the effects seen in FW membranes [reduction to 56%; Marshall et al. 1995a]. Basal Ca²⁺ influx across the opercular membrane was 48% lower in membranes from fish adapted to SW than in membranes from fish adaptated to FW. Branchial Na⁺/K⁺-ATPase activity was two times higher in SW adapted fish. Accepted: 29 October 1996     

Unidirectional Na(+) and Ca (2+) fluxes in two euryhaline teleost fishes, Fundulus heteroclitus and Oncorhynchus mykiss, acutely submitted to a progressive salinity increase

Na⁺ and Ca²⁺ regulation were compared in two euryhaline species, killifish (normally estuarine-resident) and rainbow trout (normally freshwater-resident) during an incremental salinity increase. Whole-body unidirectional fluxes of Na⁺ and Ca²⁺, whole body Na⁺ and Ca²⁺, and plasma concentrations (trout only), were measured over 1-h periods throughout a total 6-h protocol of increasing salinity meant to simulate a natural tidal flow. Killifish exhibited significant increases in both Na⁺ influx and efflux rates, with efflux slightly lagging behind efflux up to 60% SW, but net Na⁺ balance was restored by the time killifish reached 100% SW. Whole body Na⁺ did not change, in agreement with the capacity of this species to tolerate daily salinity fluctuations in its natural habitat. In contrast, rainbow trout experienced a dramatic increase in Na⁺ influx (50-fold relative to FW values), but not Na⁺ efflux between 40 and 60% SW, resulting in a large net loading of Na⁺ at higher salinities (60–100% SW), and increases in plasma Na⁺ and whole body Na⁺ at 100% SW. Killifish were in negative Ca²⁺ balance at all salinities, whereas trout were in positive Ca²⁺ balance throughout. Ca²⁺ influx rate increased two- to threefold in killifish at 80 and 100% SW, but there were no concomitant changes in Ca²⁺ efflux. Ca²⁺ flux rates were affected to a larger degree in trout, with twofold increases in Ca²⁺ influx at 40% SW and sevenfold increases at 100% SW. Again, there was no change in Ca²⁺ efflux with salinity, so plasma Ca²⁺ concentration increased in 100% SW. As the killifish is regularly submitted to increased salinity in its natural environment, it is able to rapidly activate changes in unidirectional fluxes in order to ensure ionic homeostasis, in contrast to the trout.     

Histochemical and biochemical studies of carbonic anhydrase activity in the opercular epithelium of the euryhaline teleost, Fundulus heteroclitus

Carbonic anhydrase (CAH) activity was biochemically measured and histochemically localized (at both the light and electron microscope levels) in isolated opercular membranes from teleost fish, Fundulus heteroclitus, adapted to freshwater (FW), seawater (SW), and double-strength seawater (2 x SW). The normal morphology of this membrane showed that its epithelial portion consisted of five cell types: (1) chloride cells, which have been previously implicated as responsible for the active chloride transport across the epithelium; (2) mucous cells; (3) pavement cells, which formed the major portion of the free epithelial surface; (4) supportive cells, which had an abundance of intermediate (10 nm)-type filaments suggesting a structural role for these cells; and (5) vesicular cells, which were characterized by various types of membrane-bound vesicles, including lysosomes, and numerous free ribosomes. Vesicular cells may be stem cells and/or endocrine cells. Hansson's histochemical method for CAH revealed cobalt sulfide reaction product confined to the following structures in fish from each environment: (1) chloride cells: throughout the cytoplasm and some nuclear staining; (2) mucous cells: throughout the cytoplasm, some nuclear staining, and some in mucous granules; (3) vesicular cells: confined to lysosomes, some of the vesicles, and nucleoli; (4) a small portion of the intracellular space between adjacent vesicular cells and supportive cells; and (5) supportive cells: in nucleoli and occasionally in larger membrane-bound lysosomelike structures. Acetazolamide (10(-5) M) and potassium cyanate (KCNO) (10(-1) M) in Hansson's incubation medium completely inhibited the formation of reaction product. Biochemical determination of CAH activity on vascularly perfused, isolated opercular membranes showed no statistically significant difference in enzyme activity between environmental groups. The following units of activity/mg opercular membrane protein were measured: FW: 0.63 +/- 0.02; SW: 0.43 +/- 0.08; 2 x SW: 0.64 +/- 0.09.     

Ultrastructure of the pineal organ of the killifish,Fundulus heteroclitus,with special reference to the secretory function

Summary The pineal organ of the killifish, Fundulus heteroclitus, was investigated by electron microscopy under experimental conditions; its general and characteristic features are discussed with respect to the photosensory and secretory function. The strongly convoluted pineal epithelium is usually composed of photoreceptor, ganglion and supporting cells. In addition to the well-differentiated photosensory apparatus, the photoreceptor cell contains presumably immature dense-cored vesicles (140–220 nm in diameter) associated with a well-developed granular endoplasmic reticulum in the perinuclear region and the basal process. These dense-cored vesicles appear rather prominent in fish subjected to darkness. The ganglion cell shows the typical features of a nerve cell; granular endoplasmic reticulum, polysomes, mitochondria and Golgi apparatus are scattered in the electron-lucent cytoplasm around the spherical or oval nucleus. The dendrites of these cells divide into smaller branches and form many sensory synapses with the photoreceptor basal processes. Lipid droplets appear exclusively in the supporting cell, which also contains well-developed granular endoplasmic reticulum and Golgi apparatus. Cytoplasmic protrusions filled with compact dense-cored vesicles (90–220 nm in diameter) are found in dark-adapted fish. The origin of these cytoplasmic protrusions, however, remains unresolved. Thus, the pineal organ of the killifish contains two types of dense-cored vesicles which appear predominantly in darkness. The ultrastructural results suggest that the pineal organ of fish functions not only as a photoreceptor but also as a secretory organ.We thank Dr. Grace Pickford for the fishes.     

Specific inhibitors of intracellular Ca2+ transport ATPases

Support from the National Institutes of Health and the American Heart Association is gratefully acknowledged.     

Ca2+ transport in mitochondrial and microsomal fractions from higher plants

Mitochondria from etiolated corn possess a much greater Ca²⁺ uptake capacity per mg protein than microsomes from the same source. Differences in energy requirements, sensitivity to specific inhibitors, and sedimentation properties enabled us to study both Ca²⁺ uptake mechanisms without mutual contamination. The microsomal Ca²⁺ uptake does not vary much among different plants as compared to the mitochondrial Ca²⁺ uptake; this is also true for different organs of the same plant. Mitochondrial Ca²⁺ uptake is more dependent on the age of the seedlings than microsomal uptake, because of changes in active Ca²⁺ uptake activity rather than of changes in efflux. Intactness and the oxidative and phosphorylative properties of the mitochondria remained unchanged during this time period. Na⁺ and Mg²⁺ do not induce Ca²⁺ release from mitochondria.Abbreviations ATP adenosine triphosphate - ADP adenosine diphosphate - NADH₂ -nicotinamide adenin dinucleotide, reduced form - Mops 3-(N-morpholino)propane-sulfonic acid - Tris tris-(hydroxymethyl)-aminomethane - Hepes hydroxyethylpiperazine-N-2-ethanesulfonic acid - BSA bovine serum albumin - EDTA (ethylene-dinitrilo)-tetraacetic acid - EGTA ethylene glycol-bis(-aminoethylether)-N,N-tetraacetic acid - CCCP carbonyl cyanide m-chlorophenylhydrazone - DTE 1,4-dithiothreitol     

The effect of prolactin on kidney structure of the euryhaline teleost Gasterosteus aculeatus during adaptation to fresh water

Summary The effect of injections of ovine prolactin on kidney structure was examined in the first 10 days following transfer of seawater sticklebacks to fresh water.In hormone injected animals as well as in controls the glomeruli increase slightly in size after transfer. The podocytes intensify the secretion of mucopolysaccharides, which is indicative of increased turnover of the components of the glomerular basal lamina. The nuclei of the podocytes become enlarged, while those of the juxtaglomerular cells decrease in size. These changes are related to the well known rise of the glomerular filtration rate following transfer to fresh water. Structural indications that prolactin is involved in the control of glomerular filtration were not found.The epithelial cells of the three nephronic segments and of the ureter become considerably better developed after transfer to fresh water. Cell height, nuclear and mitochondrial volume, and surface of the membranes of the basal labyrinth increase in all tubular epithelia, although not to the same extent. Increases are moderate in the first proximal segment, but increasingly higher for the second proximal segment, collecting duct and the ureter. Especially the growth of membrane surface of the basal labyrinth, site of ion transport mechanisms, is impressive. In controls, values characteristic for freshwater fishes are reached in 6 to 9 days for all parameters for cellular development. Prolactin injections greatly stimulate growth rates in all tubular epithelia: freshwater values are reached within 3 days. No further increase was found, however.     

Ca2+ transport and chemoreception in Paramecium

Intracellular Ca²⁺ levels in Paramecium must be tightly controlled, yet little is understood about the mechanisms of control. We describe here indirect evidence that a phosphoenzyme intermediate is the calmodulin-regulated plasma membrane Ca²⁺ pump and that a Ca²⁺-ATPase activity in pellicles (the complex of cell body surface membranes) is the enzyme correlate of the plasma membrane pump protein. A change in Ca²⁺ pump activity has been implicated in the chemoresponse of paramecia to some attractant stimuli. Indirect support for this is demonstrated using mutants with different modifications of calmodulin to correlate defects in chemoresponse with altered Ca²⁺ homeostasis and pump activity.Abbreviations EGTA ethyleneglycol tetra-acetate - ER endoplasmic reticulum - IBMX isobutyl methylxanthine - I _che index of chemokinesis - Mops 3-[N-morpholino] propanesulfonic acid - PEI phosphoenzyme intermediate - STEN sucrose, TRIS, EDTA, sodium chloride - TCA trichloroacetic acid - TRIS tris[hydroxymethyl] aminomethane     

Gibberellic-acid-stimulated Ca2+ accumulation in endoplasmic reticulum of barley aleurone: Ca2+ transport and steady-state levels

The steady-state levels of Ca²⁺ within the endoplasmic reticulum (ER) and the transport of ⁴⁵Ca²⁺ into isolated ER of barley (Hordeum vulgare L. cv. Himalaya) aleurone layers were studied. The Ca²⁺-sensitive dye indo-1. Endoplasmic reticulum was isolated and purified from indo-1-loaded protoplasts, and the Ca²⁺ level in the ER was measured using the Ca²⁺-sensitive dye indo-1. Endoplasmic reticulum was isolated and purified from indo-1-loaded protoplasts, and the Ca²⁺ level in the lumen of the ER was determined by the fluorescence-ratio method to be at least 3 M. Transport of ⁴⁵Ca²⁺ into the ER was studied in microsomal fractions isolated from aleurone layers incubated in the presence and absence of gibberellic acid (GA₃) and Ca²⁺. Isopycinic sucrose density gradient centrifugation of microsomal fractions isolated from aleurone layers or protoplasts separates ER from tonoplast and plasma membranes but not from the Golgi apparatus. Transport of ⁴⁵Ca²⁺ occurs primarily in the microsomal fraction enriched in ER and Golgi. Using monensin and heat-shock treatments to discriminate between uptake into the ER and Golgi, we established that ⁴⁵Ca²⁺ transport was into the ER. The sensitivity of ⁴⁵Ca²⁺ transport to inhibitors and the K_m of ⁴⁵Ca²⁺ uptake for ATP and Ca²⁺ transport in the microsomal fraction of barley aleurone cells. The rate of ⁴⁵Ca²⁺ transport is stimulated several-fold by treatment with GA₃. This effect of GA₃ is mediated principally by an effect on the activity of the Ca²⁺ transporter rather than on the amount of ER.Abbreviations CCR cytochrome-c reductase - DCCD dicyclohexylcarbodiimide - EGTA ethylene glycol bis(-aminoethyl ether)-N,N,N,N-tetraacetic acid - ER endoplasmic reticulum - FCCP carbonylcyanide p-trifluoromethoxyphenyl hydrazone - GA₃ gibberellic acid - IDPase inosine diphosphatase - Mon monensin     

Chloride transport activation by plasma osmolarity during rapid adaptation to high salinity of Fundulus heteroclitus

Transition from low salt water to sea water of the euryhaline fish, Fundulus heteroclitus, involves a rapid signal that induces salt secretion by the gill chloride cells. An increase of 65 mOsm in plasma osmolarity was found during the transition. The isolated, chloridecell-rich opercular epithelium of sea-water-adapted Fundulus exposed to 50 mOsm mannitol on the basolateral side showed a 100% increase in chloride secretion, which was inhibited by bumetanide 10^–4 m and 10^–4 m DPC (N-Phenylanthranilic acid). No effect of these drugs was found on apical side exposure. A Na⁺/H⁺ exchanger, demonstrated by NH₄Cl exposure, was inhibited by amiloride and its analogues and stimulated by IBMX, phorbol esters, and epithelial growth factor (EGF). Inhibition of the Na⁺/H⁺ exchanger blocks the chloride secretion increase due to basolateral hypertonicity. A Cl^–/HCO ₃ ^– exchanger was also found in the chloride cells, inhibited by 10^–4 m DIDS but not involved in the hyperosmotic response. Ca²⁺ concentration in the medium was critical for the stimulation of Cl^– secretion to occur. Chloride cell volume shrinks in response to hypertonicity of the basolateral side in sea-water-adapted operculi; no effect was found on the apical side. Freshwater-adapted fish chloride cells show increased water permeability of the apical side. It is concluded that the rapid signal for adaptation to higher salinities is an increased tonicity of the plasma that induces chloride cell shrinkage, increased chloride secretion with activation of the Na⁺K⁺2Cl^– cotransporter, the Na⁺/H⁺ exchanger and opening of Cl^– channels.The work was supported by the National Institutes of Health, Research Grant EYO1340 to J.A.Z. Part of this research was performed while Dr. Zadunaisky was a Scholar In Residence at the Fogarty International Center of The National Institutes of Health in Bethesda, Maryland. Ms. Dawn Roberts was a fellow of the Grass Foundation and Pew Foundation during this work. Grants from the National Science Foundation and the National Institutes of Health to the Mount Desert Island Biological Laboratory also provided assistance for this research.     

NaCl transport across the opercular epithelium of Fundulus heteroclitus is inhibited by an endothelin to NO, superoxide, and prostanoid signaling axis

Recent evidence suggests that paracrine signaling agents, such as endothelin (ET), nitric oxide (NO), superoxide (O2-), and prostanoids can modulate mammalian renal function by affecting both hemodynamic and epithelial ionic transport pathways. Since these signaling pathways have been described in fish blood vessels, we hypothesized that they may control salt transport across the gill epithelium--the primary site of ion excretion in marine teleost fishes. We found that ET, the NO donors sodium nitroprusside and spermine NONOate, and the prostanoid PGE2 each can produce a concentration-dependent reduction in the short circuit current (Isc) across the isolated opercular epithelium of the killifish (Fundulus heteroclitus), the generally accepted model for the marine teleost gill epithelium. Sarafotoxin S6c was equipotent to ET-1, suggesting that ETB receptors are involved. Incubation with NG-nitro-L-arginine methyl ester (L-NAME) or indomethacin reduced the effect of subsequent addition of SRXS6c by 17 and 89%, respectively, suggesting the presence of an ET to NO and PGE axis. The effects of l-NAME and indomethacin were not additive, but the superoxide dismutase mimetic 4-hydroxy-2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPOL) reduced the effect of SRXS6c by 34% and preincubation with l-NAME, indomethacin, and TEMPOL reduced the SRXS6c response to zero. This suggests a direct role for O2- in this axis. COX-2 appears to be the major enzyme involved in this axis because the specific COX-2 inhibitor NS-398 was twice as effective as the COX-1 inhibitor SC560 in inhibiting the SRXS6c effect. The Isc was stimulated by the EP2 agonist butaprost and inhibited by the EP(1,3) agonist sulprostone, suggesting both stimulatory and inhibitory PGE receptors in this tissue. Carbaprostacyclin (PGI2 analog), thromboxane A2, PGF(2alpha), and PGD2 did not affect the Isc. Our data are the first to suggest the importance of an ET-stimulated and NO-, O2(-)-, and PGE2-mediated signaling axis that can modify active extrusion of NaCl across the killifish opercular epithelium and, by inference, the marine teleost gill epithelium.     

Turgor regulation and cytoplasmic free Ca2+ in the algaLamprothamnium

Summary In internodal cells ofLamprothamnium succinctum, turgor regulation in response to hypotonie treatment is inhibited by lowering external Ca²⁺ concentration ([Ca²⁺]_e) from 3.9 (normal) to 0.01 (low) mM. In order to clarify whether a change in the cytoplasmic free Ca²⁺ concentration ([Ca²⁺]_c) is involved in turgor regulation, the Ca²⁺ sensitive protein aequorin was injected into the cytoplasm of internodal cells. A large transient light emission was observed upon hypotonic treatment under normal [Ca²⁺]_e but not under low [Ca²⁺]_e. Thus hypotonic treatment induces a transient increase in [Ca²⁺]_c under normal [Ca²⁺]_e but not under low [Ca²⁺]_e.Abbreviations ASW artificial sea water - _i cellular osmotic pressure - [Ca²⁺]_c cytoplasmic free Ca²⁺ concentration - EDTA ethylenediamine-tetraacetic acid - EGTA ethylenglycol-bis(-aminoethyl ether(N,N-tetraacetic acid - [Ca²⁺]_e external Ca²⁺ concentration - _e external osmotic pressure - GM glass micropipette - GP glass plate - HEPES N-2-hydroxyethylpiperazine-N-2-ethansulfonic acid - MS microscope stage - OL objective lens - PIPES piperazine-N-N-bis(2-ethanesulfonic acid) - W Weight     

Ca+2 transport across intestinal brush border membranes of the cichlid teleost Oreochromis mossambicus

Summary Brush border membranes were isolated from tilapia (Oreochromis mossambicus) intestine by the use of magnesium precipitation and differential centrifugation. The membrane preparation was enriched 17-fold in alkaline phosphatase. The membranes were 99% right-side-out oriented as indicated by the unmasking of latent glyceraldehyde-3-phosphate dehydrogenase and acetylcholine esterase activity by detergent treatment. The transport of Ca⁺² in brush border membrane vesicles was analyzed. A saturable and a nonsaturable component in the uptake of Ca⁺² was resolved. The saturable component is characterized by a K _m much lower than the Ca⁺² concentrations predicted to occur in the intestinal lumen. The nonsaturable component displays a Ca⁺² permeability too high to be explained by simple diffusion. We discuss the role of the saturable component as the rate-limiting step in transmembrane Ca⁺² movement, and suggest that the nonsaturable component reflects a transport mechanism operating well below its level of saturation.The authors wish to thank Tom Spanings for his superb organization of fish husbandry, and Maarten de Jong (Dept. of Physiology, Faculty of Medicine, University of Nijmegen) for making the automated stopped-flow apparatus available to us.     

Analytical and experimental studies on the relationship between Na+, K+, and water uptake during volume increases associated with Fundulus oocyte maturation in vitro

Summary Oocytes of marine and estuarine teleosts often undergo pronounced volume increases during the maturation phase of development that precedes ovulation and fertilization. To examine the physiological correlates of these volume increases, prematuration follicles of the saltmarsh teleost, Fundulus heteroclitus, were cultured in vitro with a maturation-inducing steroid (17-hydroxy-20-dihydroprogesterone). Mean follicle volume rose significantly (75%) during a 40-h incubation period. Similar to the situation previously found in vivo, uptake of water by the maturing follicle was responsible for this volume increase in vitro, with the water content increasing from 62% to 78% of the total follicle mass. The follicle contents of two probable osmotic effectors-Na⁺ and K⁺-also rose, the increase in K⁺ being twice that of Na⁺. The influx of K⁺ even exceeded water uptake, resulting in a net increase in the concentration of this cation. It thus appears that the influx of these cations, in particular K⁺, is a major cause of the uptake of osmotically obligated water and subsequent volume increase experienced by maturing F. heteroclitus follicles. In a search for operant mechanisms, it was found that follicle hydration, but not maturation, was strictly dependent on external K⁺ in a concentration-dependent manner. The mechanism by which K⁺ accumulates in the follicle was insensitive to ouabain, so that a typical Na⁺, K⁺-ATPase mechanism does not appear to be involved. The ability of external K⁺ to promote follicle hydration was gradually lost during the maturation process as the oocyte dissociated from the surrounding granulosa cells in preparation for ovulation. Removal of all associated somatic cells prior to maturation prevented subsequent steroid-initiated hydration but not maturation. The results suggest that K⁺ may be translocated from surrounding granulosa cells to the oocyte via gap junctions during maturation.Abbreviations GVBD germinal vesicle breakdown     

Ca2+ regulation of ion transport in the na+/ca2+ exchanger

The binding of Ca(2+) to two adjacent Ca(2+)-binding domains, CBD1 and CBD2, regulates ion transport in the Na(+)/Ca(2+) exchanger. As sensors for intracellular Ca(2+), the CBDs form electrostatic switches that induce the conformational changes required to initiate and sustain Na(+)/Ca(2+) exchange. Depending on the presence of a few key residues in the Ca(2+)-binding sites, zero to four Ca(2+) ions can bind with affinities between 0.1 to 20 μm. Importantly, variability in CBD2 as a consequence of alternative splicing modulates not only the number and affinities of the Ca(2+)-binding sites in CBD2 but also the Ca(2+) affinities in CBD1.     

The role of the plasma-membrane Ca2+-ATPase in Ca2+ homeostasis in Sinapis alba root hairs

The regulation of cytosolic Ca²⁺ has been investigated in growing root-hair cells of Sinapis alba L. with special emphasis on the role of the plasmamembrane Ca²⁺-ATPase. For this purpose, erythrosin B was used to inhibit the Ca²⁺-ATPase, and the Ca²⁺ ionophore A₂₃₁₈₇ was applied to manipulate cytosolic free [Ca²⁺] which was then measured with Ca²⁺-selective microelectrodes. (i) At 0.01 M, A₂₃₁₈₇ had no effect on the membrane potential but enhanced the Ca²⁺ permeability of the plasma membrane. Higher concentrations of this ionophore strongly depolarized the cells, also in the presence of cyanide. (ii) Unexpectedly, A₂₃₁₈₇ first caused a decrease in cytosolic Ca²⁺ by 0.2 to 0.3 pCa units and a cytosolic acidification by about 0.5 pH units, (iii) The depletion of cytosolic free Ca²⁺ spontaneously reversed and became an increase, a process which strongly depended on the external Ca²⁺ concentration, (iv) Upon removal of A₂₃₁₈₇, the cytosolic free [Ca²⁺] returned to its steady-state level, a process which was inhibited by erythrosin B. We suggest that the first reaction to the intruding Ca²⁺ is an activation of Ca²⁺ transporters (e.g. ATPases at the endoplasmic reticulum and the plasma membrane) which rapidly remove Ca²⁺ from the cytosol. The two observations that after the addition of A₂₃₁₈₇, (i) Ca²⁺ gradients as steep as-600 mV could be maintained and (ii) the cytosolic pH rapidly and immediately decreased without recovery indicate that the Ca²⁺-exporting plasma-membrane ATPase is physiologically connected to the electrochemical pH gradient, and probably works as an nH⁺/Ca²⁺-ATPase. Based on the finding that the Ca²⁺-ATPase inhibitor erythrosin B had no effect on cytosolic Ca²⁺, but caused a strong Ca²⁺ increase after the addion of A₂₃₁₈₇ we conclude that these cells, at least in the short term, have enough metabolic energy to balance the loss in transport activity caused by inhibition of the primary Ca²⁺-pump. We further conclude that this ATPase is a major Ca²⁺ regulator in stress situations where the cytosolic Ca²⁺ has been shifted from its steady-state level, as may be the case during processes of signal transduction.Abbreviations and Symbols EB erythrosin B - E_m membrane potential - pCa negative logarithm of the Ca²⁺ concentration This work was supported by the Deutche Forschungsgemeinschaft (H.F.) and the Alexander-von-Humboldt-Foundation (A.T.).     

Anti-microtubular herbicides and fungicides affect Ca2+ transport in plant mitochondria

The herbicides amiprophosmethyl (APM) trifluralin, and oryzalin as well as the fungicides methylbenzimidazolyl carbamate (MBC), O-isopropyl N-phenyl carbamate (IPC), and chlorisopropyl N-phenyl carbamate (CIPC), which are known to cause the destruction of microtubules in vivo but do not interfere with tubulin polymerization in vitro, have been examined with respect to their ability to affect Ca²⁺ transport in isolated cell organelles. In contrast to colchicine which has no effect on Ca²⁺ transport in isolated mitochondrial and microsomal fractions, all of the substances investigated caused considerable reduction of ca²⁺ net uptake into mitochondrial but not into microsomal fractions. This reduction has been shown to be due to an increase in passive Ca²⁺ efflux. These results have been extrapolated to in vivo situations where they are postulated to act by raising cytoplasmic Ca²⁺ levels.Abbreviations APM amiprophosmethyl - CIPC chlorisopropyl N-phenyl carbamate - IPC O-isopropyl N-phenyl carbamate - MBC methylbenzimidazolyl carbamate - Mops 3-(N-Morpholino) propanesulfonic acid - DMSO dimethylsulfoxide