μM AlCl3 Research Articles

Kinetics of Aluminum Uptake by Cell Suspensions of Phaseolus vulgaris L.

Summary The Kinetics of aluminum (Al) uptake by cell suspensions derived from an Al-resistant (Dade) an an Al-sensitive (Romano) cultivar of Phaseolus vulgaris L. were investigated. Uptake of Al from low-volume (3-mL) uptake solutions containing 75 μM AlCl 3 was rapid during the first 20 min with little additional absorption occurring over the remainder of the 180 min experimental period. In contrast studies with excised roots showed a longer rapid phase (30 min) which was followed by a linear phase of uptake. Differences in Al uptake between excised roots and cell suspensions appear to reflect the unique characteristics of our low-volume cell system rather than real differences in uptake between the two systems. The rate, extent, and saturable nature of uptake in this cell system suggested that depletion of Al from uptake solutions may have been responsible for the lacf of an observable linear phase of uptake. However, when the concentration of Al in uptake solutions was increased to 500 and 1000 μM, total accumulation of Al increased, while the general pattern of uptake was not affected. Moreover, mock-uptake experiments at these high concentrations ruled out the possibility of artifacts arising from formation of solid phase phase Al. While increasing the concentration of Al in uptake solutions provided a means of increasing Al available for uptake, Pyrocatechol violet analysis of monomeric Al indicate incomplete recovery of added Al particularly at concentrations above 300 μM. To overcome potential problems associate with the formation of complex Al species in solution, the total amount of Al in solution was increased using a high-volume (100-mL), low-concentration (75-μM AlC1 3 ) system. Under these conditions, a biphasic pattern of uptake was observed, with a rapid phase of uptake during the first 20 min, and a linear phase of uptake over the remainder of the 180 min uptake period. We were also able to isolate the linear phase of uptake after desorption in 9.0 mM citric acid. These results suggest that the basic pattern of Al uptake by cell suspensions (saturated vs. linear) is strongly affected by the speciation of Al, which is indirectly related to the volume of uptake solutions. The biphasic pattern of uptake observed in our high-volume system also supports the hypothesis that the kinetics of short-term Al uptake observed in excise roots reflect transport events occurring at the cellular level. We believe further investigation of the kinetics of Al uptake at the cellular level could provide a more direct means of measuring the uptake of Al across the plasma membrane in plants.

Effects of chronic dietary aluminum on local cerebral glucose utilization in rats

Beginning at 4 weeks of age normal, male, Sprague-Dawley rats were reared on Purina Laboratory Chow and drinking water containing 100 μM AlCl 3. After 2 years, local rates of cerebral glucose utilization were determined with the autoradiographic [ 14C] deoxyglucose method in the brain as a whole and in 25 brain regions in 6 treated rats and 4 age-matched controls. The results indicate that any effects of chronic aluminum in the diet on rates of cerebral glucose utilization are small. In the brain as a whole, the mean rate of glucose utilization in the aluminum-treated rats was 6% lower than that of the controls ( p = 0.09). In 21 of the 25 brain regions examined mean rates of glucose utilization were generally lower in the aluminum-treated rats but in none of the regions were the effects statistically significant.

Separation and Characterization of Two Mg 2+-ATPase Activities from the Human Erythrocyte Membrane

Two distinct Mg 2+-ATPase activities were isolated from Triton X-100-solubilized human erythrocyte membranes using a combination of calmodulin-agarose to remove Ca 2+-ATPase and ion exchange chromatography to separate the Mg 2+-ATPase activities. The activity designated Mg 2+-ATPase A was inhibited by low concentrations of vanadate (IC 50 ≍ 2 μM) and by calcium (IC 50 ≍ 1.5 mM), La 3+ (80% inhibition at 0.5 mM), and F − (IC 50 ≍ 1 mM). Inhibition by F − increased in the presence of 10 μM AlCl 3. The activity appeared to be dependent on the concentration of MgATP. Two millimolar deoxyATP supported nearly full activity and 2 mM GTP supported 40% of the activity compared to that seen with 2 mM ATP. The activity was stimicrolated twofold by the addition of 1% (w/v) phosphatidylserine. These properties are consistent with a role of this enzyme in the control of red blood cell shape, possibly through association with the ATP-dependent translocation of phosphatidylserine and phosphatidylethanolamine from the outer to the inner leaflet of the bilayer. In contrast, Mg 2+-ATPase B was slightly stimicrolated by vanadate and by calcium but was unaffected by LaCl 3, and by F − ± AlCl 3. The activity was further stimicrolated by added magnesium in excess of fixed concentrations of ATP. In addition, at a fixed concentration of 10 mM MgCl 2, the activity with increasing ATP was biphasic, with the maximicrom at 1 mM. GTP 2 mM failed to support activity while 2 mM deoxyATP supported only 30% of activity compared to that seen with 2 mM ATP. The activity was not stimicrolated by oxidized glutathione or the glutathione conjugate lithocholic acid, suggesting it is not involved in the ATP-dependent transport of such compounds out of the erythrocyte.

Comparison of the early response to aluminum stress between tolerant and sensitive wheat cultivars: Root growth, aluminum content and efflux of K+

Abstract In order to characterize the mechanism of Al tolerance (Atlas 66) and Al sensitivity (Scout 66) in two cultivars of wheat (Triticum aestivum L.), the early responses to Al stress under acidic conditions were investigated. Marked inhibition of root elongation of Scout was observed upon treatment with 10 μM AlCl3 for less than 3 h. The inhibition of root elongation of Scout was reversed within 3 days when the treated samples were transferred to a solution without Al. However, treatment for 6 h with AlCl3 repressed root elongation almost completely and irreversibly. Root elongation of Atlas was only partially inhibited by the treatment with 10 μM AlCl3 for more than 6 h. Levels of Al in two portions of roots, namely, portions 0–5 mm and 5–10 mm from the tip, were lower in Atlas than those in Scout. In Atlas the levels of Al on a fresh weight basis in both portions were very similar, while the level of Al in the portion 0–5 mm from the tip was almost double than that in the 5–10 mm portion in Scout. ...

G-proteins coupled to phosphoinositide hydrolysis in the cochlear and vestibular sensory epithelia of the rat are insensitive to cholera and pertussis toxins

In the cochlear (CSE) and vestibular sensory epithelia (VSE), phosphoinositides are hydrolyzed in response to stimulation of phospholipase C (PLC) by cholinergic muscarinic and purinergic P 2y agonists. Such receptor-mediated activation of PLC is expected to be coupled through guanine nucleotide-binding proteins (G-proteins). Although several classes of G-proteins have been identified in the inner ear, nothing is known about the type of G-proteins associated with the phosphoinositide second messenger system in CSE and VSE. Phosphoinositide hydrolysis was determined by the release of radiolabeled inositol phosphates (InsPs). Ten mM NaF plus 10 μM AlCl 3 increased basal InsPs accumulation 2-fold in both CSE and VSE of the rat. Release of InsPs was also enhanced by guanosine 5'-O-(3-thiotriphosphate) (GTP-γ-S) in saponin-permeabilized tissues. Furthermore, release of InsPs stimulated by both carbamylcholine (CCh) and adenosine 5'-O-[3-thiotriphosphate](ATP-γ-S) was significantly inhibited by 100 μM guanosine 5'-O-[2-thiodiphosphate](GDP-β-S). These results strongly suggest the involvement of G-proteins in the receptor-PLC coupling in CSE and VSE. ADP-ribosylation in membrane fractions of CSE and VSE in the presence of cholera toxin (CTX) or pertussis toxin (PTX) indicated the existence of G s- and G i-type G-proteins. However, neither CTX nor PTX affected basal or agonist-stimulated release of InsPs. These observations suggest that muscarinic and P 2y purinergic receptors are coupled to PLC via CTX- and PTX-insensitive G-proteins in CSE and VSE.

Mechanisms underlying AlCl 3 inhibition of agonist-stimulated inositol phosphate accumulation : Role of calcium, G-proteins, phospholipase C and protein kinase C

Possible mechanisms of AlCl 3-induced inhibition of agonist-stimulated inositol phosphate (IP) accumulation were investigated using rat brain cortex slices, synaptosomes or homogenates. Under conditions in which AlCl 3 inhibits carbachol (CARB)-stimulated IP accumulation (G p-mediated), AlCl 3 did not affect CARB (100 μM)-induced decreases (G i-mediated) in 30 μM forskolin-stimulated cAMP accumulation, suggesting that AlCl 3 may be specific for G p-mediated signal transduction. To determine whether AlCl 3 interfered with G p function and/or phosphatidylinositol-specific phospholipase C (PiPLC) activity, effects of AlCl 3 on CARB- and Ca 2+-stimulated IP accumulation were examined in cortical synaptosomes. AlCl 3 (500 μM) decreased CARB (1 mM)- and Ca 2+ (20 μM ionomycin)-stimulated IP accumulation to 77 and 75% of control, respectively, suggesting that AlCl 3 may not directly affect G p activity, but does inhibit PiPLC activity. In cortical homogenates, AlCl 3 (10–500 μM) inhibited hydrolysis of [ 3H]phosphatidylinositol 4,5-bisphosphate (PIP 2) by PiPLC in a concentration-dependent manner with an estimated IC 50 of 100 μM. The effects of AlCl 3 on modulation of IP accumulation by extracellular Ca 2+ and PKC were also examined as potential mechanisms. Decreasing the extracellular Ca 2+ concentration ([Ca 2+] e) from 1.0 to 0.1 mM decreased CARB-stimulated IP accumulation in slices. AlCl 3 (500 μM) decreased significantly 1 mM CARB-stimulated IP accumulation in 1.0 and 0.1 mM Ca 2+ solutions; however, the effect of AlCl 3 on IP accumulation did not depend on [Ca 2+] e. In cortical slices, inhibition of 1 mM CARB-stimulated IP accumulation by 500 μM AlCl 3 was not altered by the PKC activator phorbol 12,13-dibutyrate (PdBu, 1 μM), or the PKC inhibitor H-7 (10 μM), suggesting that AlCl 3 does not interfere with IP accumulation by activation of PKC. Other studies found that AlCl 3 (10–100 μM) inhibited PKC activity in a concentration-dependent manner in both cytosolic and membrane fractions of cortical homogenates with an estimated IC 50 of 60 μM. These results support the hypothesis that AlCl 3 inhibition of agonist-stimulated IP accumulation may be mediated by inhibition of PiPLC activity, rather than disruption of G-protein function or modulation of the IP signalling system by Ca 2+ or PKC.

Prolonged enhancement of synaptic transmission in area CA1 of rat hippocampal slices induced by NaF/AlCl 3 does not require NMDA receptor activation but is suppressed by inhibitors of phosphoinositide-mediated signalling pathways

The processes underlying the action of AlF 4 − (10 mM NaF/10 μM AlCl 3) in inducing long-lasting enhancement of synaptic transmission in area CA1 of rat hippocampal slices have been investigated. Exposure of hippocampal slices to AlF 4 − for 10 min caused population EPSP slope to rise by approximately 50% within 60 min of washing off the NaF/AlCl 3 saline. This effect was not inhibited either by APV (50 μM), or by temporary interruption of the delivery of test stimuli during and for up to 20 min after application of the AlF 4 −-containing medium. However, pretreatment of preparations with either thapsigargin (1 μM) or staurosporine (1 μM), or ommission of Ca 2+ from the AlF 4 −-containing saline (no addition of EGTA) prevented the potentiating action of NaF/AlCl 3. We conclude that the potentiating effect of AlF 4 − is via a G-protein linked to phosphoinositide turnover, and that both arms of this signalling pathway are necessary for potentiation to occur. Ca 2+ influx is also a requirement, but does not occur through the NMDA receptor.

Aluminum decreases muscarinic, adrenergic, and metabotropic receptor-stimulated phosphoinositide hydrolysis in hippocampal and cortical slices from rat brain

Effects of alumiinum chloride (AlCl 3)(0.1 to 1000 μM) on inositol phosphate (IP) accumulation stimulated by carbachol (CARB), norepinephrine (NE) or quisqualate (QUIS) were examined in rat hippocampal and cortical slices. In the absence of agonist, only 1000 μM AlCl 3 significantly reduced basal accumulation of IPs. For CARB-stimulated IP accumulation, 100 μM and greater AlCl 3 significantly inhibited IP accumulation. In cortical slices, 1000 μM AlCl 3 reduced CARB-stimulated IP accumulation by 55% and in hippocampal slices 1000 μM AlCl 3 inhibited IP accumulation by 40%. Similar effects of AlCl 3 were observed for NE-stimulated IP accumulation. In cortical slices, the concentration-response for AlCl 3 effects on agonist-stimulated IP accumulation was significantly different from that in hippocampal slices. For QUIS-stimulated accumulation of IPs, 1000 μM AlCl 3 significantly inhibited IP accumulation in hippocampal slices. However, in cortical slices a biphasic effect of AlCl 3 was observed. 500 and 1000 μM AlCl 3 significantly inhibited IP accumulation, whereas 10 and 50 μM AlCl 3 significantly enhanced QUIS-stimulated IP accumulation. In both hippocampal and cortical slices, 500 μM AlCl 3 significantly inhibited CARB-, NE- or QUIS-stimulated IP accumulation at all agonist concentrations (0.1 to 10000 μM) tested, indicating a post-receptor effect on agonist-mediated IP accumulation. Stimulation of G-proteins with NaF (5–30 mM) resulted in accumulation of IPs in hippocampal and cortical slices in the absence of added agonists. NaF (5–30 mM) plus 1 mM CARB produced increased accumulation of IPs over CARB or NaF alone. AlCl 3 inhibited CARB- and NaF-stimulated IP accumulation, suggesting that AlCl 3 decreases IP accumulation by inhibiting G-protein or phospholipase C (PLC) activity. To determine the influence of the form of aluminum salt on IP accumulation, the effects of equinolar concentrations of AlCl 3, aluminum citrate (Al(Cit)), and aluminum lactate (Al(Lac)) on CARB-stimulated IP accumulation were determined. AlCl 3 and Al(Lac) produced similar inhibition of IP accumulation, whereas Al(Cit) did not inhibit receptor-stimulated IP accumulation. These results suggest that: (1) AlCl 3 disrupts IP accumulation mediated by different pharmacological classes of receptors; (2) differences in sensitivity to AlCl 3 exist in the hippocampus and cortex; (3) AlCl 3 and AlF − 4 influence IP accumulation through separate mechanisms and AlF − 4 is not responsible for effects of AlCl 3 on receptor-stimulated IP accumulation; (4) AlCl 3 appears to exert its action on IP accumulation ‘downstream’ of the receptor and receptor-G-protein interaction; and (5) Effects of aluminum on IP accumulation are dependent upon the aluminum salts used.

Al3+-Ca2+ interactions in aluminum rhizotoxicity

The cation Al3+ is toxic to plants at micromolar concentrations and can severely inhibit root growth in solution experiments. Trivalent aluminum hydrolyzes in solution, and, apart from the Al ion, which dominates speciation below pH 5.0, various mononuclear and polynuclear hydroxy-Al species can also occur (Kinraide 1991). Accumulating evidence suggests that Al is the rhizotoxic species under the experimental conditions used in the present study (Kinraide 1991; Kinraide et al. 1992). The inhibition of Ca2+ uptake in roots by Al3+ has been proposed as a possible mechanism for Al3+ toxicity, and in this study the hypothesis was tested directly. Root growth and Ca2+ uptake were measured in 5-d-old seedlings of wheat (Triticum aestivum L. Thell) during exposure to Al3+ in a low-Ca2+ basal medium, and to Al3+ in the presence of added cations. Uptake of Ca2+ in whole roots and translocation to the shoot were measured using 45Ca2+, and localized measurements of net Ca2+ flux were also made at the root apex using the technique of microelectrode ion-flux estimation. Treatment with 2.64 μM AlCl3 in 226 μM CaCl2, at pH 4.5, severely inhibited root growth without affecting Ca2+ uptake. Addition of 30 mM Na2+, 3 mM Mg2+ or 50 μM tris(ethylenediamine)cobalt(III) to this Al3+ treatment restored root growth but significantly reduced Ca2+ uptake measured over the entire root system and at the root apex. The Al3+ and Ca2+ concentrations were adjusted so that the activities of the Al3+ and Ca2+ ions were constant in all solutions (1.5 μM and 200 μM, respectively). Root growth can be severely inhibited by Al3+ concentrations that do not affect Ca2+ uptake, while the addition of ameliorating cations depresses Ca2+ uptake. These results argue against the hypothesis that Al3+ inhibits root growth by reducing Ca2+ uptake.

Evidence for the Involvement of G Proteins in the Modulation of 22Na and 45Ca Fluxes in Myogenic L6 and Aortic Smooth Muscle Cells

Preincubation of rat skeletal muscle derived L6 myoblasts with 10 mM NaF increased, the activity of Na+/H+ exchanger, and the uptake of 45Ca by 2 and 5 folds, respectively. In cultured vascular smooth muscle cells (CVSMC) of rat aorta, NaF increased the activity of Na+ ,K+-pump, Na+/H+ exchanger, passive permeability for Na+, and 45Ca uptake by 1.6, 9, 2, and 9 folds, respectively. Both, in CVSMC and L6 cells, the effect of NaF on the Na+/H+ exchanger and 45Ca uptake were significantly augmented by 20 μM AlCl3. No effect of AlCl3 on the NaF dependent changes in ion flux was seen in rat erythrocytes. The results suggest that in L6 and CVSMC cells, the Na+/H+ exchanger and Ca++ uptake pathway(s) are activated by GTP-binding proteins.

Selection of Bradyrhizobium strains and provenances of Acacia mangium and Faidherbia albida: Relationship with their tolerance to acidity and aluminium

This work was designed to determine the role of the acidity and aluminium stress in the selection of partners in the Acacia symbioses with relevance to the persistence of the microsymbiont Bradyrhizobium in the soil and the growth and nodulation of the host plant respectively. Fifteen strains of Bradyrhizobium from Acacia mangium and Faidherbia albida formed a very homogenous acid tolerant group as indicated by their ability to grow better in a medium at pH 4.5 than in a medium at pH 6.8. By contrast, a growth experiment using an acid liquid media (pH 4.5), containing different concentrations of aluminium successfully identified strains sensitive to aluminium toxicity and those able to grow even in the presence of 100 μM AlCl3. Our results suggest that high amounts of aluminium in the soil rather than acidity (pH 4.5) were a major soil factor for selection of Bradyrhizobium strains capable of establishing a permanently high population under natural conditions. Unlike the behaviour of the microsymbiont, growth and nodulation of Acacia mangium and Faidherbia albida were not affected by aluminium, even at 100 μM, but they might be significantly affected by medium acidity (pH 4.5) depending on plant provenances. It is therefore suggested that ability of the host plant to tolerate acidity stress should be taken into account first when screening effective Acacia-Bradyrhizobium combinations for use in afforestation trials.

Short-term Responses of Soybean Roots to Aluminium

The physiological reasons for the inhibition of root elongation by Al are not yet understood. The objective of the present studies was to clarify short-term effects of Al on plasma-membrane integrity of soybean (Glycine max L.) roots using different physiological/biochemical parameters. Initial response (0–3 h) of roots to Al (50 μM AlCl3) was a reduction of K+ net-efflux. This was most pronounced at low Ca supply (25 versus 2500 μM). Using 86Rb, it could be shown that Al effects on K+ net-efflux were mainly due to reduced K+ efflux whereas effects on K+ influx were negligible. After 2 h of Al treatment, induction by Al of non-constitutive callose synthesis could be demonstrated. Lipid peroxidation was enhanced after 32 h of Al treatment especially in the presence of higher Fe(II) concentrations. Fe(III) reducing capacity of Fe-deficient plants, as an indicator of high activity of plasma membrane- bound processes, was not impaired but rather enhanced by short-term Al treatment. The results clearly indicate, that Al concentrations leading to severe inhibition of root elongation do not primarily lead to membrane disruption. However, specific membrane functions are modified by Al.

Schistosoma mansoni: Characterization of phosphoinositide response

Signal transduction pathways may have important regulatory roles in cellular events in the human parasite Schistosoma mansoni. The presence of the phosphoinositide response in S. mansoni was examined by radiolabeling intact worms with 20 μCi of [ 3H]myoinositol for 24 hr and stimulating parasites with 25 m M NaF and 10 μM AlCl 3 in the presence of 10 m M LiCl. Total inositol phosphates were increased within 2 min and maximal accumulation was achieved after 30 min. Similar results were seen with the nonhydrolyzable GTP analogues GTP γS and GppNHp while only minimal changes were detected with GMP. Neomycin inhibited NaF-induced inositol phosphate production. NaF stimulated a significant 3.6-fold increase of inositol phosphates in females compared to males. These data suggest that stimulation of guanine nucleotide-binding regulatory proteins activates phospholipase C resulting in production of inositol phosphates in S. mansoni.

Effects of Ro 5-4864 and PK 11195 in rat duodenum and vas deferens

Ro 5-4864 and PK 11195 inhibit in a concentration-dependent manner carbachol-induced contractions in rat duodenum (IC 50: 1.56 ± 0.07 × 10 −5 M and 1.18 ± 0.07 × 10 −5 M respectively). The antagonism is non-competitive and is not mediated by peripheral-type benzodiazepine receptors. The Ro 5-4864 effect is modulated by the calcium concentration of the Tyrode-Ringer solution. In the presence of 1 mM NaF/10 μM AlCl 3, Ro 5-4864 and PK 11195 do not inhibit carbachol-induced contractions. Moreover, Ro 5-4864 and PK 11195 significantly relax AlF 4 −-induced contractions, with IC 50 values of 2.01±0.12×10 −5 M and 1.28 ± 0.11 × 10 −5 M respectively. This effect is also modulated by the calcium concentration of the medium. Pertussis toxin potentiates the antagonist effects of Ro 5-4864 and PK 11195 on carbachol-induced contractions, but cholera toxin does not affect them. Ro 5-4864 and PK 11195 inhibit 45Ca 2+ uptake induced by KCl (120 mM) in rat vas deferens, but do not affect either basal 45Ca efflux or noradrenaline-induced 45Ca 2+ efflux. Only high doses of PK 11195 (above 5 × 10 −5 M) are able to produce a slight reduction of the accumulation of inositol phosphates induced by methoxamine in rat vas deferens, while Ro 5-4864 has no significant effect. Finally, Ro 5-4864 and PK 11195 reduce calcium influx, but do not seen to be the only mechanism of the antagonistic effect on carbachol-induced contractions. An alteration of other second messengers. probably cyclic monophosphate nucleotides, may be involved.

Difference in effects of sodium fluoride and cholecystokinin on diacylglycerol accumulation and calcium increase in guinea pig gastric chief cells

In isolated guinea pig gastric chief cells, sodium fluoride (NaF) stimulated a monophasic increase in diacylglycerol accumulation, while cholecystokinin (CCK) strongly stimulated its biphasic accumulation. NaF evoked an increase in initial Ca 2+ influx rate with a slow increase in intracellular free Ca 2+ concentration ([Ca 2+]i), while CCK stimulated a rapid increase in [Ca 2+]i followed by a late sustained phase of the [Ca 2+]i increase. Lanthanum chloride (La 3+) effectively blocked NaF-stimulated increase in [Ca 2+]i, but it blocked only CCK-stimulated late sustained phase of [Ca 2+]i increase. The effect of NaF on pepsinogen secretion was enhanced in the presence of 100 μM AlCl 3. Furthermore, pertussis toxin did not affect NaF-evoked diacylglycerol accumulation at all. These results suggest that NaF may activate a pertussis-toxin insensitive guanine nucleotide regulatory protein (G protein) coupled to a signal transducing mechanism which seems to be distinct from that activated by CCK, thereby inducing increases in diacylglycerol accumulation, Ca 2+ influx and pepsinogen secretion in guinea pig gastric chief cells.

Effect of monovalent ions upon G proteins coupling muscarinic receptors to phosphoinositide hydrolysis in the rat cerebral cortex

It has been suggested that K +, Li + and F1 − affect the function of G proteins coupled to signal transducing enzymes. Lithium, at concentrations which were found to reduce forskolin-stimulated adenylate cyclase activity, was without effect on either membrane [ 3H]phosphatidylinositol-4,5-bisphosphate ([ 3H]PIP 2) hydrolysis measured in the absence or presence of 5′-guanylyl-imidodiphosphate (Gpp(NH)p), or (at ⩾ 2.3 mM Li +) upon the stimulation of rat cerebral cortical inositol phospholipid breakdown by either carbachol, noradrenaline or NaF measured at either 6 or 18 mM K +. The increase in assay [K +] greatly enhanced the inositol phospholipid response to carbachol but not to NaF. The inhibitory effect of carbachol upon forskolin-stimulated adenylate cyclase was not affected by raising the [K +] from 6 to 18 mM. At 6 mM K + (both in the absence and presence of 15 μM AlCl 3), the effects of carbachol and NaF upon inositol phospholipid breakdown were essentially additive, whereas at 18 mM K +, the breakdown response to carbachol (antagonised by pirenzepine with a pA 2 value of 7.6) was similar in the absence and presence of NaF. It is concluded that in the rat cerebral cortex: (a) Li + does not affect the function of either the phosphoinositide-specific phospholipase C enzyme itself or the G p coupled to this enzyme; (b) the difference between the additivity between NaF and carbachol seen at different assay [K +] may reflect the K +-dependent changes in the tetrodotoxin-resistant and tetrodotoxin-sensitive pathways of carbachol stimulation of inositol phospholipid breakdown reported by Gurwitz and Sokolovsky (1987, Biochemistry 26, 633); and (c) the effect of K + on muscarinic receptor-coupled inositol phospholipid breakdown is not found for muscarinic receptors inhibitorily coupled to adenylate cyclase. Evidence is also presented to suggest that NaF affects the dephosphorylation of the formed [ 3H]inositol polyphosphates.

Inhibition of proton transport activity of microsomal membrane vesicles of barley roots by aluminium

Abstract The extrusion of protons from the roots of barley (Hordeum vulgare L.) seedlings in the presence of KCl decreased markedly after treatment of the samples with 1 mm AlCl3 at pH 5.5. The membrane vesicles with ATP-dependent proton transport activity were prepared from barley roots. The proton transport activity measured by the quenching of the fluorescence of quinacrine was inhibited by approximately 50% in the presence of 100 μm AlCl3 at pH 6.5 but the inhibition was only 10% at pH 7.5. The values of V max and K m for ATP related to the proton transport activity of the vesicles were affected markedly by AlCl3. The inhibition of the proton transport activity of the vesicles due to Al was effectively restored by citric acid. The results suggest that one of the mechanism of Al toxicity in plant roots is the inhibition of the proton transport activity associated with the cell membrane which in turn leads to the inhibition of proton extrusion from the roots.

Effect of long-term feeding of aluminium chloride on hexokinase and glucose-6-phosphate dehydrogenase in the brain

Rats were fed 100 μM AlCl 3 for 1 year in their drinking water, then killed and their brains homogenized in 0.1 M Tris (pH 7.4). The 800 g supernatants were assayed for Al 3+ and the activities of acetylcholine esterase (ACE), hexokinase and glucose-6-phosphate dehydrogenase (G6PDH). The concentrations of Al in the homogenates, as computed on the original brain for the control and Al fed group were 40 ng and 80 ng/g wet wt, respectively. The activity of ACE was the same in both groups but that of hexokinase and G6PDH in the Al-fed group was about 73% and 80%, respectively, of the control. Dialysis restored the G6PDH but increased the hexokinase of the control group 2-fold and that of Al-fed group 2.7-fold. Thus at this elevated level it was same in both groups. The contribution of Al from the undialysed homogenates during assay was too low to account for the inhibition. It is therefore suggested that a dialyzable inhibitor for hexokinase is normally present in the brain and that Al feeding increases its concentration to further inhibit the utilization of glucose.

The effects of Al 3+, Cd 2+ and Mn 2- on human erythrocyte choline transport

The effects of Al 3+, Cd 2+ and Mn 2+ on human erythrocyte choline transport, Na-K-ATPase, Ca-Mg-ATPase and intracellular K + levels were examined. The concentrations used were below the levels which caused significant haemolysis (⩽300 μM). All three cations inhibited concentrative choline accumulation over 3 hr [ ic 50 values at 1 μM choline were 35 μm (AlCl 3), 250 μM (CdCl 2) and 300 μm (MnCl 2)] but at the concentrations tested, none decreased initial rates of choline uptake. The effects of Cd 2− and Mn 2− (but not Al 3+) on choline accumulation were reversed by removing the cations from the extracellular medium by washing. All three cations also inhibited efflux of choline. at 1 μM choline, 30% inhibition being produced by 33 μM AlCl 3, 81μM CdCl 2 and 111 μM MnCl 2. At subhaemolytic concentrations, only CdCl 2 inhibited Na-K-ATPase, ( ic 50 = 147 μ M) and none of the cations significantly inhibited Ca-Mg-ATPase. Intracellular K − levels were only reduced by the highest concentration of AlCl 3 used (100 μM). These results suggest that inhibition of choline accumulation and efflux in erythrocytes by Al 3−, Cd 2+ and Mn 2− is not explicable solely in terms of either inhibition of Ca-Mg-ATPase, or inhibition of Na-K-ATPase causing reduced intracellular K −. Our conclusions are similar to those previously obtained using synaptosomes and provide support for the hypothesis that inhibition of choline transport by Al 3+ may contribute to a number of disease states.