丙咪嗪对人血小板胞浆游离钙的影响

目的：观察丙咪嗪体外对人血小板胞浆游离钙的影响。方法：用Ｆｕｒａ２－ＡＭ荧光钙离子指示剂在体外测定人血小板胞浆游离钙离子浓度。结果：丙咪嗪明显抑制凝血酶诱导的血小板胞浆游离钙的升高，ＩＣ５０（ｉｎｈｉｂｉｔｏｒｙｃｏｎｃｅｎｔｒａｔｉｏｎｏｆ５０％）为２０．０μｍｏｌ／Ｌ。丙咪嗪对凝血酶诱导的内钙释放亦有明显抑制作用（Ｐ＜０．０１）。结论：丙咪嗪不仅明显抑制人血小板胞外Ｃａ２＋跨膜内流而且抑制胞内储存Ｃａ２＋释放     

Store-operated calcium entry modulates neuronal network activity in a model of chronic epilepsy

Store-operated Ca²⁺ entry (SOCE) over the plasma membrane is activated by depletion of intracellular Ca²⁺ stores and has only recently been shown to play a role in CNS processes like synaptic plasticity. However, the direct effect of SOCE on the excitability of neuronal networks in vitro and in vivo has never been determined. We confirmed the presence of SOCE and the expression of the calcium sensors STIM1 and STIM2, which convey information about the calcium load of the stores to channel proteins at the plasma membrane, in neurons and astrocytes. Inhibition of SOCE by pharmacological agents 2-APB and ML-9 reduced the steady-state neuronal Ca²⁺ concentration, reduced network activity, and increased synchrony of primary neuronal cultures grown on multi-electrode arrays, which prompted us to elucidate the relative expression of STIM proteins in conditions of pathologic excitability. Both proteins were increased in brains of chronic epileptic rodents and strongly expressed in hippocampal specimens from medial temporal lobe epilepsy patients. Pharmacologic inhibition of SOCE in chronic epileptic hippocampal slices suppressed interictal spikes and rhythmized epileptic burst activity. Our results indicate that SOCE modulates the activity of neuronal networks in vitro and in vivo and delineates SOCE as a potential drug target.     

Study of cell deformability by a simple method

Cell deformability plays an important role in many immunological processes, such as phagocyte chemotaxis and endocytosis. The most widely used method of assay consists in aspirating cells into glass micropipettes and measuring the length of the protrusion induced by a given pressure, or the minimum pressure required to drive cells into the micropipette. This procedure requires specialized equipment and delicate manipulation. The present report describes a simpler procedure: cells are centrifuged in petri dishes floating on a water cushion, then fixed and coated with 0.8 μm diameter latex beads, which allows rapid and accurate determination of their height. This method is compared with the micropipette technique by studying lymphocyte and macrophage-like cell lines in physiological medium and in the presence of a divalent cation chelator or a microfilament inhibitor. In addition to simplicity, the main advantages of this technique are that (i) many cells may be examined within a reasonable period of time, which allows testing of heterogenous cell populations, and (ii) unexpectedly, centrifugation was quite harmless under our experimental conditions, since it did not impair cell proliferative ability nor phagocytic ability.It is concluded that the method may be used in clinical laboratories to explore phagocyte dysfunctions, as well as in experimental studies.     

Pre-hospital tracheal intubation versus esophageal gastric tube airway use: a prospective study

A prospective study compared the respiratory effectiveness of the endotracheal tube (ET) with that of the esophageal gastric tube airway (EGTA) for victims of nontraumatic cardiac arrest in the pre-hospital setting. Arterial blood gases were obtained within 3 minutes of hospital arrival, and survival (defined as discharge from the hospital) was determined. During EGTA ventilation, mean pH was 7.12 +/- 0.2, mean P02 was 77 +/- 92 mm Hg, and mean PC02 was 78.2 +/- 42.9 mm Hg; the survival rate was 4.5%. During ET ventilation, mean pH was 7.34 +/- 0.2, mean P02 was 265 +/- 151 mm Hg, mean PC02 was 35 +/- 20.5 mm Hg; the survival rate was 7%. The authors conclude that endotracheal intubation remains the procedure of choice for airway management in the victim of cardiopulmonary arrest.     

The role of Ca2+ in the time-dependent pepsinogen secretion of frog oesophageal peptic glands stimulated by bombesin

Time- and dose-related stimulation of pepsinogen secretion by bombesin was studied in perifused dispersed peptic glands from the oesophagus of the American bullfrog Rana catesbeiana. The dose response to bombesin was monophasic between 10^-10 and 10^-7 M, with an EC₅₀ of 10^-9 M. Time-dependent secretion was closely monitored at 1–2 min intervals. Though there was overlap, we could discriminate an early response at ? 2 min (phase I) and a delayed or sustained response at ± 2 min (phase II) on the basis of responses in the presence and absence of external Ca²⁺. Phase I was relatively independent of external [Ca²⁺] and coincided with ⁴⁵Ca efflux following a dose-dependent increase in cytosolic [Ca²⁺], measured by Fura-2AM. Phase II was sustained at ? 80% of control at an external [Ca²⁺] of 1–5 ,μM, but was eliminated by adding 0.5–1 Mm EGTA. Bombesin caused a sustained Ca²⁺ influx and, when this was prevented by EGTA, the response to successive stimulations by bombesin and by acetylcholine was greatly attenuated. The phorbol ester, 12–O-tetradecanoyl phorbol 13–acetate, which stimulates secretion at high concentrations, was used as background at a thresehold concentration of 10^-7 M, which did not by itself stimulate secretion. At this concentration, 12–O-tetradecanoyl phorbol 13–acetate potentiated the responses to bombesin and to acetylcholine. These results define the different Ca²⁺ dependencies of the immediate and sustained secretory responses to bombesin, but indicate a complex relationship of stimulation responses to Ca²⁺ homeostasis in various agonist-sensitive Ca²⁺ pools.     

Simultaneous isolation of cytoplasmic endoribonuclease and exoribonuclease of Trypanosoma brucei

An endoribonuclease and an exoribonuclease have been isolated simultaneously from the cytoplasm of Trypanosoma brucei by hydroxyapatite column chromatography. The endoribonuclease produced oligonucleotides from poly(adenylic acid) with 5'-phosphate and 3'-OH termini. The exoribonuclease produced only ribonucleoside 5'-phosphates from poly(adenylic acid). The relative rates of degradation of synthetic homopolynucleotides by the endoribonuclease under standard conditions were in the order poly(adenylic acid) greater than poly(uridylic acid) poly(cytidylic acid); for the exoribonuclease the order was poly(adenylic acid) poly(uridylic acid) greater than poly(cytidylic acid). Natural transfer and ribosomal RNAs were also degraded by both enzymes, while DNA was resistant to them. The optimal pH of activity for each enzyme was 7.5-8.0. Both ribonucleases require Ca2+ for maximum enzymatic activity.     

Voltage -dependent suppression of calcium current by caffeine in single smooth muscle cells of the guinea-pig urinary bladder

The suppressive action of caffeine on l,-type Ca current (Ica) in smooth muscle cells of the guinea-pig urinary bladder was investigated using the whole-cell patch clamp technique. Caffeine (5–30 mM) suppressed Ica, the effect having two phases: a rapid and transient suppression of Ica, which was followed by a sustained suppression. When intracellular Ca⁺ was strongly buffered by the Ca⁺ chelator EGTA (20 mM) or BAPTA (5 mM) in the patch pipette, the transient suppression of Ica was abolished, whereas the sustained effect remained. Similarly, inclusion of both 10 mM procaine and 1 mg/ml heparin in the patch pipette blocked the transient suppression of Ica, but did not block the sustained effect. The degree of the sustained effect of caffeine on Ica was dose-dependent with a _d of 20 mM. Application of the cyclic AMP analogue, 8-bromo-cyclic AMP (100 M) or forskolin (10 M) to the bath failed to mimick the sustained suppression of Ica, suggesting that inhibition of phosphodiesterase activity was not involved in the caffeine action. The steady-state activation curve remained unchanged by 10 mM caffeine but the steady-state inactivation curve was significantly shifted in the negative direction by 15.6 mV in 1.8 mM Ca²⁺ solution or by 10 mV in 1.8 mM Ba²⁺ solution. From these results it appears that caffeine inhibits L-type Ica via two mechanisms: (1) it releases Ca²⁺ from an internal store causing a transient Ca²⁺-mediated inactivation of the Ca channel; (2) it inhibits Ca channel via a mechanism that does not require such a Ca²⁺ release. It is possible that caffeine suppresses Ica through a preferential binding to the inactivated state of l-type Ca channel.     

Cleavage of cytoskeletal proteins by two forms of Ca2+ activated neutral proteases in human platelets

Two forms of calcium activated neutral proteases (CANPs) with different affinity to Ca²⁺ were partially purified from the soluble fraction of human platelets; one (μ-CANP¹) required 6 μM Ca²⁺ and the other (m-CANP¹) did 900 μM Ca²⁺ for the respective half maximal activity. Human platelets were found to contain μ-CANP predominantly in contrast to our previous study on bovine platelets, in which μ- and m-CANP were equally distributed. Among platelet protein preparations examined for possible endogenous substrates of platelet CANPs, actin binding protein (ABP) and 230 K protein were proteolysed completely by μ- or m-CANP in the presence of the respective optimal concentration of Ca²⁺. As far as μ-CANP is concerned, this is the first demonstration of proteolysis of endogenous substrates, which implies possible involvement of μ-CANP in stimulus-linked platelet reaction coupled with an increase in intracellular Ca²⁺ to micromolar concentration. Furthermore, microtubules associated proteins (MAPs) of bovine brain was proteolysed by μ- or m-CANP of human platelet in the similar manner, indicating functional ubiquity of CANPs.