Stimulating capacity of blast cells from patients with chronic myelocytic leukaemia, in blastic crisis in ''one-way'' mixed lymphoycte reaction: lack of evidence for T lymphoblastic conversion.

It has long been suggested that the blastic transformation in some patients with Ph1-positive chronic myelocytic leukaemia (CML) may be lymphoid in nature. It has recently been postulated that some patients with CML may undergo a T lymphoblastic crisis because the leukaemic blasts from these patients have high terminal deoxynucleotidyl transferase (TdT) activity and that some patients may undergo a non-T/non-B lymphoblastic crisis since leukaemic blasts from a majority of morphologically lymphoid type CML-BC cases react with antiserum specific for non-T/non-B acute lymphoblastic leukaemia (ALL). The present study shows that leukaemic blasts from each of six patients with Ph1-positive chronic myelocytic leukaemia-blastic crisis (CML-BC) exerted a strong stimulation on allogeneic lymphocytes in 'one-way' mixed lymphocyte reaction. There was no apparent difference in stimulating capacity between morphologically myeloid type (four cases) and lymphoid type (two cases). The stimulating capacity of leukaemic blasts from patients with CML-BC was quite similar to that of blasts from all patients with acute myeloblastic leukaemia (AML) and from some patients with non-T/non-B type ALL. Leukaemic blasts from a patient with T-cell type ALL and cultured leukaemic T lymphoblastoid cells (2 lines) consistently failed to stimulate while cultured leukaemic null-cells (4 lines) consistently exerted a strong stimulation in 'one-way' mixed lymphocyte reaction. These observations suggest that leukaemic cells from patients with CML-BC, morphologically lymphoblastic type, are not T lymphoblasts although the possibility that these cells are non-T/non-B lymphoblasts cannot be ruled out entirely.     

Leukaemic T Cells from Patients with Chronic Lymphocytic Leukaemia of T-Cell Origin Respond to Staphylococcus aureus Enterotoxin Superantigens

We investigated the in vitro responsiveness of peripheral blood lymphocytes from two patients with T-cell chronic lymphocytic leukaemia (T-CLL) to Staphylococcus aureus enterotoxin (SE) superantigens. T-cell receptor (TcR) αβ(Vβ 7.1)-expressing CD4⁺ leukaemic T cells from patient HE (white blood cell count 480,000/μl) proliferated in response to SEA and, only at 1000-fold higher concentrations, to SEB, SED, and SEE. CD4⁺ CD8⁺ TcRαβ (Vβ 12.1)-expressing leukaemic T cells from patient KO (white blood cell count 120,000/μl) were activated by SEB but not by the other tested SEs. In both instances, the activation of leukaemic T cells by SE was dependent on the presence of HLA-DR⁺ cells. Southern blot analysis of TcRβ gene rearrangement confirmed that the proliferating cells were derived from the leukaemic T-cell clone and not from contaminating normal T cells. These data indicate that leukaemic T cells from patients with T-CLL exert a clonally variable responsiveness to SE superantigens. We conclude that recognition of specific antigen and subsequent signal transduction can be initiated via the TcR of leukaemic T-CLL cells.     

Mixed cultures of human foetal and adult cells

Normal and leukaemic lymphocytes from healthy donors and patients with chronic lymphatic leukaemia exhibit increased DNA synthesis in `one way' cultures with foetal cells. Leukaemic blast cells do not react. Foetal thymocytes showed a marked response to both normal and leukaemic cells in mixed cultures. Leukaemic lymphocytes in particular caused a marked increase in [³H]thymidine uptake, suggesting a wider antigenic disparity between the leukaemic and thymic cells. Lesser increases of DNA synthesis were produced in foetal liver cells and none in spleen cells.     

The Effect of Calcium Mobilization on LPS-Induced IL-1β Production Depends on the Differentiation Stage of the Monocytes/Macrophages

The role of elevated intracellular Calcium concentration [Ca²⁺]_i in the LPS-induced activation of interleukin-1β (IL-1β) production was examined in cells representing different stages of myeloid differentiation (undifferentiated monocytic leukaemia cell line THP-1, THP-1 cells induced to adherent, macrophage-like cells by phorbol ester treatment and normal peripheral blood-derived adherent monocytes). LPS did not elevate the [Ca²⁺]¹, as measured by the Fura-2 fluorescence technique. When these cells were stimulated with LPS in the presence of the calcium ionophore A23187, a clear increase in the IL- 1β protein production was observed in the undifferentiated THP-1 cells but not in the more differentiated cell types. This ionophore-induced increase was also seen in the IL-1β mRNA levels. Thus these data confirm the previous findings demonstrating that elevation of (Ca-²⁺]₁ is not involved in the LPS-dependent signal transmission. However, the LPS-induced signals are greatly potentiated by the elevated [Ca²⁺]_i, but only in undifferentiated monocytic cells.     

The Effects of Mitogens, IL-2 and Anti-CD3 Antibody on the T-Cell Receptor Vβ Repertoire

Phytohaemagglutinin (PHA), Concanavalin A (Con A), interleukin-2 (IL-2), and monoclonal antibodies to CD3 (CD3MoAbs) are used for the assessment of the T-cell receptor (TCR) BV gene family expression in autoimmune disorders and multiple sclerosis, and to produce clones for assessment of cytokine profiles in progressive human immunodeficiency virus infection. The authors examined the effects of these stimulants on the TCR Vβ repertoire of resting and blastic CD4⁺ and CD8⁺ normal human peripheral blood lymphocytes, using three-colour cytofluorometry and a panel of anti-TCR Vβ monoclonal antibodies. IL-2 was associated with an increased percentage of blastic CD4⁺ cells expressing V_β5.1 (from median of 3.7% to 8.0%, P  = 0.0002) and blastic CD8⁺ cells expressing V_β5.3 (1.0 to 1.5%, P  = 0.0039). CD3MoAb caused a slight increase in V_β6.7 + blastic CD4⁺ cells (4.5 to 6.9%, P  = 0.0078). PHA did not alter the Vβ repertoire of blastic cells. Con A caused skewing in CD8⁺ blastic cells, toward expression of V_β5.2/5.3 (3.1 to 8.1%) and V_β5.3 (0.8 to 4.8%) ( P  = 0.0020). Thus, IL-2 stimulation causes slight alterations in the Vβ repertoire that should be taken into account in certain research settings. Con A produced skewing in CD8⁺ blastic cells suggesting that, in the presence of CD8, either Con A binds selectively to certain Vβ or the three-dimensional complex created by Con A's binding to other T-cell surface molecules induces preferential V_β5 stimulation.     

Dissociation of the store-operated calcium current ICRAC and the Mg-nucleotide-regulated metal ion current MagNuM

Rat basophilic leukaemia cells (RBL-2H3-M1) were used to study the characteristics of the store-operated Ca²⁺ release-activated Ca²⁺ current ( I _CRAC) and the magnesium-nucleotide-regulated metal cation current (MagNuM) (which is conducted by the LTRPC7 channel). Pipette solutions containing 10 m m BAPTA and no added ATP induced both currents in the same cell, but the time to half-maximal activation for MagNuM was about two to three times slower than that of I _CRAC. Differential suppression of I _CRAC was achieved by buffering free [Ca²⁺]_i to 90 n m and selective inhibition of MagNuM was accomplished by intracellular solutions containing 6 m m Mg.ATP, 1.2 m m free [Mg²⁺]_i or 100 μ m GTP-γ-S, allowing investigations on these currents in relative isolation. Removal of extracellular Ca²⁺ and Mg²⁺ caused both currents to be carried significantly by monovalent ions. In the absence or presence of free [Mg²⁺]_i, I _CRAC carried by monovalent ions inactivated more rapidly and more completely than MagNuM carried by monovalent ions. Since several studies have used divalent-free solutions on either side of the membrane to study selectivity and single-channel behaviour of I _CRAC, these experimental conditions would have favoured the contribution of MagNuM to monovalent conductance and call for caution in interpreting results where both I _CRAC and MagNuM are activated.     

Inhibition of human lymphocyte stimulation by steroid hormones: cytokinetic mechanisms.

The steroid hormones estradiol, progesterone and testosterone, in addition to cortisol, inhibited stimulation of human peripheral blood lymphocytes by phytohaemagglutinin (PHA) and Con A. This effect upon lymphocyte transformation was assayed by three methods: quantitation of [3H]thymidine incorporation into acid precipitable material, microscopic assessment of blastic transformation and determination of the labelling index. Addition of steroid hormones at the initiation of culture resulted in a marked inhibition in all three parameters, which was observed with lower concentrations of cortisol than the other hormones. The inhibition was not attributable to cell death and could be partially reversed by removing hormones from the incubation medium after culture for 48-72 hr. Late addition of steroid hormones, 52 hr after addition of mitogen and 18 hr prior to pulse-labelling with [3H]thymidine, also resulted in reduced [3H]thymidine incorporation, accompanied by a nearly 50% reduction in the labelling indices and only a minimal decrease in the per cent transformed cells. Inhibition of lympohcyte stimulation by steroid hormones operates by the following cytokinetic mechanisms: (1) suppressed recruitment of cells from G3 to G1 phase of the cell cycle, as indicated by the diminished per cent blasts; (2) inhibition of progression from G1 phase into S phase, as evidenced by the reduced ratio [labelling index/blasts]; and, in the case of estradiol and progesterone, (3) reduced rate of DNA replication or altered intracellular [3H]thymidine specific activity as shown by the decreased ([3H]thymidine incorporation/labelling index) ratio. Late addition of steroid hormones to stimulated cultures reduced the per cent of cells in S phase, but did not revert previously transformed cycling lymphocytes to the G3 state.     

Capacitative calcium entry supports calcium oscillations in human embryonic kidney cells

Treatment of human epithelial kidney (HEK293) cells with low concentrations of the muscarinic agonist methacholine results in the activation of complex and repetitive cycling of intracellular calcium ([Ca²⁺]_i), known as [Ca²⁺]_i oscillations. These oscillations occur with a frequency that depends on the concentration of methacholine, whereas the magnitude of the [Ca²⁺]_i spikes does not. The oscillations do not persist in the absence of extracellular Ca²⁺, leading to the conclusion that entry of Ca²⁺ across the plasma membrane plays a significant role in either their initiation or maintenance. However, treatment of cells with high concentrations of GdCl₃, a condition which limits the flux of calcium ions across the plasma membrane in both directions, allows sustained [Ca²⁺]_i oscillations to occur. This suggests that the mechanisms that both initiate and regenerate [Ca²⁺]_i oscillations are intrinsic to the intracellular milieu and do not require entry of extracellular Ca²⁺. This would additionally suggest that, under normal conditions, the role of calcium entry is to sustain [Ca²⁺]_i oscillations. By utilizing relatively specific pharmacological manoeuvres we provide evidence that the Ca²⁺ entry that supports Ca²⁺ oscillations occurs through the store-operated or capacitative calcium entry pathway. However, by artificial introduction of a non-store-operated pathway into the cells (TRPC3 channels), we find that other Ca²⁺ entry mechanisms can influence oscillation frequency in addition to the store-operated channels.     

Inhibition of normal lymphocyte transformation by plasma and lymphocyte factors in chronic lymphatic leukaemia

Phytohaemagglutinin transformation of normal lymphocytes was significantly inhibited by the plasma and by extracts of lymphocytes in six out of eight cases of untreated chronic lymphatic leukaemia compared with similar material from matched control patients. Since the concentration of inhibitor in lymphocytes was much greater than that in plasma and both inhibitors were thermolabile, it is possible that the plasma inhibitor may be produced by the leukaemic cells. These findings indicate modifications of normal T-cell function by constituents of neoplastic lymphocytes but the effect may be due to exaggeration of a normal mechanism in which the surface properties of T cells are modified by products of B cells.     

T-Lymphocyte Functions in Acute Leukaemia Patients with Severe Chemotherapy-Induced Cytopenia: Characterization of Clonogenic T-Cell Proliferation

Intensive chemotherapy for acute leukaemia is followed by a period of severe chemotherapy-induced leukopenia. We used a limiting dilution assay to investigate whether remaining CD4⁺ and CD8⁺ T lymphocytes derived from such leukopenic patients could be activated and undergo clonogenic proliferation. The activation signal in our model was accessory cells (irradiated normal peripheral blood mononuclear cells) + phytohaemagglutinin (PHA) + interleukin-2 (IL-2). During severe leukopenia a majority of circulating lymphocytes were CD4⁺ T cells. Clonogenic proliferating T lymphocytes were detected for all patients. Higher frequencies of clonogenic cells were detected in the CD8⁺ subset as compared to the CD4⁺ subset. However, for both subsets frequencies of proliferating cells were decreased compared with healthy individuals. The CD4⁺ and CD8⁺ lymphocytes were also capable of proliferation in response to alloactivation, and accessory cells mainly containing acute myelogenous leukaemia blast were efficient as accessory cells for activation. For the CD4⁺ cells, increased proliferation was detected in the presence of acute myelogenous leukaemia (AML) blasts compared with normal accessory cells. Based on our results we conclude that: (1) although acute leukaemia patients with therapy-induced leukopenia have both a quantitative and a qualitative T-cell defect, (2) the remaining T-cell population includes a subset capable of clonogenic proliferation. However, (3) proliferation of the clonogenic CD4⁺ cells can be modulated by AML blasts.     

CD66c antigen expression is myeloid restricted in normal bone marrow but is a common feature of CD10+ early-B-cell malignancies

Abstract: CD66c is a surface (and intracellular) molecule bound to the membrane by a glycosyl-phosphatidylinositol anchor. While its expression on peripheral granulocytes is well recognized, less is known about its distribution in early steps of normal and neoplastic hematopoiesis. We analyzed by flow cytometry cell surface expression of CD66c on bone marrow cells from 4 healthy subjects and on bone marrow or peripheral blood cells from 127 patients with newly diagnosed hematologic malignancies: 70 de novo acute myeloid leukemias (AML), 6 refractory anemias with excess of blasts in transformation, 3 myeloid and 3 lymphoid blastic phases of chronic mye-logenous leukemia, 33 B-lineage and 6 T-lineage acute lymphoblastic leukemias (B- and T-ALL), and 3 B-cell and 3 T-cell non-Hodgkin's lymphomas in the leukemic phase. We found that in normal bone marrow CD66c expression was myeloid restricted, reaching its highest level on promyelocytes. As for de novo AML, slight expression of CD66c was found on 6/25 (24%) AML-M4 and only occasionally in other subgroups. In 9 out of 10 cases of acute promyelocytic leukemia, CD66c was totally absent, but antigen expression was easily detectable following in vitro exposure to all- trans retinoic add. Among lymphoid malignancies, CD10⁺ early-B-ALL consistently expressed the molecule (20/23 cases, or 87%) whereas both CD10^- early-B ALL and Smlg⁺ B-ALL completely lacked it. Finally, dual staining with CD66c and CD10 proved to be a suitable tool for distinguishing even low percentages of residual leukemic cells (CD10⁺/CD66c⁺) from normal regenerating early-B cells (CD10⁺/CD66c^-) in CD10⁺ early-B-ALL induced into remission.     

Leukaemia antigens. Stimulation of lymphocytes in mixed culture by cells from HL-A identical siblings

The frequency with which peripheral blood leucocytes (PBL) from ninety-one patients with acute leukaemia would stimulate non-reciprocally lymphocytes from their normal genotypically HL-A identical siblings was compared with that observed between cells of eighty-two pairs of normal HL-A identical siblings. The mean stimulation index (SI) between cells of normal sibling pairs was 1·06±0·55, and significant stimulation, i.e. SI[unk]2·16 was observed in only 7/164 reactions. Similarly, PBL from only 2/28 patients with leukaemia in complete remission stimulated cells from 2/44 siblings. By contrast, PBL from 17/63 patients with leukaemia in relapse stimulated cells from 21/91 of their HL-A identical siblings. The results in marrow relapse were not significantly influenced by type of leukaemia nor by presence or absence of leukaemic cells in the PBL. The results suggest that PBL from some patients with acute leukaemia in relapse but not in remission possess antigens recognizable by lymphocytes from normal HL-A identical siblings, and that such antigens may be present on morphologically normal as well as leukaemic cells.     

Action potential fidelity during normal and epileptiform activity in paired soma–axon recordings from rat hippocampus

Although action potential initiation and propagation are fundamental to nervous system function, there are few direct electrophysiological observations of propagating action potentials in small unmyelinated fibres, such as the axons within mammalian hippocampus. To circumvent limitations of previous studies that relied on extracellular stimulation, we performed dual recordings: whole-cell recordings from hippocampal CA3 pyramidal cell somas and extracellular recordings from their axons, up to 800 μm away. During brief spike trains under normal conditions, axonal spikes were more resistant to amplitude reduction than somatic spikes. Axonal amplitude depression was greatest at the axon initial segment < 150 μm from the soma, and initiation occurred ∼75 μm from the soma. Although prior studies, which failed to verify spike initiation, suggested substantial axonal depression during seizure-associated extracellular K⁺ ([K⁺]_o) rises, we found that 8 m m [K⁺]_o caused relatively small decreases in axonal spike amplitude during brief spike trains. However, during sustained, epileptiform spiking induced in 8 m m [K⁺]_o, axonal waveforms decreased significantly in peak amplitude. During epileptiform spiking, bursts of two or more action potentials > 20 Hz failed to propagate in most cases. In normal [K⁺]_o at 25 and 32°C, spiking superimposed on sustained somatic depolarization, but not spiking alone, produced similar axonal changes as the epileptiform activity. These results highlight the likely importance of steady-state inactivation of axonal channels in maintaining action potential fidelity. Such changes in axonal propagation properties could encode information and/or serve as an endogenous brake on seizure propagation.     

Investigation and Standardization of the Conditions for Micro-Lymphocyte Cultures

Experiments were performed to establish optimal conditions for specific and non-specific lymphocyte transformation in vitro , in cultures containing 1–3 × 10⁵ human lymphocytes. The stimulation as measured by ¹⁴C-thymidine incorporation was evaluated when using various methods of lymphocyte purification and different numbers of contaminating erythrocytes, different kinds of tubes, serum from different sources, and varying PHA concentrations.
Furthermore, the dependence of the ¹⁴C-thymidine incorporation on the thymidine concentration in the medium and on the total number of lymphocytes present per culture was determined.
Two methods of harvesting the cells to measure the ¹⁴C-thymidine incorporation were compared.
On the basis of this analysis, conditions for maximal sensitivity and reproducibility could be defined in measuring lymphocyte reactivity to PHA, allogeneic lymphocytes and antigens in a miniaturized culture system.     

Cell-Mediated Cytotoxicity for Melanoma Tumor Cells: Detection by a (3H) Proline Release Assay

An in vitro lymphocyte-mediated cytotoxicity assay using [³H]proline-labeled target cells is described. The assay, modified from an original procedure of Bean et al., assesses the release of [³H]proline by filtering the total culture fluid containing both trypsinized tumor cells and effector cells. Filtration is performed with a semiautomatic harvesting device using low suction pressure and large-diameter glass filters. Pretreatment of filters with whole serum diminishes adsorption of cell-free radioactive material considerably and thus increases the sensitivity of the assay. Nearly 100% of the radioactivity could be recovered with this harvesting device. The technique allowed the detection of cytolytic activities of lymphocytes after 6 h of incubation. Lymphocytes from patients with primary malignant melanoma showed a significantly higher cytolytic reactivity (P < 0.001) than normal donors' lymphocytes against three different melanoma cell lines. In a series of parallel experiments on 36 patients and IS normal donors, this modification of the [³H]proline test was compared with three different assays: the conventional microcytotoxicity test of Takasugi and Klein, the original [³H]proline microcytotoxicity test of Bean et al., and the viability count of tumor cells.     

Separation of T lymphocytes from normal individuals and patients with B lymphocyte chronic lymphocytic leukaemia.

Previous studies have shown that lymphocytes from patients with chronic lymphocytic leukaemia have a diminished response to mitogens which stimulate T cells. Chronic lymphocytic leukaemia is most often a disease of accumulating B cells so that T lymphocytes are diluted by large numbers of leukaemic cells. Direct comparison with the responses of normal lymphocytes to mitogenic stimulation is therefore suspect. To circumvent this difficulty, a method of isolating T cells from normal individuals and patients with chronic lymphocytic leukaemia was developed. Lymphocytes containing an average of 16.1 per cent B cells from normal individuals were applied to IgG-anti-IgG-coated Degalan bead columns and held at 4 degrees for 2 hours. The eluted cells contained less than 2 per cent B cells. When chronic lymphocytic leukaemic lymphocytes, containing an average of 68.6 per cent B cells, were applied to IgG-anti-IgG columns, the eluted cells contained 36.4 per cent B cells. To improve the purification of T lymphocytes, columns of uncoated Degalan beads were used to remove non-specifically adherent cells. Eluted lymphocytes were then applied to IgG-anti-IgG columns. This resulted in the recovery of purified populations of T cells with less than 2 per cent contamination with B cells. Patients with chronic lymphocytic leukaemia were found to have lymphocytes with either a normal density or a low density of surface immunoglobulins. B cells were successfully removed from lymphocyte suspensions in all cases of chronic lymphocytic leukaemia with a normal density of lymphocyte surface immunoglobulins. In the three cases of chronic lymphocytic leukaemia with low density surface immunoglobulins, separation by this method was unsuccessful. However, an enriched T-cell population was obtained when leukaemic lymphocytes which had lost all detectable surface immunoglobulins were passed through a column coated with heat-aggregated IgG.     

Ca2+-NFATc1 signaling is an essential axis of osteoclast differentiation

Summary:  Osteoclasts are unique, multinucleated giant cells that decalcify and degrade the bone matrix. They originate from hematopoietic cells and their differentiation is dependent on a tumor necrosis factor (TNF) family cytokine, receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL), as well as macrophage-colony stimulating factor (M-CSF). Recent studies have unveiled the precise molecular mechanism underlying osteoclastogenesis. In particular, the discovery of nuclear factor of activated T cells c1 (NFATc1), the master regulator of osteoclastogenesis, has proven to be a breakthrough in this field. NFATc1 is activated by Ca²⁺ signaling induced by the activation of the immunoglobulin-like receptor signaling associated with immunoreceptor tyrosine-based activation motif (ITAM)-harboring adapters. The long-lasting Ca²⁺ oscillation, which is evident during osteoclastogenesis, may ensure the robust induction of NFATc1 through an autoamplification mechanism. Thus, intracellular Ca²⁺ is a critical attribute of osteoclastogenic signaling. In addition, osteoclasts are exposed to a very high extracellular Ca²⁺ concentration ([Ca²⁺]_o) in the bone microenvironment and respond to the change in [Ca²⁺]_o by increasing the intracellular Ca²⁺, which regulates diverse cellular functions. Investigation of the molecular mechanisms underlying the regulation of intracellular Ca²⁺ dynamics may open up new directions for therapeutic strategies in bone disease.     

Discrete store-operated calcium influx into an intracellular compartment in rabbit arteriolar smooth muscle

This study tested the hypothesis that store-operated channels (SOCs) exist as a discrete population of Ca²⁺ channels activated by depletion of intracellular Ca²⁺ stores in cerebral arteriolar smooth muscle cells and explored their direct contractile function. Using the Ca²⁺ indicator fura-PE3 it was observed that depletion of sarcoplasmic reticulum (SR) Ca²⁺ by inhibition of SR Ca²⁺-ATPase (SERCA) led to sustained elevation of [Ca²⁺]_i that depended on extracellular Ca²⁺ and slightly enhanced Mn²⁺ entry. Enhanced background Ca²⁺ influx did not explain the raised [Ca²⁺]_i in response to SERCA inhibitors because it had marked gadolinium (Gd³⁺) sensitivity, which background pathways did not. Effects were not secondary to changes in membrane potential. Thus SR Ca²⁺ depletion activated SOCs. Strikingly, SOC-mediated Ca²⁺ influx did not evoke constriction of the arterioles, which were in a resting state. This was despite the fura-PE3-indicated [Ca²⁺]_i rise being greater than that evoked by 20 m m [K⁺]_o (which did cause constriction). Release of endothelial vasodilators did not explain the absence of SOC-mediated constriction, nor did a change in Ca²⁺ sensitivity of the contractile proteins. We suggest SOCs are a discrete subset of Ca²⁺ channels allowing Ca²⁺ influx into a 'non-contractile' compartment in cerebral arteriolar smooth muscle cells.     

Intracellular calcium signalling in magnocellular neurones of the rat supraoptic nucleus: understanding the autoregulatory mechanisms

Oxytocin and vasopressin, released at the soma and dendrites of neurones, bind to specific autoreceptors and induce an increase in [Ca²⁺]₁. In oxytocin cells, the increase results from a mobilisation of Ca²⁺ from intracellular stores, whereas in vasopressin cells, it results mainly from an influx of Ca²⁺ through voltage-dependent channels. The response to vasopressin is coupled to phospholipase C and adenylyl-cyclase pathways which are activated by V₁ (V_1a and V_1b)- and V₂-type receptors respectively. Measurements of [Ca²⁺]₁ in response to V_1a and V₂ agonists and antagonists suggest the functional expression of these two types of receptors in vasopressin neurones. The intracellular mechanisms involved are similar to those observed for the action of the pituitary adenylyl-cyclase-activating peptide (PACAP). Isolated vasopressin neurones exhibit spontaneous [Ca²⁺]₁ oscillations and these are synchronised with phasic bursts of electrical activity. Vasopressin modulates these spontaneous [Ca²⁺]₁ oscillations in a manner that depends on the initial state of the neurone, and such varied effects of vasopressin may be related to those observed on the electrical activity of vasopressin neurones in vivo.     

Lysine metabolism in acute leukemia]

The metabolism of 14C-lysine by leukaemic cells in acute myeloblastic, myelomonocytic, lymphoblastic and chronic myeloid leukaemia with blast crisis was studied. The investigations included lysine metabolism to CO2, lipids, organic acids and nucleotides and its incorporation into cellular proteins. The obtained results were compared with determinations carried out in granulocytes and lymphocytes of healthy subjects. Cells in acute leukaemias metabolized 14C-lysine in a similar range. In relation to normal cells the range of lysine metabolism to lipids in the leukaemic cells was significantly higher (p less than 0.01), while that of organic acids was significantly lower (p less than 0.05). The activity of 14C-lysine metabolism depended on the number of blast cells in the sample and the type of acute leukaemia. Neoplastic cells in blast crisis and in acute myeloblastic leukaemia incorporated more actively 14C-lysine into proteins than cells in acute myelomonocytic and acute lymphoblastic leukaemia (p less than 0.05). Similar differences in lysine metabolism were observed between myelomonocytes and blast cells from acute lymphoblastic leukaemia (p less than 0.05).