K channels of human alveolar macrophages

The activation of macrophages has been reported to be associated with Ca-activated K permeability change. In order to study this permeability change in human alveolar macrophages, we examined alveolar macrophages electrophysiologically at a single channel level. We observed two types of Ca-activated K channel currents having conductances of 218 +/- 2 and 32 +/- 0.6 picosiemens in symmetrical 154 mmol/L KCl solutions. The characteristics, such as voltage dependency and Ca sensitivity, as well as channel conductance, were different between these two types of channel currents. Quinine (a blocker of Ca-activated K conductance), 0.5 mmol/L, reduced these channel currents by 45 +/- 8% and 31 +/- 8%. Quinine, 0.5 mmol/L, also inhibited chemiluminescence and leukotriene B4 release by 82 +/- 6 to 88 +/- 3% and 88 +/- 2%, respectively. These results suggest the presence of two types of Ca-activated K channels, which may be related to the release of inflammatory mediators from human alveolar macrophages.     

Effects of caffeine on potassium currents in isolated rat ventricular myocytes

Rapid exposure of cardiac muscle to high concentrations of caffeine releases Ca(2+) from the sarcoplasmic reticulum (SR). This Ca(2+) is then extruded from the cell by the Na(+)/Ca(2+) exchanger. Measurement of the current carried by the exchanger (I(Na/Ca)) can therefore be used to estimate of the Ca(2+) content of the SR. Previous studies have shown that caffeine, however, can also inhibit K(+) currents. We therefore investigated whether the inhibitory effects of caffeine on these currents could contaminate measurements of I(Na/Ca). Caffeine caused partial inhibition of the inward rectifier K(+) current (I(K1)): the outward current at -40 mV was 1.15+/-0.24 pA/pF in control and decreased to 0.34+/-0.15 pA/pF in the presence of 10 mmol/l caffeine (P<0.05, n=15). This was similar to the effect of caffeine on the holding current observed at -40 mV in the absence of K(+) channel block and could therefore account for the contaminating effects of caffeine observed during measurements of I(Na/Ca). Moreover, caffeine also partially inhibited the transient outward ( I(to)) and the delayed rectifier (I(K)) K(+) currents.     

ATP-sensitive potassium channels in freshly dissociated adult rat striatal neurons: activation by metabolic inhibitors and the dopaminergic receptor agonist quinpirole

The characteristics of adenosine 5'-triphosphate (ATP)-sensitive K+ channels in acutely isolated striatal neurons from adult rats were examined. Neurons had a resting membrane potential of -53.9+/-1.2 mV (n=66), with evoked or spontaneous action potentials firing at 10+/-0.7 Hz, and large inwards and outwards whole-cell currents. In cell-attached patches with a high [K+] in the pipette, a voltage-independent, ATP-insensitive 16.5+/-1.5 pS channel was observed in 375 out of 452 cells. Bath application of Na+-azide (0.5-2 mM) to 108 neurons revealed another 145.7+/-3.5 pS (LKATP) channel in 65 neurons; this channel was blocked by tolbutamide. The LKATP channel exhibited a high open probability (Po, 0.8+/-0.05) at 0 mV pipette potential. Varying the pipette [K+] shifted the reversal potential of LKATP, showing the channel's K+ selectivity. Cytoplasmic ATP (ATPi) reversibly inhibited LKATP, with an inhibitory constant (Ki) of 0.12 mM. LKATP was sensitive to intracellular Ca2+ but insensitive to iberiotoxin. In 25% of cell-attached patches, the presence of quinpirole in the pipette opened a third type of channel (90.6+/-1.7 pS, termed D2KATP). Sulpiride, a dopamine D2-receptor antagonist, inhibited D2KATP. ATPi reversibly inhibited D2KATP, with a Ki of 0.212 mM. The Na+-azide- or quinpirole-induced current caused a tolbutamide-sensitive membrane hyperpolarization and a marked reduction in action potential frequency. We propose that ATP-sensitive K+ channels play a metabolism-dependent role in striatal neurons.     

Blood plasma and saliva levels of magnesium and other bivalent cations in patients with parotid gland tumors.

The plasma and saliva cations in parotid malignant tumors of stages II-III were studied in 31 patients before surgical therapy and in 27 control group volunteers. The magnesium (t-Mg), calcium (t-Ca), copper (t-Cu) and zinc (t-Zn) concentrations in plasma were determined, and t-Mg and t-Ca in saliva. Our results showed that salivary and plasma t-Mg concentrations were significantly higher in patients with parotid malignant tumors in comparison to control group (saliva: 0.25 +/- 0.04 mmol/L versus 0.14 +/- 0.03/L, p < 0.01; plasma: 1.05 +/- 0.06 mmol/L versus 0.86 +/- 0.05 mmol/L, p < 0.05). The t-Ca plasma concentrations were lower for patients with parotid malignant tumors by 20-22% in comparison to the control group (p < 0.05). Plasma and salivary t-Mg/t-Ca molar ratios are respectively 0.38 and 0.12 for control group, and respectively 0.61 and 0.31 for patients with parotid gland tumors. The t-Zn plasma concentration for patients with parotid malignant tumors (0.017 +/- 0.010 mmol/L) was significantly lower (p < 0.05) in comparison to control group (0.024 +/- 0.011 mmol/L). Plasma t-Cu/t-Zn molar ratio is respectively 0.68 for control group and 1.12 for patients with parotid gland tumors. The mechanism responsible for the increase of salivary magnesium as a consequence of the development of tumoral tissue needs to be clarified.     

Outwardly rectifying chloride current in rabbit osteoclasts is activated by hyposmotic stimulation.

1. We characterized chloride currents in freshly isolated rabbit osteoclasts using whole-cell and single channel patch-clamp recording configurations. Depolarization activated an outwardly rectifying current in 40-50% of cells, distinct from the inwardly rectifying K+ current we have previously reported in osteoclasts. 2. The outwardly rectifying current persisted under conditions where all K+ currents were blocked. Furthermore, the outward current was reversibly inhibited by Cl- transport blockers 4-acetamido-4'-isothiocyanostilbene-2,2'-disulphonic acid (SITS); 4,4'-diisothiocyanostilbene-2,2'-disulphonic acid (DIDS); 4,4'-dinitrostilbene-2,2'-disulphonic acid (DNDS); and niflumic acid. The blocked current had a reversal potential close to the predicted chloride equilibrium potential and was dependent on the chloride concentration gradient. 3. In those osteoclasts in which outwardly rectifying current was not initially apparent, exposure to hyposmotic extracellular solution resulted in its reversible activation. The induced current was due to Cl-, based on its reversal close to the chloride equilibrium potential and sensitivity to blockade by Cl- channel inhibitors. The hyposmotically induced current could be activated in Ca(2+)-free solutions containing 0.2 mM EGTA. 4. When studied in the current-clamp configuration, hyposmotic stimulation caused depolarization from -76 +/- 5 to -5 +/- 6 mV (mean +/- S.D., n = 7). 5. Unitary Cl- currents were recorded in the cell-attached patch configuration at positive potentials. Single channels had a slope conductance of 19 +/- 3 pS (n = 5). Reduction of the external [Cl-] shifted the current-voltage relationship in the positive direction, supporting the conclusion that these were Cl- currents. Like the whole-cell currents, single channel Cl- currents were activated by exposure of cells to hyposmotic bathing solution. 6. We conclude that rabbit osteoclasts express an outwardly rectifying Cl- current that can be activated by osmotic stress. Cl- channels may play a role in cell volume regulation and may also provide conductive pathways for dissipating the potential difference that arises from electrogenic proton transport during bone resorption.     

阻断MaxiK 通道对人单核细胞源性巨噬细胞向泡沫细胞分化的抑制作用

目的研究高电导钙离子激活钾通道(MaxiK通道)在人单核细胞源性巨噬细胞向泡沫细胞分化的过程中,其mRNA和蛋白的表达及作用。方法采用密度梯度离心加贴壁黏附法,从男性健康志愿者的外周血中分离单核细胞,经5d培养后分化为巨噬细胞。在建立人巨噬细胞源性泡沫细胞模型的基础上,采用免疫细胞化学染色法、RT-PCR及蛋白印迹,研究MaxiK通道α-亚单位的表达,并观察其特异性阻断剂-Paxilline对摄取氧化型低密度脂蛋白(OxLDL)的巨噬细胞中胆固醇代谢的影响。结果将巨噬细胞同30mg/LOxLDL于37℃孵育60h后,细胞体积增大,并有许多红色的脂质颗粒沉积于细胞质内,细胞内的总胆固醇(TC)、游离胆固醇(FC)及胆固醇酯(CE)的含量均显著增加,CE/TC从(14.437±6.781)%提高到(57.946±3.507)%(n=7,P<0.05);而MaxiK通道α-亚单位mRNA及蛋白的表达水平没有明显改变(P>0.05)。5和10μmol/L的Paxilline能显著减少巨噬细胞内TC、FC及CE的含量,CE/TC分别降至(41.217±5.584)%和(18.017±11.559)%(n=7,P<0.05),细胞内沉积的红色脂质颗粒也明显减少。结论阻断MaxiK通道能抑制人单核细胞源性巨噬细胞向泡沫细胞分化。     

Protein kinase C suppresses spontaneous, transient, outwards K+ currents through modulation of the Na/Ca exchanger in guinea-pig gastric myocytes

The effect of protein kinase C (PKC) on the Ca2+-activated K+ current (IK,Ca) in guinea-pig gastric myocytes was studied using the whole-cell voltage-clamp technique. At a holding potential of 0 mV, IK,Ca, recorded as spontaneous, transient, outwards currents (STOCs), was markedly inhibited, both in mean amplitude (54 +/- 5%) and frequency (60 +/- 8%) by 1 microM phorbol 12, 13 dibutyrate (PDBu, n = 6). These effects were antagonized by pretreatment with 10 nM bisindolylmaleimide I (n = 5), a selective inhibitor of PKC. The possibility that the inhibition of STOCs was due to direct channel inhibition by PKC was addressed using inside-out or open-cell-attached patch-clamp techniques, the latter established using beta-escin. PDBu did not alter the conductance or open probability of the KCa channel in any mode, suggesting that PKC does not inhibit the KCa channel directly. To study the involvement of the Na/Ca exchanger in the inhibition of STOCs by PDBu, its operation was prevented by replacing Na+ in the internal solution by tris(hydroxymethyl)aminomethane (TRIS) and external Na+ by equimolar K+ and Ca2+-activated inwards K+ currents recorded at a holding potential of 0 mV. Neither the mean amplitude (96 +/- 8%) nor the frequency of these currents was inhibited significantly by 1 microM PDBu (n = 5). Like PDBu, 5 microM 2-(2-[4-(4-nitrobenzyloxy)phenyl]ethyl) isothiourea methanesulphonate (KB-R7943), a selective inhibitor of the reverse mode Na/Ca exchanger, also inhibited the mean amplitude (45 +/- 6%) and frequency (26 +/- 2%) of STOCs at the holding potential of 0 mV (n=6). The results suggest that the suppression of STOCs by PKC is mediated by inhibition of the Na/Ca exchanger.     

Prediction of equilibrated urea in children on chronic hemodialysis

Urea rebound (UR) after hemodialysis (HD) requires the use of equilibrated urea (Ceq) instead of immediate end-dialysis urea (Ct) for correct quantification of HD, which is impractical. A new formula for predicting Ceq in children is suggested in our study. Thirty eight standard pediatric HD sessions (single pool Kt/V = 1.70 +/- 0.35, K = 4.65 +/- 1.14 ml/min/kg, UF coeff. = 3.2-6.2 ml/h/mm Hg, t = 3.80 +/- 0.46 h) in 15 children (M: 6, F: 9), ages 14.5 +/- 3.28 years were analyzed. Blood samples were taken: before, 70 min from the start, at the end, and 60 min after the end of HD sessions. After correlating UR (20.32 +/- 7.74%) to various HD parameters, we found that it was mainly determined by HD efficiency parameters. Therefore we correlated Ceq to HD efficiency parameters (Ct, urea reduction ratio, Kt/V, and K/V) and found a very high correlation between Ct and Ceq (r = 0.973). Linear regression analysis was used to further investigate this relationship, and a new formula to predict Ceq from Ct was obtained (Ceq = 1.085 Ct + 0.729, R2 = 0.946, SE = 0.49, absolute residuals = 0.38 +/- 0.29 mmol/L). In a validation study (10 HD sessions with new set of urea blood samples) the results obtained by the new formula were compared with measured values of Ceq and those obtained by the Smye formulae. Values predicted by the new formula (9.91 +/- 2.92 mmol/L) were not significantly different from the measured values (10.33 +/- 3.44 mmol/L). Absolute error of the new formula was 0.78 +/- 0.73 mmol/L, median 0.65; ie., 6.93 +/- 5.3%, median 7.7%. Ceq predicted by the Smye formulae (10.95 +/- 4.18 mmol/L) also did not significantly differ from the measured values, but absolute error of predicted values was markedly higher (1.21 +/- 0.90 mmol/L, median 0.89; 11.73 +/- 7.72%, median 10.11%; p < 0.05). When predicted Ceq was used for calculating equilibrated Kt/V (eKt/V), the new formula resulted in lower absolute error (0.09 +/- 0.07, median 0.08) than the Smye method (0.14 +/- 0.08, median 0.12). We conclude that our simple formula is sufficiently accurate in predicting Ceq in standard pediatric HD and that it is more accurate than the existing Smye formulae, while requiring only pre- and post-HD urea samples. We suggest the use of the new formula for predicting Ceq, which can then be used instead of Ct for a more accurate estimation of double pool Kt/V, URR, V, and PCR.     

Effects of lysophosphatidylcholine on resting potassium conductance of isolated guinea pig ventricular cells

We studied the effects of lysophosphatidylcholine (LPC), a toxic metabolite of ischemia, on the inward rectifier potassium channel current in isolated guinea pig ventricular cells. LPC (10-50 microM) added to the external solution decreased the resting membrane potential and occasionally induced repetitive action potential discharges, with or without loss of repolarization. In voltage clamp studies, LPC (20 microM) decreased the conductance at the levels of resting potentials (approximately equal to -80 mV) from 26 +/- 8 nS to 16 +/- 3 nS (mean and SD, n = 4) within 10 min. Prolonged application of LPC (greater than 12 min) produced transient inward currents after depolarizing clamp pulses, thereby suggesting that the LPC elevated intracellular Ca2+ concentrations. The effect of LPC on the single inward rectifier K channel current was examined using the patch clamp technique in a cell-attached mode. LPC decreased the single channel conductance, depending on the concentration (5-100 microM). The slope conductance in the presence of 150 mM K+ in the pipette decreased from 45 +/- 7 pS (control) to 32 +/- 17, 20 +/- 19, and 14 +/- 10 pS for 5, 20 and 100 microM LPC, respectively. LPC induced little change with regard to probability of the channel opening. These results suggest that LPC depolarizes membrane by decreasing single channel conductance of the inward rectifier K channel. This reduction partially contributes to the alleged LPC-induced abnormal automaticities and conduction disturbances in the heart.     

Mechanosensitive channel currents recorded in rat microvascular endothelial cells with whole-cell mode.

Using the whole-cell patch-clamp technique, we recorded delayed outward currents in rat brain microvascular endothelial cells (BRMECs), which were nearly completely inhibited by 20 mmol/L extracellular TEA-Cl and 5 mM extracellular CsCl. Whole-cell currents were elicited under voltage clamp condition by 2100 ms depolarizing voltage pulses applied every 7 s between -100 to 90 mV in 10 mV increment from a holding potential of -100 mV. The currents were defined as delayed rectified K+ currents (IKv), which were inhibited in a concentration-dependent manner by bath application of TEA-Cl, with an IC50 approximately 2.0 mM, similar to that reported on IKv in other preparations. In the present of mechanical force, outward currents were increased in amplitude as compared with controls. These mechanical force induced currents were also defined as IKv, which are different from previous described mechanosensitive currents with characteristic of inward rectifier.     

K+ currents generated by NMDA receptor activation in rat hippocampal pyramidal neurons

Long lasting outward currents mediated by Ca2+-activated K+ channels can be induced by Ca2+ influx through N-methyl-D-aspartate (NMDA)-receptor channels in voltage-clamped hippocampal pyramidal neurons. Using specific inhibitors, we have attempted to identify the channels that underlie these outward currents. At a holding potential of -50 mV, applications of 1 mM NMDA to the soma of cultured hippocampal pyramidal neurons induced the expected inward currents. In 44% of cells tested, these were followed by outward currents (average amplitude 60 +/- 7 pA) that peaked 2.5 s after the initiation of the inward NMDA currents and decayed with a time constant of 1.4 s. In 43% of those cells exhibiting an outward current, SK channel inhibitors, UCL 1848 (100 nM) and apamin (100 nM) abolished the outward current. In the remainder of the cells, the outward currents were either insensitive or only partly inhibited (44 +/- 4%) by 100 nM UCL 1848. In these cells, the outward currents were reduced by the slow afterhyperpolarization (sAHP) inhibitors, muscarine (3 microM; 43 +/- 9%), UCL 1880 (3 microM; 34 +/- 10%), and UCL 2027 (3 microM; 57 +/- 6%). Neither the BK channel inhibitor, charybdotoxin (100 nM), nor the Na+/K+ ATPase inhibitor, ouabain (100 microM), reduced these outward currents. Irrespective of the pharmacology, the time course of the outward current did not differ. Interestingly, no correlation was observed between the presence of a slow apamin-insensitive afterhyperpolarization and an outward current insensitive to SK channel blockers following NMDA-receptor activation. It is concluded that an NMDA-mediated rise in [Ca2+]i can result in the activation of apamin-sensitive SK channels and of the channels that underlie the sAHP. The activation of these channels may, however, depend on their location relative to NMDA receptors as well as on the spatial Ca2+ buffering within individual neurons.     

P2X currents in peritoneal macrophages of wild type and P2X4 -/- mice

In this study the ATP-induced (P2X) currents in isolated peritoneal macrophages of wild type (WT) and P2X(4) knockout (P2X(4)(-/-)) mice were studied by means of whole-cell patch clamp in order to (1) survey the P2X currents of native macrophages and (2) to investigate the expression of P2X(4)-like currents in the WT versus P2X(4)(-/-) mice. Three types of currents were observed in the isolated macrophages: (1) in approximately 10% of both WT and P2X(4)(-/-) macrophages a fast activating and inactivating P2X1-like current was recorded with low concentrations (0.1-1 microM) of ATP; (2) 85% of wild type and 100% of P2X(4)(-/-) macrophages exhibited a non-desensitizing P2X(7)-like current activated at high concentrations of ATP (10mM). The identity of the P2X(7) current was confirmed using the specific blocker A-740003; (3) 88.6% of the WT but none of the P2X(4)(-/-) macrophages showed a small P2X(4)-like current that desensitized slowly upon ATP application at intermediate concentrations (3-300 microM). Several observations indicated that the slowly desensitizing current in WT macrophages was P2X(4). The EC50 value of 5.3 microM ATP was as expected for P2X(4) and the current induced by 3-300 microM ATP was absent in P2X(4)(-/-) mice. Upon application of 3 microM ivermectin, a P2X(4)-selective modulator, the amplitude of this current was increased and the desensitization was inhibited in WT cells. In addition, this current was facilitated by 10 microM Zn(2+) but inhibited by Cu(2+) (in contrast to P2X(2)). We conclude that the P2X(4) and P2X(7) currents are functionally expressed in recruited peritoneal macrophages of WT mice and that the P2X(4)-like current is absent in P2X(4)(-/-) mice.     

Major potassium conductance in type I hair cells from rat semicircular canals: characterization and modulation by nitric oxide

Mammalian vestibular organs have two types of hair cell, type I and type II, which differ morphologically and electrophysiologically. Type I hair cells alone express an outwardly rectifying current, I(K, L), which activates at relatively negative voltages. We used whole cell and patch configurations to study I(K,L) in hair cells isolated from the sensory epithelia of rat semicircular canals. I(K,L) was potassium selective, blocked by 4-aminopyridine, and permeable to internal cesium. It activated with sigmoidal kinetics and was half-maximally activated at -74.5 +/- 1.6 mV (n = 35; range -91 to -50 mV). It was a very large conductance (91 +/- 8 nS at -37 mV; 35 nS/pF for a cell of average size). Patch recordings from type I cells revealed a candidate ion channel with a conductance of 20-30 pS. Because I(K,L) was activated at the resting potential, the cells had low input resistances (R(m)): median 25 MOmega at -67 mV versus 1.3 GOmega for type II cells. Consequently, injected currents comparable to large transduction currents (300 pA) evoked small (相似文献   

Inhibition by bis(7)-tacrine of native delayed rectifier and KV1.2 encoded potassium channels

Bis(7)-tacrine [bis(7)-tetrahydroaminacrine] acts as an AChE inhibitor and also exerts modulatory effects on many ligand-gated ion channels and voltage-gated Ca(2+) and K(+) channels. It has been reported previously that tacrine and some other AChE inhibitors suppressed I(K(A)) in central and peripheral neurons. The present study aimed to explore whether bis(7)-tacrine could modulate the function of native delayed rectifier potassium channels in DRG neurons and K(V)1.2 encoded potassium channels expressed in oocytes. We found that both delayed rectifier potassium currents (I(K(DR))) in rat DRG neurons and the currents recorded from oocytes expressing K(V)1.2 (I(K(K(V)1.2))) were suppressed by bis(7)-tacrine, the potency of which was two orders greater than that of tacrine. The IC(50) values for bis(7)-tacrine and tacrine inhibition of I(K(KD)) in DRG neurons were 0.72+/-0.05 and 58.3+/-3.7 microM, respectively; while the two agents inhibited I(K(K(V)1.2)) in oocytes with an IC(50) of 0.24+/-0.06 and 102.1+/-21.5 microM, respectively. The possible mechanism for bis(7)-tacrine inhibition of I(K(A)) and I(K(K(V)1.2)) was identified as the suppression of their activation, inactivation.     

Effects of HC 20-511 (ketotifen) on chemiluminescence of human neutrophils

The effects of ketotifen on the chemiluminescence (CL) and chemotaxis of human neutrophils were studied in vitro. Stimulations of neutrophils by concanavalin A (Con A), calcium ionophore A23187 (CI) and formyl-methionyl-leucyl-phenylalanine (FMLP) were strongly suppressed in contrast with that by zymosan, which was only slightly inhibited in CL assay; the inhibitory effect of ketotifen was dose dependent. The addition of ketotifen even after stimulations of CI and FMLP also inhibited CL response. The neutrophils which were activated by incubation with FMLP (10^–7 M) at 37°C for 60 min emitted much greater light in CI-induced CL assay than those without the pretreatment with FMLP. Such enhanced CL of activated cells was also markedly suppressed by ketotifen. On the other hand, ketotifen did not show any inhibitory effect on the direct movement of neutrophils by FMLP (10^–7 M) at the concentrations which inhibited CL responses. These unique pharmacological activities of ketotifen are encouraging for its potential clinical usage as an antiinflammatory agent in some disorders associated with neutrophils as well as for its experimental usefulness for the analysis of various functions of neutrophils.     

Regional citrate anticoagulation for hemodialysis: calcium-free vs. calcium containing dialysate - a randomized trial

BACKGROUND: The majority of citrate protocols for hemodialysis (HD) use calcium (Ca)-free dialysate, a limited number use dialysate with Ca, aiming to simplify the procedure. This randomized clinical study sought to compare the anticoagulant effect of citrate using Ca-free dialysate and dialysate with Ca 1.25 mmol/L. METHODS: Fifty HD procedures (in 5 chronic HD patients treated by chronic citrate anticoagulation) were randomly assigned to Ca-free dialysate (25 procedures) or Ca-1.25 dialysate (25 procedures), both with Mg 0.5 mmol/L, Na 138 mmol/L, and bicarbonate 28 mmol/L. Ca-free HD: 15% Na3 citrate 80 ml/hour was infused into the arterial line, and 1 M CaCl2, 14 ml/hour into the venous line. Ca-1.25 group: 15% Na3 citrate 100 ml/hour, 1 M CaCl2 2-4 ml/hour. Polyflux H dialyzers were used. Antithrombotic effect was assessed visually after HD at 3 points: dialyzer, arterial, and venous bubble traps, using a score of 5 (no clotting) to 1 (total clotting). RESULTS: Ca-free group: arterial bubble trap score 4.7 +/- 0.5, dialyzer 4.5 +/- 0.6, venous bubble trap 4.8 +/- 0.6. Ionized calcium (iCa) at dialyzer inlet 0.34 +/- 0.17, outlet 0.21 +/- 0.06 mmol/L. All HDs were completed successfully. Ca-1.25 group: arterial bubble trap score 4.7 +/- 0.5 (NS), dialyzer 2.6 +/- 1.04 (p<0.01), venous bubble trap 2.4 +/- 0.9 (p<0.01). Volume of clot in venous bubble trap was 1.9 +/- 1.8 mL (range 0.5-6 mL). iCa at dialyzer inlet 0.24 +/- 0.05 mmol/L (p<0.05), outlet 0.63 +/- 0.11 mmol/L (p<0.01). Four of 25 HD procedures (16%) were prematurely terminated due to threatening dialyzer clotting, in 6/25 HD procedures (24%), the venous line was changed (p<0.01). CONCLUSION: Citrate anticoagulation with Ca-1.25 dialysate resulted in significantly worse anticoagulation of dialyzer and venous bubble trap compared with Ca-free dialysate, despite higher citrate dose.     

Analysis of single K(ATP) channels in mammalian dentate gyrus granule cells

ATP-sensitive potassium (K(ATP)) channels are heteromultimer complexes of subunits from members of the inwardly rectifying K(+) channel and the ATP-binding cassette protein superfamilies. K(ATP) channels couple metabolic state to membrane excitability, are distributed widely, and participate in a variety of physiological functions. Understood best in pancreatic beta cells, where their activation inhibits insulin release, K(ATP) channels have been implicated also in postischemia cardio- and neuroprotection. The dentate gyrus (DG) is a brain region with a high density of K(ATP) channels and is relatively resistant to ischemia/reperfusion-induced cell death. Therefore we were interested in describing the characteristics of single K(ATP) channels in DG granule cells. We recorded single K(ATP) channels in 59/105 cell-attached patches from DG granule cells in acutely prepared hippocampal slices. Single-channel openings had an E(K) close to 0 mV (symmetrical K(+)) and were organized in bursts with a duration of 19.3 +/- 1.6 (SE) ms and a frequency of 3.5 +/- 0.8 Hz, a unitary slope conductance of 27 pS, and a low, voltage-independent, probability of opening (P(open), 0.04 +/- 0.01). Open and closed dwell-time histograms were fitted best with one (tau(open) = 1.3 +/- 0.2 ms) and the sum of two (tau(closed,fast) = 2.6 +/- 0.9 ms, tau(closed,slow) = 302.7 +/- 67. 7 ms) exponentials, respectively, consistent with a kinetic model having at least a single open and two closed states. The P(open) was reduced ostensibly to zero by the sulfonylureas, glybenclamide (500 nM, 2/6; 10 microM,11/14 patches) and tolbutamide (20 microM, 4/6; 100 microM, 4/4 patches). The blocking dynamics for glybenclamide included transition to a subconductance state (43.3 +/- 2.6% of control I(open channel)). Unlike glybenclamide, the blockade produced by tolbutamide was reversible. In 5/5 patches, application of diazoxide (100 microM) increased significantly P(open) (0.12 +/- 0.02), which was attributable to a twofold increase in the frequency of bursts (8.3 +/- 2.0 Hz). Diazoxide was without effect on tau(open) and tau(closed,fast) but decreased significantly tau(closed,slow) (24.4 +/- 2.6 ms). We observed similar effects in 6/7 patches after exposure to hypoxia/hypoglycemia, which increased significantly P(open) (0.09 +/- 0.03) and the frequency of bursts (7.1 +/- 1.7 Hz) and decreased significantly tau(closed,slow) (29.5 +/- 1.8 ms). We have presented convergent evidence consistent with single K(ATP) channel activity in DG granule cells. The subunit composition of K(ATP) channels native to DG granule cells is not known; however, the characteristics of the channel activity we recorded are representative of Kir6.1/SUR1, SUR2B-based channels.     

ClC-3氯通道在二甲双胍抑制鼻咽癌细胞周期进程中的作用

目的:探讨ClC-3氯通道在二甲双胍抑制鼻咽癌细胞周期进程中的作用。方法:采用不同浓度二甲双胍处理低分化鼻咽癌细胞CNE-2Z,CCK-8法检测细胞活力,流式细胞术检测细胞周期分布,Western blot法检测ClC-3氯通道蛋白表达,全细胞膜片钳技术检测细胞氯电流。构建高表达ClC-3氯通道蛋白的质粒pEZ-M03-ClC-3转染CNE-2Z细胞,流式细胞术检测ClC-3氯通道对细胞周期分布的影响。结果:5、10和20 mmol/L浓度的二甲双胍均可有效抑制CNE-2Z细胞的活力。10 mmol/L二甲双胍可阻抑CNE-2Z细胞周期于G0/G1期,并抑制CNE-2Z细胞氯电流及ClC-3氯离子通道蛋白的表达。ClC-3氯通道蛋白高表达可逆转二甲双胍对CNE-2Z细胞周期分布的影响。结论:二甲双胍抑制鼻咽癌CNE-2Z细胞周期进程可能与抑制ClC-3氯通道功能和蛋白表达有关。     

The effect of azelastine and some other antiasthmatic and antiallergic drugs on calmodulin and protein kinase C

The antiallergic and antiasthmatic drug, azelastine, interacts strongly with calmodulin (but not bovine serum albumin) as determined by an indirect assay; it also moderately inhibited the Ca²⁺-calmodulindependent enzyme bovine brain phosphodiesterase. Ketotifen was less active than azelastine in both assays of calmodulin reactivity and both drugs were less active than the recognized calmodulin inhibitor, W-7. Neither azelastine nor ketotifen had any inhibitory effect on the Ca²⁺- and phospholipid-dependent protein kinase C. A number of other commonly employed antiallergic and antiasthmatic drugs were essentially inactive in the calmodulin assays and had no or marginal inhibitory effect on protein kinase C.